Effect of increasing maximal aerobic exercise on serum muscles enzymes in professional field hockey players.

PubMed

Hazar, Muhsin; Otag, Aynur; Otag, Ilhan; Sezen, Mehmet; Sever, Ozan

2014-11-04

Exercise results in oxidative enzyme increase and micro-injuries in skeletal muscles. The aim of this study was to investigate the effect of maximal aerobic exercise on serum muscle enzymes in professional field hockey players. This study aims to determine the effect of increasing maximal aerobic exercise on creatine kinase (CK), creatine kinase-MB (CK-MB), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) serum levels. 31 young professional field hockey players (13 female and 18 male players) volunteered for this study. All participants underwent the shuttle run test. Blood samples were taken from each participant before the shuttle run test. Post test blood samples were taken immediately after exercise and one hour after respectively. Pre and post test CK, CK-MB, AST and ALT values were measured by means of auto analyzer using original kits. The acute post test measure of the CK level increased in male (p=0.002) and female (p=0.00) sportsmen. CK-MB values obtained one hour after the exercise was lower than those before the exercise in males (p=0.02). In females (p=0.017) and males (p=0.05) AST activity significantly increased immediately after exercise and decreased to resting activity 1 h recovery. ALT significantly increased immediately after exercise in female (p=0.03) and male (p=0.00) athletes and after 1 h recovery ALT activities decreased below resting values. The timing and severity of exercise used in our study increased CK values, decreased CK-MB values and AST, ALT values increased in female and male field hockey players.

Effect of diode-laser and AC magnetic field of bovine serum albumin nanospheres loaded with phthalocyanine and magnetic particles.

PubMed

Simioni, Andreza Ribeiro; Rodrigues, Marcilene M A; Primo, Fernando L; Morais, Paulo C; Tedesco, Antonio Claudio

2011-04-01

This study reports on the development and characterization of bovine serum albumin (BSA) nanospheres containing Silicon(IV) phthalocyanine (NzPc) and/or maghemite nanoparticles (MNP), the latter introduced via ionic magnetic fluid (MF). The nanosized BSA-loaded samples were designed for synergic application while combining Photodynamic Therapy and Hyperthermia. Incorporation of MNP in the albumin-based template, allowing full control of the magnetic content, was accomplished by adding a highly-stable ionic magnetic fluid sample to the albumin suspension, following heat denaturing. The material's evaluation was performed using Zeta potential measurements and scanning electron microscopy. The samples were characterized by steady-state techniques and time-resolved fluorescence. The in vitro assay, using human fibroblasts, revealed no cytotoxic effect in all samples investigated, demonstrating the potential of the tested system as a synergistic drug delivery system.

Evaluation of the rapid plasma reagin "teardrop" card test for screening of syphilis in field conditions.

PubMed

Van Dyck, E; Van de Velden, L; Ndoye, I; Piot, P; Meheus, A

1993-01-01

The availability of simple diagnostic methods may contribute to more efficient control of sexually transmitted diseases (STDs) in developing countries. For the detection of syphilis, a simple rapid plasma reagin (RPR) "teardrop" assay for finger-prick blood samples was developed in 1962. The reliability of this test is compared with RPR, Treponema pallidum hemagglutination assay (TPHA), and fluorescent treponemal antibody absorption (FTA-Abs) assays performed on venous blood samples. To evaluate the potential usefulness of the finger-stick RPR teardrop assay for diagnosis of syphilis in settings with poor medical resources. Pregnant women evaluated at two health centers in Pikine, Senegal were tested for STDs. The RPR teardrop assay was performed on plasma from blood samples obtained by finger prick, and standard RPR, TPHA, and FTA-Abs procedures were performed on serum obtained by vein puncture. The sensitivity and specificity of the finger-prick RPR teardrop assay were 69.7% and 96.5%, respectively, and its reactivity was correlated with RPR serum antibody titer. The finger-prick RPR teardrop assay is not a reliable alternative to the classic serum RPR test.

Monoclonal antibodies specific to sailfish serum albumin: development of an assay for the identification of fish species in the field.

PubMed

Rossi, E A; Shepard, S R; Poyer, J C; Hartmann, J X

1992-06-01

Balb/c mice were immunized with albumin purified from sailfish (Istiophorus albicans) serum. Hybridomas were produced and screened by ELISA for reactivity with the purified albumins of sailfish, blue marlin (Makaira nigricans) and white marlin (Tetrapturus albidus). Monoclonal antibodies (MAbs) from 16 different clones exhibited activity against sailfish albumin. Thirteen of the MAbs showed cross-reactivity with the marlin species. Three MAbs exhibited distinct specificity for sailfish albumin. One of these species specific MAbs (M2D1) was conjugated to horseradish peroxidase (HRP) in order to construct an ELISA for identification of sailfish from serum. The ELISA for sailfish correctly identified eight sailfish from 26 billfish serum samples. The MAb-peroxidase conjugate was highly specific toward sailfish in that no reaction against heterologous species was detected.

Development of a qualitative indirect ELISA for the measurement of rabies virus-specific antibodies from vaccinated dogs and cats.

PubMed

Cliquet, F; McElhinney, L M; Servat, A; Boucher, J M; Lowings, J P; Goddard, T; Mansfield, K L; Fooks, A R

2004-04-01

A protocol suitable for the detection of rabies virus-specific antibodies in serum samples from companion animals using an enzyme linked immunosorbent assay (ELISA) is described. This method has been used successfully for the qualitative assessment of rabies virus-specific antibodies in serum samples from a cohort of vaccinated dogs and cats. In two initial field studies, a variable population of field samples from the Veterinary Laboratories Agency (VLA), United Kingdom were tested. In the first study (n = 1000), the number of false-positive and false-negative results was 11 samples (1.1%) and 67 samples (6.7%), respectively. In the second study (n = 920), the number of false-positive and false-negative results was 7 samples (0.8%) and 52 samples (5.7%). In a third study, undertaken at l'Agence Française de Sécurité Sanitaire des Aliments (AFSSA), Nancy, France (n = 440), 1 false-positive sample (0.23%) and 91 (20.7%) false-negative samples were identified. Data generated using this prototype ELISA indicate a strong correlation for specificity when compared to the gold standard fluorescent antibody virus neutralisation (FAVN) test. Although the ELISA has a lower sensitivity than the FAVN test, it is a useful tool for rapidly screening serum samples from vaccinated companion animals. Using a cut-off value of 0.6 EU/ml, the sensitivity (R = % from VLA and 79% from AFSSA) and specificity (R = 97.3%) indices between the ELISA compared favourably with data generated using the FAVN test. The major advantages of the ELISA test are that it is a qualitative tool that can be completed in four hours, does not require the use of live virus and can be performed without the need for specialised laboratory containment. This contrasts with 4 days using conventional rabies antibody virus neutralisation assays. Using the current format, the ELISA assay described would be a valuable screening tool for the detection of rabies antibodies from vaccinated domestic animals in combination with other Office International des Epizooties (OIE) accepted serological tests.

Marine Mammal Necropsy: An Introductory Guide for Stranding Responders and Field Biologists

DTIC Science & Technology

2007-09-01

the researcher or lab for required tissues and proper sample storage protocols (chill, fix, freeze and/or place in viral transport media). The most...tissues and fluids such as: liver, kidney, serum, aqueous humor, stom- ach contents, intestinal contents, feces, and urine . Tissue samples can be stored...refer to the Figure (2-1) for further explanation on frozen sample storage . The first label is written in black Sharpie on a 1 - 2 square inch piece of

Characterization of immobilization methods of antiviral antibodies in serum for electrochemical biosensors

NASA Astrophysics Data System (ADS)

Huy, Tran Quang; Hanh, Nguyen Thi Hong; Van Chung, Pham; Anh, Dang Duc; Nga, Phan Thi; Tuan, Mai Anh

2011-06-01

In this paper, we describes different methods to immobilize Japanese encephalitis virus (JEV) antibodies in human serum onto the interdigitated surface of a microelectrode sensor for optimizing electrochemical detection: (1) direct covalent binding to the silanized surface, (2) binding to the silanized surface via a cross-linker of glutaraldehyde (GA), (3) binding to glutaraldehyde/silanized surface via goat anti-human IgG polyclonal antibody and (4) binding to glutaraldehyde/silanized surface via protein A (PrA). Field emission scanning electron microscopy, Fourier transform infrared spectrometry, and fluorescence microscopy are used to verify the characteristics of antibodies on the interdigitated surface after the serum antibodies immobilization. The analyzed results indicate that the use of protein A is an effective choice for immobilization and orientation of antibodies in serum for electrochemical biosensors. This study provides an advantageous immobilization method of serum containing antiviral antibodies to develop electrochemical biosensors for preliminary screening of viruses in clinical samples from outbreaks.

Metabolomics-Based Discovery of Small Molecule Biomarkers in Serum Associated with Dengue Virus Infections and Disease Outcomes

PubMed Central

Voge, Natalia V.; Perera, Rushika; Mahapatra, Sebabrata; Gresh, Lionel; Balmaseda, Angel; Loroño-Pino, María A.; Hopf-Jannasch, Amber S.; Belisle, John T.; Harris, Eva; Blair, Carol D.; Beaty, Barry J.

2016-01-01

Background Epidemic dengue fever (DF) and dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) are overwhelming public health capacity for diagnosis and clinical care of dengue patients throughout the tropical and subtropical world. The ability to predict severe dengue disease outcomes (DHF/DSS) using acute phase clinical specimens would be of enormous value to physicians and health care workers for appropriate triaging of patients for clinical management. Advances in the field of metabolomics and analytic software provide new opportunities to identify host small molecule biomarkers (SMBs) in acute phase clinical specimens that differentiate dengue disease outcomes. Methodology/Principal Findings Exploratory metabolomic studies were conducted to characterize the serum metabolome of patients who experienced different dengue disease outcomes. Serum samples from dengue patients from Nicaragua and Mexico were retrospectively obtained, and hydrophilic interaction liquid chromatography (HILIC)-mass spectrometry (MS) identified small molecule metabolites that were associated with and statistically differentiated DHF/DSS, DF, and non-dengue (ND) diagnosis groups. In the Nicaraguan samples, 191 metabolites differentiated DF from ND outcomes and 83 differentiated DHF/DSS and DF outcomes. In the Mexican samples, 306 metabolites differentiated DF from ND and 37 differentiated DHF/DSS and DF outcomes. The structural identities of 13 metabolites were confirmed using tandem mass spectrometry (MS/MS). Metabolomic analysis of serum samples from patients diagnosed as DF who progressed to DHF/DSS identified 65 metabolites that predicted dengue disease outcomes. Differential perturbation of the serum metabolome was demonstrated following infection with different DENV serotypes and following primary and secondary DENV infections. Conclusions/Significance These results provide proof-of-concept that a metabolomics approach can be used to identify metabolites or SMBs in serum specimens that are associated with distinct DENV infections and disease outcomes. The differentiating metabolites also provide insights into metabolic pathways and pathogenic and immunologic mechanisms associated with dengue disease severity. PMID:26913918

Evaluation of the presence of porcine reproductive and respiratory syndrome virus in pig meat and experimental transmission following oral exposure

PubMed Central

2004-01-01

Abstract A study was performed to evaluate the presence of porcine reproductive and respiratory syndrome virus (PRRSV) in pig meat collected at slaughterhouses and its potential transmission to pigs via pig meat. A total of 1039 blood samples were collected from pigs upon their arrival at the abattoir. The following day, meat samples (n = 1027) were collected from the carcasses of these same pigs. Samples originated from 2 Canadian slaughterhouses, 1 situated in the province of Quebec and the other situated in the province of Manitoba. Serum samples were tested for antibodies to PRRSV and both serum and meat samples were also tested for PRRSV nucleic acid by polymerase chain reaction (PCR). Seropositivity to PRRSV for all serum samples was 74.3%. Furthermore 45 (4.3%) of the total serum samples and 19 (1.9%) of the 1027 meat samples were positive for PRRSV by PCR. Sequence analysis of open reading frame (ORF) 5 performed on 15 of the 19 PRRSV strains identified in pig meat indicated that 9 were field strains and 6 were vaccine-like (98% to 99.7% nucleotide homology with the Ingelvac RespPRRS/Repro vaccine). One of these 6 strains presented an intermediate 2-6-2 restriction fragment length polymorphism (RFLP) cut pattern and the others showed the characteristic 2-5-2 RFLP pattern of the vaccine strain. All strains sequenced were determined to be North American strains. In only 1 of the 19 PRRSV-positive meat samples could PRRSV be isolated. To test the potential infectivity of meat samples containing residual PRRSV, 11 of the PCR-positive meat samples (weighing 1.05 to 1.8 kg) were each used in feeding experiments of 2 PRRSV antibody-negative specific pathogen-free pigs of 9 wk of age. Samples were cut into several pieces and fed to each pair of pigs on 2 consecutive days. Each pig pair was housed in a separate cubicle and serum samples were collected at –7, 0, 7, 14, and 20 to 21 days post exposure. Seven pig pairs were found to be infected by PRRSV following ingestion of meat samples, including meat samples containing vaccine-like virus, as judged by the demonstration of PRRSV antibodies and/or PRRSV nucleic acid in the serum. In summary, the present study indicated that low residual quantities of PRRSV may be found in a small percentage of pig meat collected at slaugtherhouses. Furthermore, when this meat was fed raw to pigs in the experimental setting designed, pigs could be infected by PRRSV. PMID:15581220

Application of metabonomics on an experimental model of fibrosis and cirrhosis induced by thioacetamide in rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Constantinou, Maria A.; Department of Forensic Medicine and Toxicology, Medical School, University of Athens, 75, Mikras Asias str., 11527 Athens; Theocharis, Stamatios E.

2007-01-01

Metabonomics has already been used to discriminate different pathological states in biological fields. The metabolic profiles of chronic experimental fibrosis and cirrhosis induction in rats were investigated using {sup 1}H NMR spectroscopy of liver extracts and serum combined with pattern recognition techniques. Rats were continuously administered with thioacetamide (TAA) in the drinking water (300 mg TAA/L), and sacrificed on 1st, 2nd, and 3rd month of treatment. {sup 1}H NMR spectra of aqueous and lipid liver extracts, together with serum were subjected to Principal Component Analysis (PCA). Liver portions were also subjected to histopathological examination and biochemical determination of malondialdehyde (MDA).more » Liver fibrosis and cirrhosis were progressively induced in TAA-treated rats, verified by the histopathological examination and the alterations of MDA levels. TAA administration revealed a number of changes in the {sup 1}H NMR spectra compared to control samples. The performance of PCA in liver extracts and serum, discriminated the control samples from the fibrotic and cirrhotic ones. Metabolic alterations revealed in NMR spectra during experimental liver fibrosis and cirrhosis induction, characterize the stage of fibrosis and could be illustrated by subsequent PCA of the spectra. Additionally, the PCA plots of the serum samples presented marked clustering during fibrosis progression and could be extended in clinical diagnosis for the management of cirrhotic patients.« less

Use of ProteinChip technology for identifying biomarkers of parasitic diseases: the example of porcine cysticercosis (Taenia solium).

PubMed

Deckers, N; Dorny, P; Kanobana, K; Vercruysse, J; Gonzalez, A E; Ward, B; Ndao, M

2008-12-01

Taenia solium cysticercosis is a significant public health problem in endemic countries. The current serodiagnostic techniques are not able to differentiate between infections with viable cysts and infections with degenerated cysts. The objectives of this study were to identify specific novel biomarkers of these different disease stages in the serum of experimentally infected pigs using ProteinChip technology (Bio-Rad) and to validate these biomarkers by analyzing serum samples from naturally infected pigs. In the experimental sample set 30 discriminating biomarkers (p<0.05) were found, 13 specific for the viable phenotype, 9 specific for the degenerated phenotype and 8 specific for the infected phenotype (either viable or degenerated cysts). Only 3 of these biomarkers were also significant in the field samples; however, the peak profiles were not consistent among the two sample sets. Five biomarkers discovered in the sera from experimentally infected pigs were identified as clusterin, lecithin-cholesterol acyltransferase, vitronectin, haptoglobin and apolipoprotein A-I.

Serum corticosteroid binding globulin expression is modulated by fasting in polar bears (Ursus maritimus).

PubMed

Chow, Brian A; Hamilton, Jason; Cattet, Marc R L; Stenhouse, Gordon; Obbard, Martyn E; Vijayan, Mathilakath M

2011-01-01

Polar bears (Ursus maritimus) from several subpopulations undergo extended fasting during the ice-free season. However, the animals appear to conserve protein despite the prolonged fasting, though the mechanisms involved are poorly understood. We hypothesized that elevated concentrations of corticosteroid binding globulin (CBG), the primary cortisol binding protein in circulation, lead to cortisol resistance and provide a mechanism for protein conservation during extended fasting. The metabolic state (feeding vs. fasting) of 16 field sampled male polar bears was determined based on their serum urea to creatinine ratio (>25 for feeding vs. <5 for fasting). There were no significant differences in serum cortisol levels between all male and female polar bears sampled. Serum CBG expression was greater in lactating females relative to non-lactating females and males. CBG expression was significantly higher in fasting males when compared to non-fasting males. This leads us to suggest that CBG expression may serve as a mechanism to conserve protein during extended fasting in polar bears by reducing systemic free cortisol concentrations. This was further supported by a lower serum glucose concentration in the fasting bears. As well, a lack of an enhanced adrenocortical response to acute capture stress supports our hypothesis that chronic hunger is not a stressor in this species. Overall, our results suggest that elevated serum CBG expression may be an important adaptation to spare proteins by limiting cortisol bioavailability during extended fasting in polar bears. Copyright © 2010 Elsevier Inc. All rights reserved.

Multicomponent analysis of radiolytic products in human body fluids using high field proton nuclear magnetic resonance (NMR) spectroscopy

NASA Astrophysics Data System (ADS)

Grootveld, Martin C.; Herz, Herman; Haywood, Rachel; Hawkes, Geoffrey E.; Naughton, Declan; Perera, Anusha; Knappitt, Jacky; Blake, David R.; Claxson, Andrew W. D.

1994-05-01

High field proton Hahn spin-echo nuclear magnetic resonance (NMR) spectroscopy has been employed to investigate radiolytic damage to biomolecules present in intact human body fluids. γ-Radiolysis of healthy or rheumatoid human serum (5.00 kGy) in the presence of atmospheric O 2 gave rise to reproducible elevations in the concentration of NMR-detectable acetate which are predominantly ascribable to the prior oxidation of lactate to pyruvate by hydroxyl radical (·OH) followed by oxidative decarboxylation of pyruvate by radiolytically-generated hydrogen peroxide (H 2O 2) and/or further ·OH radical. Increases in the serum levels of non-protein-bound, low-molecular-mass components such as citrate and glutamine were also observed subsequent to γ-radiolysis, an observation which may reflect their mobilisation from protein binding-sites by ·OH radical, superoxide anion and/or H 2O 2. Moreover, substantial radiolytically-mediated elevations in the concentration of serum formate were also detectable. In addition to the above modifications, γ-radiolysis of inflammatory knee-joint synovial fluid (SF) generated a low-molecular-mass oligosaccharide species derived from the radiolytic fragmentation of hyaluronate. The radiolytically-mediated production of acetate in SF samples was markedly greater than that observed in serum samples, a consequence of the much higher levels of ·OH radical-scavenging lactate present. Indeed, increases in SF acetate concentration were detectable at doses as low as 48 Gy. We conclude that high field proton NMR analysis provides much useful information regarding the relative radioprotectant abilities of endogenous components and the nature, status and levels of radiolytic products generated in intact biofluids. We also suggest that NMR-detectable radiolytic products with associated toxicological properties (e.g. formate) may play a role in contributing to the deleterious effects observed following exposure of living organisms to sources of ionising radiation.

Field application of serodiagnostics to identify elephants with Tuberculosis prior to case confirmation by culture

USDA-ARS?s Scientific Manuscript database

Three serologic methods for antibody detection in elephant tuberculosis (TB), multiantigen print immunoassay (MAPIA), ElephantTB STAT-PAK kit, and DPP VetTB test, were validated prospectively using serial serum samples from 14 captive elephants in 5 countries which were diagnosed with TB by positive...

Comparison of biochemical values in serum and plasma, fresh and frozen plasma, and hemolyzed samples from orange-winged Amazon parrots (Amazona amazonica).

PubMed

Hawkins, Michelle G; Kass, Philip H; Zinkl, Joseph G; Tell, Lisa A

2006-06-01

To the authors' knowledge, on the basis of sample type, storage condition, or hemolysis, differences in serum and plasma biochemical values have not been evaluated in orange-winged Amazon parrots (Amazona amazonica). The purpose of this study was to compare values for biochemical analytes in serum vs plasma, fresh vs frozen plasma, and nonhemolyzed vs hemolyzed samples in orange-winged Amazon parrots. We also compared differences in serum and plasma yield from whole-blood aliquots. Fifteen biochemical analytes were evaluated in paired serum and plasma, fresh and frozen plasma, nonhemolyzed and hemolyzed serum and plasma samples from orange-winged Amazon parrots (n = 10) using a wet reagent analyzer. Hemolysis was assessed qualitatively (visually) and quantitatively (hemoglobin [Hgb] measured spectrophotometrically). Serum and plasma yields from 500-microl whole-blood aliquots were determined from centrifuged samples. Analyte values significantly differed among sample groups, but were still within published reference intervals, with the exception of increases in potassium concentration in markedly hemolyzed serum and plasma samples. Clinically important changes in hemolyzed serum and plasma samples included increases in potassium, phosphorus, and albumin concentrations and lactate dehydrogenase activity. The degree of hemolysis assigned qualitatively did not correlate with quantitative Hgb concentration. A significantly greater yield of plasma (288 +/- 13 microL) than serum (241 +/- 44 microL) was obtained. Significant differences may occur in different sample types, however, only changes in potassium, phosphorus, albumin, and lactate dehydrogenase values in hemolyzed samples were considered clinically relevant. Lack of agreement between qualitative and quantitative Hgb concentration indicates the unreliability of visual estimation. Based on higher sample yield, and lack of clinically relevant differences from serum, plasma is a better sample choice for clinical chemistry analysis in birds.

A comparison of serum and plasma cytokine values using a multiplexed assay in cats.

PubMed

Gruen, Margaret E; Messenger, Kristen M; Thomson, Andrea E; Griffith, Emily H; Paradise, Hayley; Vaden, Shelly; Lascelles, B D X

2016-12-01

Degenerative joint disease (DJD) is highly prevalent in cats, and pain contributes to morbidity. In humans, alterations of cytokine concentrations have been associated with joint deterioration and pain. Similar changes have not been investigated in cats. Cytokine concentrations can be measured using multiplex technology with small samples of serum or plasma, however, serum and plasma are not interchangeable for most bioassays. Correlations for cytokine concentrations between serum and plasma have not been evaluated in cats. To evaluate the levels of detection and agreement between serum and plasma samples in cats. Paired serum and plasma samples obtained from 38 cats. Blood was collected into anti-coagulant free and EDTA Vacutainer ® tubes, serum or plasma extracted, and samples frozen at -80°C until testing. Duplicate samples were tested using a 19-plex feline cytokine/chemokine magnetic bead panel. Agreement between serum and plasma for many analytes was high, however correlation coefficients ranged from -0.01 to 0.97. Results from >50% of samples were below the lower limit of quantification for both serum and plasma for nine analytes, and for an additional three analytes for plasma only. While serum and plasma agreement was generally good, detection was improved using serum samples. Copyright © 2016 Elsevier B.V. All rights reserved.

The relationships between sixteen perfluorinated compound concentrations in blood serum and food, and other parameters, in the general population of South Korea with proportionate stratified sampling method.

PubMed

Kim, Hee-Young; Kim, Seung-Kyu; Kang, Dong-Mug; Hwang, Yong-Sik; Oh, Jeong-Eun

2014-02-01

Serum samples were collected from volunteers of various ages and both genders using a proportionate stratified sampling method, to assess the exposure of the general population in Busan, South Korea to perfluorinated compounds (PFCs). 16 PFCs were investigated in serum samples from 306 adults (124 males and 182 females) and one day composite diet samples (breakfast, lunch, and dinner) from 20 of the serum donors, to investigate the relationship between food and serum PFC concentrations. Perfluorooctanoic acid and perfluorooctanesulfonic acid were the dominant PFCs in the serum samples, with mean concentrations of 8.4 and 13 ng/mL, respectively. Perfluorotridecanoic acid was the dominant PFC in the composite food samples, ranging from

Extended Gate Field-Effect Transistor Biosensors for Point-Of-Care Testing of Uric Acid.

PubMed

Guan, Weihua; Reed, Mark A

2017-01-01

An enzyme-free redox potential sensor using off-chip extended-gate field effect transistor (EGFET) with a ferrocenyl-alkanethiol modified gold electrode has been used to quantify uric acid concentration in human serum and urine. Hexacyanoferrate (II) and (III) ions are used as redox reagent. The potentiometric sensor measures the interface potential on the ferrocene immobilized gold electrode, which is modulated by the redox reaction between uric acid and hexacyanoferrate ions. The device shows a near Nernstian response to uric acid and is highly specific to uric acid in human serum and urine. The interference that comes from glucose, bilirubin, ascorbic acid, and hemoglobin is negligible in the normal concentration range of these interferents. The sensor also exhibits excellent long term reliability and is regenerative. This extended gate field effect transistor based sensor is promising for point-of-care detection of uric acid due to the small size, low cost, and low sample volume consumption.

Method and apparatus for automated processing and aliquoting of whole blood samples for analysis in a centrifugal fast analyzer

DOEpatents

Burtis, C.A.; Johnson, W.F.; Walker, W.A.

1985-08-05

A rotor and disc assembly for use in a centrifugal fast analyzer. The assembly is designed to process multiple samples of whole blood followed by aliquoting of the resultant serum into precisely measured samples for subsequent chemical analysis. The assembly requires minimal operator involvement with no mechanical pipetting. The system comprises: (1) a whole blood sample disc; (2) a serum sample disc; (3) a sample preparation rotor; and (4) an analytical rotor. The blood sample disc and serum sample disc are designed with a plurality of precision bore capillary tubes arranged in a spoked array. Samples of blood are loaded into the blood sample disc by capillary action and centrifugally discharged into cavities of the sample preparation rotor where separation of serum and solids is accomplished. The serum is loaded into the capillaries of the serum sample disc by capillary action and subsequently centrifugally expelled into cuvettes of the analyticaly rotor for conventional methods. 5 figs.

Method and apparatus for automated processing and aliquoting of whole blood samples for analysis in a centrifugal fast analyzer

DOEpatents

Burtis, Carl A.; Johnson, Wayne F.; Walker, William A.

1988-01-01

A rotor and disc assembly for use in a centrifugal fast analyzer. The assembly is designed to process multiple samples of whole blood followed by aliquoting of the resultant serum into precisely measured samples for subsequent chemical analysis. The assembly requires minimal operator involvement with no mechanical pipetting. The system comprises (1) a whole blood sample disc, (2) a serum sample disc, (3) a sample preparation rotor, and (4) an analytical rotor. The blood sample disc and serum sample disc are designed with a plurality of precision bore capillary tubes arranged in a spoked array. Samples of blood are loaded into the blood sample disc in capillary tubes filled by capillary action and centrifugally discharged into cavities of the sample preparation rotor where separation of serum and solids is accomplished. The serum is loaded into the capillaries of the serum sample disc by capillary action and subsequently centrifugally expelled into cuvettes of the analytical rotor for analysis by conventional methods.

Fast and accurate determination of K, Ca, and Mg in human serum by sector field ICP-MS.

PubMed

Yu, Lee L; Davis, W Clay; Nuevo Ordonez, Yoana; Long, Stephen E

2013-11-01

Electrolytes in serum are important biomarkers for skeletal and cellular health. The levels of electrolytes are monitored by measuring the Ca, Mg, K, and Na in blood serum. Many reference methods have been developed for the determination of Ca, Mg, and K in clinical measurements; however, isotope dilution thermal ionization mass spectrometry (ID-TIMS) has traditionally been the primary reference method serving as an anchor for traceability and accuracy to these secondary reference methods. The sample matrix must be separated before ID-TIMS measurements, which is a slow and tedious process that hindered the adoption of the technique in routine clinical measurements. We have developed a fast and accurate method for the determination of Ca, Mg, and K in serum by taking advantage of the higher mass resolution capability of the modern sector field inductively coupled plasma mass spectrometry (SF-ICP-MS). Each serum sample was spiked with a mixture containing enriched (44)Ca, (26)Mg, and (41)K, and the (42)Ca(+):(44)Ca(+), (24)Mg(+):(26)Mg(+), and (39)K(+):(41)K(+) ratios were measured. The Ca and Mg ratios were measured in medium resolution mode (m/Δm ≈ 4 500), and the K ratio in high resolution mode (m/Δm ≈ 10 000). Residual (40)Ar(1)H(+) interference was still observed but the deleterious effects of the interference were minimized by measuring the sample at K > 100 ng g(-1). The interferences of Sr(++) at the two Ca isotopes were less than 0.25 % of the analyte signal, and they were corrected with the (88)Sr(+) intensity by using the Sr(++):Sr(+) ratio. The sample preparation involved only simple dilutions, and the measurement using this sample preparation approach is known as dilution-and-shoot (DNS). The DNS approach was validated with samples prepared via the traditional acid digestion approach followed by ID-SF-ICP-MS measurement. DNS and digested samples of SRM 956c were measured with ID-SF-ICP-MS for quality assurance, and the results (mean ± expanded uncertainty in mg dL(-1) unit) for Ca (DNS = 10.14 ± 0.13, digested = 10.11 ± 0.10), Mg (DNS = 2.093 ± 0.008, digested = 2.098 ± 0.007), and K (DNS = 15.48 ± 0.11, digested = 15.50 ± 0.28) were in good agreement with the certified values (Ca = 10.17 ± 0.06, Mg = 2.084 ± 0.023, K = 15.55 ± 0.13). Major sources of uncertainty are sample measurement, spike calibration, and instrument factor including mass discrimination of the spectrometer and the detector deadtime.

Use of dried blood spots for the determination of serum concentrations of tamoxifen and endoxifen.

PubMed

Jager, N G L; Rosing, H; Schellens, J H M; Beijnen, J H; Linn, S C

2014-07-01

The anti-estrogenic effect of tamoxifen is suggested to be mainly attributable to its metabolite (Z)-endoxifen, and a minimum therapeutic threshold for (Z)-endoxifen in serum has been proposed. The objective of this research was to establish the relationship between dried blood spot (DBS) and serum concentrations of tamoxifen and (Z)-endoxifen to allow the use of DBS sampling, a simple and patient-friendly alternative to venous sampling, in clinical practice. Paired DBS and serum samples were obtained from 50 patients using tamoxifen and analyzed using HPLC-MS/MS. Serum concentrations were calculated from DBS concentrations using the formula calculated serum concentration = DBS concentration/([1-haematocrit (Hct)] + blood cell-to-serum ratio × Hct). The blood cell-to-serum ratio was determined ex vivo by incubating a batch of whole blood spiked with both analytes. The average Hct for female adults was imputed as a fixed value. Calculated and analyzed serum concentrations were compared using weighted Deming regression. Weighted Deming regression analysis comparing 44 matching pairs of DBS and serum samples showed a proportional bias for both analytes. Serum concentrations were calculated using [Tamoxifen] serum, calculated  = [Tamoxifen] DBS /0.779 and [(Z)-Endoxifen] serum, calculated = [(Z)-Endoxifen] DBS /0.663. Calculated serum concentrations were within 20 % of analyzed serum concentrations in 84 and 100 % of patient samples for tamoxifen and (Z)-endoxifen, respectively. In conclusion, DBS concentrations of tamoxifen and (Z)-endoxifen were equal to serum concentrations after correction for Hct and blood cell-to-serum ratio. DBS sampling can be used in clinical practice.

[Expression of high mobility group box-1 in the lung tissue and serum of patients with pulmonary tuberculosis].

PubMed

Yang, Xiao-min; Yang, Hua

2013-07-01

To explore the expression of high mobility group box-1 (HMGB1) in the lung tissue and serum of patients with pulmonary tuberculosis and to explore its relationship with tumor necrosis factor (TNF)-α and interleukin(IL)-1β. Sixty samples of lung tissues were obtained from patients with pulmonary tuberculosis who had underwent pneumonectomy in Department of Chest Surgery, First Affiliated Hospital of Zunyi Medical College from June 2010 to December 2011. At the same period, 40 normal lung samples were also obtained from patients with pulmonary contusion and lung cancer by surgical resections as the control group. The mRNA expressions of HMGB1 was detected by reverse transcription-polymerase chain reaction (RT-PCR), and the protein level of HMGB1 was measured by immunohistochemical staining of tissue microarrays in lung tissue. Blood samples were taken from 89 patients with active pulmonary tuberculosis (pulmonary tuberculosis group), including hematogenous disseminated pulmonary tuberculosis (type II) in 35 cases and secondary pulmonary tuberculosis (type III) in 54 cases, and 50 healthy volunteers (control group). Furthermore, the 54 patients with secondary pulmonary tuberculosis were divided into different subgroups according to cavity formation and the lung fields involved: patients without lung cavity (35 cases) vs those with lung cavity (19 cases), patients with involvement of <2 lung fields (31 cases) vs ≥ 2 lung fields (23 cases). Serum concentration of HMGB1, TNF-α and IL-1β were detected by ELISA. Two sample t-test was used to compare date among groups, liner correlation analysis was established for correlation analysis. The average optical density of HMGB1 in pulmonary tuberculosis (69 ± 29) was significantly higher than that in normal lung tissue (22 ± 12) (t = 2.389, P < 0.05). The mRNA relative transcript levels of HMGB1 in pulmonary tuberculosis (786 ± 86) was significantly higher than that in normal lung tissue (202 ± 60) (t = 3.872, P < 0.01). The serum concentration of HMGB1, TNF-α and IL-1β in the pulmonary tuberculosis group were (5.0 ± 3.2) µg/L, (118 ± 77) ng/L and (33 ± 20) ng/L, respectively, which were significantly higher than those in the control group [(1.7 ± 1.0) µg/L, (40 ± 11) ng/L and (18 ± 12) ng/L, respectively], the respective t values being -0.928, 4.268 and 11.064, all P < 0.01. In the subgroup of patients with hematogenous disseminated pulmonary tuberculosis, the serum concentration of HMGB1 and TNF-α[ (6.4 ± 3.3) µg/L, (147 ± 89) ng/L] were significantly higher than those in patients with secondary pulmonary tuberculosis [(4.1 ± 2.7) µg/L, (85 ± 37) ng/L] (t = 3.643 and t = 3.111, both P < 0.01). HMGB1 were correlated positively with TNF-α and IL-1β (r = 0.722 and r = 0.620, P < 0.01, respectively, n = 89) in the pulmonary tuberculosis group. Overexpression of HMGB1 in the lung tissue and serum of patients with pulmonary tuberculosis may play an important role in the inflammatory response of pulmonary tuberculosis. The measurement of serum HMGB1 is useful to evaluate the severity of disease.

Measurement of titanium in hip-replacement patients by inductively coupled plasma optical emission spectroscopy.

PubMed

Harrington, Chris F; McKibbin, Craig; Rahanu, Monika; Langton, David; Taylor, Andrew

2017-05-01

Background Patients with metal-on-metal hip replacements require testing for cobalt and chromium. There may also be a need to test for titanium, which is used in the construction of the femoral stem in total hip replacements. It is not possible to use quadrupole inductively coupled plasma mass spectrometry due to interferences. Methods Titanium was measured using inductively coupled plasma optical emission spectroscopy using the emission line at 336.1 nm and Y (internal standard) at 371.0 nm. Internal quality control materials were prepared for blood and serum and concentrations assigned using a sector field-inductively coupled plasma mass spectrometer. A candidate whole blood certified reference material was also evaluated. Results The method had detection and quantitation limits of 0.6 and 1.9 µg/L, respectively. The respective bias (%) and measurement uncertainty ( U) (k = 2) were 3.3% and 2.0 µg/L (serum) and - 1.0% and 1.4 µg/L (whole blood). The respective repeatability and intermediate precision (%) were 5.1% and 10.9% (serum) and 2.4% and 8.6% (whole blood). The concentration of titanium was determined in patients' samples, serum (median = 2.4 µg/L, n = 897) and whole blood (median = 2.4 µg/L, n = 189). Serum is recommended for monitoring titanium in patients, since the concentration is higher than in whole blood and the matrix less problematic. In hip fluid samples, the concentrations were much higher (mean 58.5 µg/L, median 5.1 µg/L, n = 83). Conclusions A method based on inductively coupled plasma optical emission spectroscopy was developed and validated for measuring titanium in clinical samples.

The importance of sample collection when using single cytokine levels and systemic cytokine profiles as biomarkers--a comparative study of serum versus plasma samples.

PubMed

Tvedt, Tor Henrik Anderson; Rye, Kristin Paulsen; Reikvam, Håkon; Brenner, Annette K; Bruserud, Øystein

2015-03-01

Cytokines, soluble adhesion molecules and metalloproteinases can be detected in human serum or plasma samples. Such systemic levels are widely used as biomarkers in epidemiological and clinical studies. We prepared serum samples and three types of plasma samples (EDTA, heparin, citric acid) from 20 healthy individuals. The levels of 31 cytokines, four soluble adhesion molecules and eight matrix metalloproteinases were analyzed by Luminex technology. Most mediators showed detectable levels in both plasma and serum. Several mediators that can be released by platelets showed increased serum levels, especially CCL5 and CD40L, but for the other mediators the serum levels did not correlate with peripheral blood platelet counts and for these last mediators serum and plasma levels often showed strong correlations. The use of bivalirudin for anticoagulation significantly increased and citric acid combined with platelet inhibitors (ticagrelor, acetylsalicylic acid plus prostaglandin E2) did not alter plasma levels of platelet-store mediators compared with citric acid alone. The impact of sample preparation differed between mediators; for many mediators strong correlations were seen between serum and plasma levels even when absolute levels differed. Soluble adhesion molecule levels showed only minor differences between samples. Unsupervised hierarchical clustering suggested that the effect of sampling/preparation was strongest for serum and heparin plasma samples. Careful standardization of sample preparation is usually necessary when analyzing systemic mediator levels, and differences caused by sample preparation should be considered as a possible explanation if studies show conflicting results. Copyright © 2015 Elsevier B.V. All rights reserved.

Proteins and antibodies in serum, plasma, and whole blood-size characterization using asymmetrical flow field-flow fractionation (AF4).

PubMed

Leeman, Mats; Choi, Jaeyeong; Hansson, Sebastian; Storm, Matilda Ulmius; Nilsson, Lars

2018-05-29

The analysis of aggregates of therapeutic proteins is crucial in order to ensure efficacy and patient safety. Typically, the analysis is performed in the finished formulation to ensure that aggregates are not present. An important question is, however, what happens to therapeutic proteins, with regard to oligomerization and aggregation, after they have been administrated (i.e., in the blood). In this paper, the separation of whole blood, plasma, and serum is shown using asymmetric flow field-flow fractionation (AF4) with a minimum of sample pre-treatment. Furthermore, the analysis and size characterization of a fluorescent antibody in blood plasma using AF4 are demonstrated. The results show the suitability and strength of AF4 for blood analysis and open new important routes for the analysis and characterization of therapeutic proteins in the blood.

Melatonin secretion and puberty in female lambs exposed to environmental electric and magnetic fields

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, J.M. Jr.; Stormshak, F.; Thompson, J.M.

This study determined whether chronic exposure of female lambs to the electric and magnetic fields (EMF) of a high voltage transmission line can alter pineal secretion of melatonin and the normal occurrence of puberty. Twenty female Suffolk lambs were assigned randomly in equal numbers to a control and a treatment group. Treatment from 2 to 10 mo of age consisted of continuous exposure within the electrical environment of a 500-kV transmission line (mean electric field 6 kV/m, mean magnetic field 40 mG). Treated lambs were penned directly beneath the transmission line; control lambs were maintained in a pen of similarmore » construction 229 m from the line where EMF were at ambient levels (mean electric field < 10 V/m, mean magnetic field < 0.3 mG). Melatonin was analyzed by RIA in serum of blood samples collected at 0.5-3-h intervals over eight 48-h periods. To assess attainment of puberty, serum concentrations of progesterone were determined by RIA from blood samples collected twice weekly beginning at 19 wk of age. Concentrations of circulating melatonin in control and treated lambs were low during daylight hours and increased during nighttime hours. The characteristic pattern of melatonin secretion during nighttime (amplitude, phase, and duration) did not differ between control and treatment groups. Age at puberty and number of subsequent estrous cycles also did not differ between groups. These data suggest that chronic exposure of developing female sheep to 60-Hz environmental EMF does not affect the mechanisms underlying the generation of the circadian pattern of melatonin secretion or the mechanisms involved in the onset of reproductive activity.« less

Effect of delayed serum separation and storage temperature on serum glucose concentration in horse, dog, alpaca, and sturgeon.

PubMed

Collicutt, Nancy B; Garner, Bridget; Berghaus, Roy D; Camus, Melinda S; Hart, Kelsey

2015-03-01

Although delays between blood sample collection and analysis are common in veterinary medicine, the effect of prolonged serum-clot contact time on serum glucose concentration is not well established and species differences have not been elucidated. The objective was to investigate the effect of storage time and temperature on serum glucose concentration in stored whole blood samples from horse, dog, alpaca, and sturgeon. Whole blood specimens were divided into 7 no-additive tubes and serum was separated from one sample within one hour, serving as the reference sample. The remaining samples were stored at 4°C and 25°C, then centrifuged and serum glucose measured by automated analysis at 2, 4, and 8 hours postcollection. Glucose concentrations were compared using linear mixed models. The decline in serum glucose concentration for all samples stored at 4°C was not statistically significant, except for the 8-hour samples from sturgeon and dog. At 25°C, serum glucose concentration was comparable to reference values at 2 hours in sturgeon and alpaca, but significantly lower at 4 and 8 hours in those species, and at all time points in equine and canine specimens, being most prominent after 8 hours of storage in canine specimens. Storage at 4°C limits serum glucose decline for at least 4 hours in all species tested and up to 8 hours in specimens of horse and alpaca. At 25°C, serum-clot contact time should not exceed 1 hour in equine and canine samples, and 2 hours in specimens from alpaca and sturgeon. © 2014 American Society for Veterinary Clinical Pathology.

Changes to Serum Sample Tube and Processing Methodology Does Not Cause Inter-Individual Variation in Automated Whole Serum N-Glycan Profiling in Health and Disease

PubMed Central

Shubhakar, Archana; Kalla, Rahul; Nimmo, Elaine R.; Fernandes, Daryl L.; Satsangi, Jack; Spencer, Daniel I. R.

2015-01-01

Introduction Serum N-glycans have been identified as putative biomarkers for numerous diseases. The impact of different serum sample tubes and processing methods on N-glycan analysis has received relatively little attention. This study aimed to determine the effect of different sample tubes and processing methods on the whole serum N-glycan profile in both health and disease. A secondary objective was to describe a robot automated N-glycan release, labeling and cleanup process for use in a biomarker discovery system. Methods 25 patients with active and quiescent inflammatory bowel disease and controls had three different serum sample tubes taken at the same draw. Two different processing methods were used for three types of tube (with and without gel-separation medium). Samples were randomised and processed in a blinded fashion. Whole serum N-glycan release, 2-aminobenzamide labeling and cleanup was automated using a Hamilton Microlab STARlet Liquid Handling robot. Samples were analysed using a hydrophilic interaction liquid chromatography/ethylene bridged hybrid(BEH) column on an ultra-high performance liquid chromatography instrument. Data were analysed quantitatively by pairwise correlation and hierarchical clustering using the area under each chromatogram peak. Qualitatively, a blinded assessor attempted to match chromatograms to each individual. Results There was small intra-individual variation in serum N-glycan profiles from samples collected using different sample processing methods. Intra-individual correlation coefficients were between 0.99 and 1. Unsupervised hierarchical clustering and principal coordinate analyses accurately matched samples from the same individual. Qualitative analysis demonstrated good chromatogram overlay and a blinded assessor was able to accurately match individuals based on chromatogram profile, regardless of disease status. Conclusions The three different serum sample tubes processed using the described methods cause minimal inter-individual variation in serum whole N-glycan profile when processed using an automated workstream. This has important implications for N-glycan biomarker discovery studies using different serum processing standard operating procedures. PMID:25831126

Changes to serum sample tube and processing methodology does not cause Intra-Individual [corrected] variation in automated whole serum N-glycan profiling in health and disease.

PubMed

Ventham, Nicholas T; Gardner, Richard A; Kennedy, Nicholas A; Shubhakar, Archana; Kalla, Rahul; Nimmo, Elaine R; Fernandes, Daryl L; Satsangi, Jack; Spencer, Daniel I R

2015-01-01

Serum N-glycans have been identified as putative biomarkers for numerous diseases. The impact of different serum sample tubes and processing methods on N-glycan analysis has received relatively little attention. This study aimed to determine the effect of different sample tubes and processing methods on the whole serum N-glycan profile in both health and disease. A secondary objective was to describe a robot automated N-glycan release, labeling and cleanup process for use in a biomarker discovery system. 25 patients with active and quiescent inflammatory bowel disease and controls had three different serum sample tubes taken at the same draw. Two different processing methods were used for three types of tube (with and without gel-separation medium). Samples were randomised and processed in a blinded fashion. Whole serum N-glycan release, 2-aminobenzamide labeling and cleanup was automated using a Hamilton Microlab STARlet Liquid Handling robot. Samples were analysed using a hydrophilic interaction liquid chromatography/ethylene bridged hybrid(BEH) column on an ultra-high performance liquid chromatography instrument. Data were analysed quantitatively by pairwise correlation and hierarchical clustering using the area under each chromatogram peak. Qualitatively, a blinded assessor attempted to match chromatograms to each individual. There was small intra-individual variation in serum N-glycan profiles from samples collected using different sample processing methods. Intra-individual correlation coefficients were between 0.99 and 1. Unsupervised hierarchical clustering and principal coordinate analyses accurately matched samples from the same individual. Qualitative analysis demonstrated good chromatogram overlay and a blinded assessor was able to accurately match individuals based on chromatogram profile, regardless of disease status. The three different serum sample tubes processed using the described methods cause minimal inter-individual variation in serum whole N-glycan profile when processed using an automated workstream. This has important implications for N-glycan biomarker discovery studies using different serum processing standard operating procedures.

IL8 and IL16 levels indicate serum and plasma quality.

PubMed

Kofanova, Olga; Henry, Estelle; Quesada, Rocio Aguilar; Bulla, Alexandre; Linares, Hector Navarro; Lescuyer, Pierre; Shea, Kathi; Stone, Mars; Tybring, Gunnel; Bellora, Camille; Betsou, Fay

2018-02-09

Longer pre-centrifugation times alter the quality of serum and plasma samples. Markers for such delays in sample processing and hence for the sample quality, have been identified. Twenty cytokines in serum, EDTA plasma and citrate plasma samples were screened for changes in concentration induced by extended blood pre-centrifugation delays at room temperature. The two cytokines that showed the largest changes were further validated for their "diagnostic performance" in identifying serum or plasma samples with extended pre-centrifugation times. In this study, using R&D Systems ELISA kits, EDTA plasma samples and serum samples with a pre-centrifugation delay longer than 24 h had an IL16 concentration higher than 313 pg/mL, and an IL8 concentration higher than 125 pg/mL, respectively. EDTA plasma samples with a pre-centrifugation delay longer than 48 h had an IL16 concentration higher than 897 pg/mL, citrate plasma samples had an IL8 concentration higher than 21.5 pg/mL and serum samples had an IL8 concentration higher than 528 pg/mL. These robust and accurate tools, based on simple and commercially available ELISA assays can greatly facilitate qualification of serum and plasma legacy collections with undocumented pre-analytics.

Serum samples can be substituted by plasma samples for the diagnosis of paratuberculosis.

PubMed

Goodridge, Amador; Correa, Ricardo; Castro, Paul; Escobar, Cecilia; de Waard, Jacobus H

2013-10-01

Employing plasma samples rather than serum samples for serological paratuberculosis diagnosis is practical, especially when bovine TB is assessed in the same cattle herd with the gamma interferon bovine avian (IFN-γ BA) test. We demonstrate that antibody titers in serum and plasma samples, utilizing the PARACHECK(®) ELISA kit, are highly comparable (Cohen's kappa test, k=0.955). We conclude that serum can be replaced with plasma in this commercially available antibody detection assay resulting in working hour savings for sampling and blood sample work-up and cost reductions for materials and sample storage. Copyright © 2013 Elsevier B.V. All rights reserved.

Detection of Human Immunodeficiency Virus Type 1 (HIV-1) RNA in Pools of Sera Negative for Antibodies to HIV-1 and HIV-2

PubMed Central

Morandi, Pierre-Alain; Schockmel, Gérard A.; Yerly, Sabine; Burgisser, Philippe; Erb, Peter; Matter, Lukas; Sitavanc, Radan; Perrin, Luc

1998-01-01

A total of 234 pools were prepared from 10,692 consecutive serum samples negative for antibodies to human immunodeficiency virus type 1 (HIV-1) and HIV-2 collected at five virological laboratories (average pool size, 45 serum samples). Pools were screened for the presence of HIV-1 RNA by a modified commercial assay (Amplicor HIV-1 Monitor test) which included an additional polyethylene glycol (PEG) precipitation step prior to purification of viral RNA (PEG Amplicor assay). The sensitivity of this assay for HIV-1 RNA detection in individual serum samples within pools matches that of standard commercial assays for individual serum samples, i.e., 500 HIV-1 RNA copies per ml. Five pools were identified as positive, and each one contained one antibody-negative, HIV-1 RNA-positive serum sample, corresponding to an average of 1 infected sample per 2,138 serum samples. Retrospective analysis revealed that the five HIV-1 RNA-positive specimens originated from individuals who had symptomatic primary HIV-1 infection at the time of sample collection and who were also positive for p24 antigenemia. We next assessed the possibility of performing the prepurification step by high-speed centrifugation (50,000 × g for 80 min) of 1.5-ml pools containing 25 μl of 60 individual serum samples, of which only 1 contained HIV-1 RNA (centrifugation Amplicor assay). The sensitivity of this assay also matches the sensitivities of standard commercial assays for HIV-1 RNA detection in individual serum samples. The results demonstrate that both assays with pooled sera can be applied to the screening of large numbers of serum samples in a time- and cost-efficient manner. PMID:9620372

Magnetic solid phase extraction of gemfibrozil from human serum and pharmaceutical wastewater samples utilizing a β-cyclodextrin grafted graphene oxide-magnetite nano-hybrid.

PubMed

Abdolmohammad-Zadeh, Hossein; Talleb, Zeynab

2015-03-01

A magnetic solid phase extraction method based on β-cyclodextrin (β-CD) grafted graphene oxide (GO)/magnetite (Fe3O4) nano-hybrid as an innovative adsorbent was developed for the separation and pre-concentration of gemfibrozil prior to its determination by spectrofluorometry. The as-prepared β-CD/GO/Fe3O4 nano-hybrid possesses the magnetism property of Fe3O4 nano-particles that makes it easily manipulated by an external magnetic field. On the other hand, the surface modification of GO by β-CD leads to selective separation of the target analyte from sample matrices. The structure and morphology of the synthesized adsorbent were characterized using powder X-ray diffraction, Fourier transform infrared spectroscopy, and field emission scanning electron microscopy. The experimental factors affecting the extraction/pre-concentration and determination of the analyte were investigated and optimized. Under the optimized experimental conditions, the calibration graph was linear in the range between 10 and 5000 pg mL(-1) with a correlation coefficient of 0.9989. The limit of detection and enrichment factor for gemfibrozil were 3 pg mL(-1) and 100, respectively. The maximum sorption capacity of the adsorbent for gemfibrozil was 49.8 mg g(-1). The method was successfully applied to monitoring gemfibrozil in human serum and pharmaceutical wastewaters samples with recoveries in the range of 96.0-104.0% for the spiked samples. Copyright © 2014 Elsevier B.V. All rights reserved.

No effect of mobile phone-like RF exposure on patients with atopic dermatitis.

PubMed

Johansson, Amanda; Forsgren, Sture; Stenberg, Berndt; Wilén, Jonna; Kalezic, Nebojsa; Sandström, Monica

2008-07-01

This study investigates the effect of exposure to a mobile phone-like radiofrequency (RF) electromagnetic field on people with atopic dermatitis (AD). Fifteen subjects with AD were recruited and matched with 15 controls without AD. The subjects were exposed for 30 min to an RF field at 1 W/kg via an indoor base station antenna attached to a 900 MHz GSM mobile phone. Blood samples for ELISA analysis of the concentration of substance P (SP), tumor necrosis factor receptor 1 (TNF R1), and brain derived neurotrophic factor (BDNF) in serum were drawn before and after the provocation (exposure/sham). Baseline heart rate and heart rate variability, local blood flow, and electrodermal activity were also recorded. No significant differences between the subject groups were found for baseline neurophysiological data. The cases displayed a serum concentration of TNF R1 significantly higher than the control subjects and a significantly lower serum concentration of BDNF in the baseline condition. For SP there was no difference between groups. However, no effects related to RF exposure condition were encountered for any of the measured substances. As to symptoms, a possible correlation with exposure could not be evaluated, due to too few symptom reports. The result of the study does not support the hypothesis of an effect of mobile phone-like RF exposure on serum levels of SP, TNF R1, and BDNF in persons with AD.

Comparison of Schmallenberg virus antibody levels detected in milk and serum from individual cows.

PubMed

Daly, Janet M; King, Barnabas; Tarlinton, Rachael A; Gough, Kevin C; Maddison, Ben C; Blowey, Roger

2015-03-11

Schmallenberg virus (SBV) is a recently emerged virus of ruminants in Europe. Enzyme-linked immunosorbent assays (ELISA) are commonly used to detect SBV-specific antibodies in bulk tank milk samples to monitor herd exposure to infection. However, it has previously been shown that a bulk tank milk sample can test positive even though the majority of cows within the herd are seronegative for SBV antibodies. Development of a pen-side test to detect antibodies in individual milk samples would potentially provide a cheaper test (for which samples are obtained non-invasively) than testing individual serum samples by ELISA. Therefore, the aim of this study was to investigate the agreement between antibody levels measured in milk and serum. Corresponding milk and serum samples from 88 cows in two dairy herds in the UK were tested for presence of immunoglobulin G antibodies to SBV using a commercially-available indirect ELISA. A serum neutralisation test (NT) was also performed as a gold standard assay. The ELISA values obtained for the bulk tank milk samples corresponded with the mean values for individual milk samples from each herd (bulk tank milk values were 58% and 73% and mean individual milk values 50% and 63% for herds A and B, respectively). Of the 88 serum samples tested in the NT, 82 (93%) were positive. Although at higher antibody levels, the ELISA values tended to be higher for the individual milk samples than for the corresponding serum samples, the positive predictive value for milk samples was 98% and for serum samples 94%. The serum ELISA was more likely to give false positive results around the lower cut-off value of the assay. The results indicate that testing of individual milk samples for antibodies against SBV by ELISA could be used to inform decisions in the management of dairy herds such as which, if any, animals to vaccinate.

Comparison of digoxin concentration in plastic serum tubes with clot activator and heparinized plasma tubes.

PubMed

Dukić, Lora; Simundić, Ana-Maria; Malogorski, Davorin

2014-01-01

Sample type recommended by the manufacturer for the digoxin Abbott assay is either serum collected in glass tubes or plasma (sodium heparin, lithium heparin, citrate, EDTA or oxalate as anticoagulant) collected in plastic tubes. In our hospital samples are collected in plastic tubes. Our hypothesis was that the serum sample collected in plastic serum tube can be used interchangeably with plasma sample for measurement of digoxin concentration. Our aim was verification of plastic serum tubes for determination of digoxin concentration. Concentration of digoxin was determined simultaneously in 26 venous blood plasma (plastic Vacuette, LH Lithium heparin) and serum (plastic Vacuette, Z Serum Clot activator; both Greiner Bio-One GmbH, Kremsmünster, Austria) samples, on Abbott AxSYM analyzer using the original Abbott Digoxin III assay (Abbott, Wiesbaden, Germany). Tube comparability was assessed using the Passing Bablok regression and Bland-Altman plot. Serum and plasma digoxin concentrations are comparable. Passing Bablok intercept (0.08 [95% CI = -0.10 to 0.20]) and slope (0.99 [95% CI = 0.92 to 1.11]) showed there is no constant or proportional error. Blood samples drawn in plastic serum tubes and plastic plasma tubes can be interchangeably used for determination of digoxin concentration.

Comparison of digoxin concentration in plastic serum tubes with clot activator and heparinized plasma tubes

PubMed Central

Dukić, Lora; Šimundić, Ana-Maria; Malogorski, Davorin

2014-01-01

Introduction: Sample type recommended by the manufacturer for the digoxin Abbott assay is either serum collected in glass tubes or plasma (sodium heparin, lithium heparin, citrate, EDTA or oxalate as anticoagulant) collected in plastic tubes. In our hospital samples are collected in plastic tubes. Our hypothesis was that the serum sample collected in plastic serum tube can be used interchangeably with plasma sample for measurement of digoxin concentration. Our aim was verification of plastic serum tubes for determination of digoxin concentration. Materials and methods: Concentration of digoxin was determined simultaneously in 26 venous blood plasma (plastic Vacuette, LH Lithium heparin) and serum (plastic Vacuette, Z Serum Clot activator; both Greiner Bio-One GmbH, Kremsmünster, Austria) samples, on Abbott AxSYM analyzer using the original Abbott Digoxin III assay (Abbott, Wiesbaden, Germany). Tube comparability was assessed using the Passing Bablok regression and Bland-Altman plot. Results: Serum and plasma digoxin concentrations are comparable. Passing Bablok intercept (0.08 [95% CI = −0.10 to 0.20]) and slope (0.99 [95% CI = 0.92 to 1.11]) showed there is no constant or proportional error. Conclusion: Blood samples drawn in plastic serum tubes and plastic plasma tubes can be interchangeably used for determination of digoxin concentration. PMID:24627723

Negative interference by rheumatoid factor in alpha-fetoprotein chemiluminescent microparticle immunoassay.

PubMed

Wang, Hui; Bi, Xiaohui; Xu, Lei; Li, Yirong

2017-01-01

Background Rheumatoid factor causes positive interference in multiple immunoassays. Recently, negative interference has also been found in immunoassays in the presence of rheumatoid factor. The chemiluminescent microparticle immunoassay is widely used to determine serum alpha-fetoprotein. However, it is not clear whether the presence of rheumatoid factor in the serum causes interference in the chemiluminescent microparticle immunoassay of alpha-fetoprotein. Methods Serum alpha-fetoprotein was determined using the ARCHITECT alpha-fetoprotein assay. The estimation of alpha-fetoprotein recovery was carried out in samples prepared by diluting high-concentration alpha-fetoprotein serum with rheumatoid factor-positive or rheumatoid factor-negative serum. Paramagnetic microparticles coated with hepatitis B surface antigen-anti-HBs complexes were used to remove rheumatoid factor from the serum. Results The average recovery of alpha-fetoprotein was 88.4% and 93.8% in the rheumatoid factor-positive and rheumatoid factor-negative serum samples, respectively. The recovery of alpha-fetoprotein was significantly lower in the rheumatoid factor-positive serum samples than in the rheumatoid factor-negative serum samples. In two of five rheumatoid factor-positive samples, a large difference was found (9.8%) between the average alpha-fetoprotein recoveries in the serially diluted and initial recoveries. Fourteen rheumatoid factor-positive serum samples were pretreated with hepatitis B surface antigen-anti-HBs complex-coated paramagnetic microparticles. The alpha-fetoprotein concentrations measured in the pretreated samples increased significantly. Conclusions It was concluded that the alpha-fetoprotein chemiluminescent microparticle immunoassay is susceptible to interference by rheumatoid factor, leading to significantly lower results. Eliminating the incidence of negative interference from rheumatoid factor should be an important goal for immunoassay providers. In the meantime, laboratorians must remain alert to the negative interference by rheumatoid factor, and in some cases, pretreat rheumatoid factor-positive samples with blocking or absorbing reagents.

Quantitative proteomic analysis for high-throughput screening of differential glycoproteins in hepatocellular carcinoma serum

PubMed Central

Gao, Hua-Jun; Chen, Ya-Jing; Zuo, Duo; Xiao, Ming-Ming; Li, Ying; Guo, Hua; Zhang, Ning; Chen, Rui-Bing

2015-01-01

Objective Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths. Novel serum biomarkers are required to increase the sensitivity and specificity of serum screening for early HCC diagnosis. This study employed a quantitative proteomic strategy to analyze the differential expression of serum glycoproteins between HCC and normal control serum samples. Methods Lectin affinity chromatography (LAC) was used to enrich glycoproteins from the serum samples. Quantitative mass spectrometric analysis combined with stable isotope dimethyl labeling and 2D liquid chromatography (LC) separations were performed to examine the differential levels of the detected proteins between HCC and control serum samples. Western blot was used to analyze the differential expression levels of the three serum proteins. Results A total of 2,280 protein groups were identified in the serum samples from HCC patients by using the 2D LC-MS/MS method. Up to 36 proteins were up-regulated in the HCC serum, whereas 19 proteins were down-regulated. Three differential glycoproteins, namely, fibrinogen gamma chain (FGG), FOS-like antigen 2 (FOSL2), and α-1,6-mannosylglycoprotein 6-β-N-acetylglucosaminyltransferase B (MGAT5B) were validated by Western blot. All these three proteins were up-regulated in the HCC serum samples. Conclusion A quantitative glycoproteomic method was established and proven useful to determine potential novel biomarkers for HCC. PMID:26487969

Validation of serological tests for the detection of antibodies against Treponema pallidum in nonhuman primates.

PubMed

Knauf, Sascha; Dahlmann, Franziska; Batamuzi, Emmanuel K; Frischmann, Sieghard; Liu, Hsi

2015-03-01

There is evidence to suggest that the yaws bacterium (Treponema pallidum ssp. pertenue) may exist in non-human primate populations residing in regions where yaws is endemic in humans. Especially in light of the fact that the World Health Organizaiton (WHO) recently launched its second yaws eradication campaign, there is a considerable need for reliable tools to identify treponemal infection in our closest relatives, African monkeys and great apes. It was hypothesized that commercially available serological tests detect simian anti-T. pallidum antibody in serum samples of baboons, with comparable sensitivity and specificity to their results on human sera. Test performances of five different treponemal tests (TTs) and two non-treponemal tests (NTTs) were evaluated using serum samples of 57 naturally T. pallidum-infected olive baboons (Papio anubis) from Lake Manyara National Park in Tanzania. The T. pallidum particle agglutination assay (TP-PA) was used as a gold standard for comparison. In addition, the overall infection status of the animals was used to further validate test performances. For most accurate results, only samples that originated from baboons of known infection status, as verified in a previous study by clinical inspection, PCR and immunohistochemistry, were included. All tests, TTs and NTTs, used in this study were able to reliably detect antibodies against T. pallidum in serum samples of infected baboons. The sensitivity of TTs ranged from 97.7-100%, while specificity was between 88.0-100.0%. The two NTTs detected anti-lipoidal antibodies in serum samples of infected baboons with a sensitivity of 83.3% whereas specificity was 100%. For screening purposes, the TT Espline TP provided the highest sensitivity and specificity and at the same time provided the most suitable format for use in the field. The enzyme immune assay Mastblot TP (IgG), however, could be considered as a confirmatory test.

Detection of Canine Distemper Virus Nucleoprotein RNA by Reverse Transcription-PCR Using Serum, Whole Blood, and Cerebrospinal Fluid from Dogs with Distemper

PubMed Central

Frisk, A. L.; König, M.; Moritz, A.; Baumgärtner, W.

1999-01-01

Reverse transcription-PCR (RT-PCR) was used to detect canine distemper virus (CDV) nucleoprotein (NP) RNA in serum, whole blood, and cerebrospinal fluid (CSF) samples from 38 dogs with clinically suspected distemper. Results were correlated to clinical findings, anti-CDV neutralizing antibody titers, postmortem findings, and demonstration of CDV NP antigen by immunohistochemistry. The specificity of the RT-PCR was ensured by amplification of RNA from various laboratory CDV strains, restriction enzyme digestion, and Southern blot hybridization. In 29 of 38 dogs, CDV infection was confirmed by postmortem examination and immunohistochemistry. The animals displayed the catarrhal, systemic, and nervous forms of distemper. Seventeen samples (serum, whole blood, or CSF) from dogs with distemper were tested with three sets of primers targeted to different regions of the NP gene of the CDV Onderstepoort strain. Expected amplicons were observed in 82, 53, and 41% of the 17 samples, depending upon the primer pair used. With the most sensitive primer pair (primer pair I), CDV NP RNA was detected in 25 of 29 (86%) serum samples and 14 of 16 (88%) whole blood and CSF samples from dogs with distemper but not in body fluids from immunohistochemically negative dogs. Nucleotide sequence analysis of five RT-PCR amplicons from isolates from the field revealed few silent point mutations. These isolates exhibited greater homology to the Rockborn (97 to 99%) than to the Onderstepoort (95 to 96%) CDV strain. In summary, although the sensitivity of the RT-PCR for detection of CDV is strongly influenced by the location of the selected primers, this nucleic acid detection system represents a highly specific and sensitive method for the antemortem diagnosis of distemper in dogs, regardless of the form of distemper, humoral immune response, and viral antigen distribution. PMID:10523566

Specific IgA and IgG antibodies in paired serum and breast milk samples in human strongyloidiasis.

PubMed

Mota-Ferreira, Daniela M L; Gonçalves-Pires, Maria do Rosário F; Júnior, Alvaro Ferreira; Sopelete, Mônica C; Abdallah, Vânia O S; Costa-Cruz, Julia M

2009-02-01

Strongyloidiasis, caused by the nematode Strongyloides stercoralis, is one of the major worldwide parasitic infections in humans. Breastfeeding may offer a potential protection against this infection. Feces, serum and milk samples were obtained from 90 lactating women from Clinical Hospital of Universidade Federal de Uberlândia, Brazil. The fecal samples were collected for parasitological diagnosis and the serum and milk samples were examined for specific S. stercoralis IgA and IgG antibodies using the indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assay (ELISA). Fecal examination showed that the rate of prevalence of S. stercoralis infection in the lactating women was 4.4%. IFAT manifested a 16.7% positivity rate for specific IgA antibody in serum and a 28.9% rate in milk samples; specific IgG was 41.1% in serum and 25.5% in milk samples. According to ELISA the positivity rate for specific IgA antibody was 21.1% in serum and 42.2% in milk samples; specific IgG was 40% in serum and 18.9% in milk samples. In serum samples, these immunological tests showed a concurrence of 91.1% and 94.4%, respectively, in detecting specific IgA and IgG antibodies. In milk samples, they showed a concurrence of 70% and 78.9%, respectively, in detecting specific IgA and IgG antibodies. There was a statistically significant difference between concordant and discordant results of immunological tests (P<0.0001). IFAT and ELISA highly concurred in their detection of specific S. stercoralis IgA and IgG antibodies in serum and in milk samples reconfirming prior studies that the serological method is a complement to the direct diagnosis of the parasite, and suggesting that immunological methods using milk samples can also be helpful. Furthermore, in endemic areas, infants may acquire antibodies to S. stercoralis from breast milk, possibly, contributing to the enhancement of specific mucosal immunity against this parasite.

A contribution for the definition of serum chemistry values in captive adults Antillean manatees (Trichechus manatus manatus Linnaeus, 1758).

PubMed

Silva, F M O; Vergara-Parente, J E; Gomes, J K N; Teixeira, M N; Lima, R P

2007-04-01

Serum chemistry analyses represents a fundamental tool for the diagnosis and understanding of diseases in marine mammals. Although several studies are being conducted within the field of clinical pathology, haematological and serum chemistry data for Antillean manatees are deficient. The purpose of this study was to determine serum chemistry values for captive Antillean manatees within the CMA/Ibama facility in Brazil. Serum samples were obtained from five captive adult Antillean manatees fed with seagrass and analysed for aspartate aminotransferase, alanine aminotransferase, bilirubin, alkaline phosphatase, urea, creatinine, glucose, triglycerides, cholesterol, total protein, albumin, globulin, phosphate, chloride, calcium and uric acid. Blood chemistry parameters were determined using a semi-automatic analyzer. Maximum, minimum, mean and standard deviations were calculated for each serum chemistry parameter. Differences on the values of males and females were verified using an unpaired Student's t-test. All the parameters analysed were similar between sexes, with exception of AP, which was higher in females (191.43 +/- 31.86 U/l). Alanine aminotransferase and uric acid values for Trichechus manatus manatus are reported for the first time in this paper. This study is the first to report serum chemistry parameter values for long-term captive male and female Antillean manatees. Therefore, the lower values of albumin, phosphate, chloride, cholesterol and triglycerides obtained here highlight the importance of clinical pathology during health monitoring of captive marine mammals.

Infection dynamics and acute phase response of an Actinobacillus pleuropneumoniae field isolate of moderate virulence in pigs.

PubMed

Gómez-Laguna, Jaime; Islas, Armando; Muñoz, Dennis; Ruiz, Alvaro; Villamil, Aura; Carrasco, Librado; Quezada, Manuel

2014-10-10

Actinobacillus pleuropneumoniae, the causative agent of porcine contagious pleuropneumonia (PCP), causes significant economic losses associated mainly with growth stunting of animals. Although serotypes can be distinguished according to their virulence, most of the studies are focused in A. pleuropneumoniae infections with virulent serotypes. There is little information regarding the role of acute phase proteins (APPs) and proinflammatory cytokines in infections with isolates of mild or moderate virulence. Thus, the present study aims to evaluate the kinetics of infection with an A. pleuropneumoniae serotype 6 (Ap6) field isolate of moderate virulence and the changes in the serum concentration of specific antibodies and different APPs and proinflammatory cytokines. Control animals showed no clinical signs or lesions throughout the study. Infected animals showed increased rectal temperature, respiratory distress and depression from 24hpi, and typical gross and microscopic lesions of PCP from 6hpi onwards. Ap6 was isolated from nasal swabs of four out of five inoculated animals at 24hpi, and from nasal swabs, tonsil and lung samples from all inoculated animals at 72hpi. Specific antibodies against Ap6 or changes in the serum concentration of IL-1β, IL-10 and TNF-α were not detected throughout the study. The serum concentration of IL-6 increased from 6hpi as well as serum A amyloid, C-reactive protein and haptoglobin from 24hpi onwards. Our results highlight the onset of the acute phase response after the infection with a field isolate of A. pleuropneumoniae of moderate virulence from 24hpi onwards which may be of interest in the study of the pathogenesis of this disease. Copyright © 2014 Elsevier B.V. All rights reserved.

Immunization against East Coast Fever in field cattle with low infectivity Theileria parva stabilate--preliminary assessment.

PubMed

Mbassa, G K; Kweka, L E; Dulla, P N

1998-05-01

Two Theileria parva sporozoite stabilates stored at -196 degrees C, then at -70 degrees C for six weeks (stabilate 1) and more than six months (stabilate 2) were inoculated into four eight-month old male calves, 1 and 2 (stabilate 1), and 3 and 4 (stabilate 2). Calves 1 and 2 developed pyrexia, enlargement of lymph nodes, and the former died of East Coast Fever. Calves 3 and 4 showed slight enlargement of lymph nodes without fever. Lymph node smears from all calves (from day 10 to 20 post-inoculation) showed lymphoblasts, phagocytic macrophages, and schizonts. Piroplasms were detected in erythrocytes in blood smears from calves 1 and 2 but not in calves 3 and 4. Calves 2, 3 and 4 recovered without any treatment while calf 1 died of East Coast Fever on day 20. Serum samples from recovered calves taken on day 30 of the experiment were positive for antischizont antibodies to T. parva at 1:640 dilution, but pre-inoculation serum samples were negative. Stabilate 2 was used to immunize 64 Boran, Friesian, Ayrshire and crosses with Zebu cattle in four herds with 25% reduction of oxytetracycline dose. All the animals except one calf recovered without any severe reactions. The latter died of disease other than ECF after the monitoring period was over (day 24). Day 30 post-inoculation serum samples were positive for T. parva antischizont antibodies. A follow-up of the remaining animals for over one year revealed no further ECF incidences in these herds. This experiment shows the loss of infectivity of the vaccine stored at temperatures higher than -196 degrees C. dependent on the duration. However, despite the lack of clinical signs in calves 3 and 4, there was cellular response and antibody production, and the stabilate for vaccine against East Coast fever can thus be stored prior to use at higher than -196 degrees C and still maintain capability to produce antibodies in field cattle, eliminating the use of oxytetracycline and monitoring. The vaccine will be cheaper and easier to use and the requirement for liquid nitrogen in the field reduced and the scale of application of the vaccine widened.

[Intraocular and serum antibody titers to Leptospira in 150 horses with equine recurrent uveitis (ERU) subjected to vitrectomy].

PubMed

Wollanke, B; Gerhards, H; Brem, S; Kopp, H; Meyer, P

1998-04-01

Between February 1993 and July 1997, 150 horses suffering from recurrent uveitis were subjected to parsplana vitrectomy. In these horses, antibody titers to Leptospira serovars were determined in serum samples and in samples from diluted vitreous collected during vitrectomy. Although the vitreous samples were diluted with 250 ml of balanced salt solution, in 86 of the 150 vitreous samples (= 57%) the antibody titers were higher than in the serum samples. Additionally, serum samples from 77 horses suffering from ERU, but which were not subjected to vitrectomy, and serum samples from 97 horses with clinically normal eyes were analyzed for antibodies to Leptospira serovars. Among the 227 horses with ERU (150 treated surgically, 77 treated conservatively) 50 horses (50 of 227 = 22%) had serum antibody titers to Leptospira serovars of > or = 1:800. Among the 97 horses with clinically normal eyes, 24 horses (24 of 97 = 25%) had serum antibody titers to Leptospira serovars of > or = 1:800. In undiluted vitreous samples from 20 horses with clinically normal eyes, no antibody titers to Leptospira serovars could be detected. Among the 150 horses with ERU, 90 animals (90 of 150 = 60%) had antibody titers of > or = 1:100 in the diluted vitreous samples, the difference being highly significant (p < 0.001). The findings are discussed in relation to the etiology of recurrent uveitis in horses.

Dielectrophoretic Isolation and Detection of cfc-DNA Nanoparticulate Biomarkers and Virus from Blood

PubMed Central

Sonnenberg, Avery; Marciniak, Jennifer Y.; McCanna, James; Krishnan, Rajaram; Rassenti, Laura; Kipps, Thomas J.; Heller, Michael J.

2015-01-01

Dielectrophoretic (DEP) microarray devices allow important cellular nanoparticulate biomarkers and virus to be rapidly isolated, concentrated and detected directly from clinical and biological samples. A variety of sub-micron nanoparticulate entities including cell free circulating (cfc) DNA, mitochondria and virus can be isolated into DEP high-field areas on microelectrodes, while blood cells and other micron-size entities become isolated into DEP low-field areas between the microelectrodes. The nanoparticulate entities are held in the DEP high-field areas while cells are washed away along with proteins and other small molecules which are not affected by the DEP electric fields. DEP carried out on 20 µL of whole blood obtained from Chronic Lymphocytic Leukemia (CLL) patients showed a considerable amount of SYBR Green stained DNA fluorescent material concentrated in the DEP high-field regions. Whole blood obtained from healthy individuals showed little or no fluorescent DNA materials in the DEP high-field regions. Fluorescent T7 bacteriophage virus could be isolated directly from blood samples, and fluorescently stained mitochondria could be isolated from biological buffer samples. Using newer DEP microarray devices, high molecular weight (hmw) DNA could be isolated from serum and detected at levels as low as 8–16 ng/mL. PMID:23436471

Evaluation of transmission infrared spectroscopy and digital and optical refractometers to identify low immunoglobulin G concentrations in alpaca serum

PubMed Central

Elsohaby, Ibrahim; Burns, Jennifer J.; Riley, Christopher B.; McClure, J. Trenton

2017-01-01

This study aimed to evaluate the digital Brix and optical serum total protein (STP) refractometers for measuring concentrations of serum immunoglobulin G (IgG) in alpacas and compare them to IgG concentrations measured by the reference method of radial immunodiffusion (RID) assay. The appropriate cutoff point for Brix and STP refractometers and the transmission infrared (TIR) spectroscopy method was determined for low IgG concentrations (< 10 g/L). Serum samples were collected from alpacas (N = 169) and tested by both refractometers. The correlation between Brix % and STP was high [correlation coefficient (r) = 0.99]. However, the correlation coefficients between Brix % and STP with serum RID-IgG concentrations were only 0.56 and 0.55, respectively. Twenty-one (12.4%) of 169 alpaca serum samples had IgG concentrations of < 10 g/L. Using receiver operator characteristic curve (ROC) analysis, the optimal cutoff points for the TIR assay, digital Brix, and optical STP refractometers for assessing low IgG (RID < 10 g/L) were 13 g/L, 8.8%, and 50 g/L, respectively. The TIR assay showed higher sensitivity (Se = 95.2%) and specificity (Sp = 96.8%) than either the digital Brix (Se = 90.5% and Sp = 65.5%) or optical STP (Se = 81% and Sp = 73.7%) refractometers for assessing alpacas with low IgG. In conclusion, the Brix and STP refractometers lack accuracy in measuring alpaca IgG concentrations, but may be useful for screening animals for low serum IgG. However, the TIR assay with a cutoff point of 13 g/L was more appropriate for identifying low IgG than either refractometer. Another study that focuses on neonatal crias is recommended in order to evaluate the usefulness of these assays for field diagnosing of failure of transfer of passive immunity (FTPI). PMID:28725112

Evaluation of transmission infrared spectroscopy and digital and optical refractometers to identify low immunoglobulin G concentrations in alpaca serum.

PubMed

Elsohaby, Ibrahim; Burns, Jennifer J; Riley, Christopher B; McClure, J Trenton

2017-07-01

This study aimed to evaluate the digital Brix and optical serum total protein (STP) refractometers for measuring concentrations of serum immunoglobulin G (IgG) in alpacas and compare them to IgG concentrations measured by the reference method of radial immunodiffusion (RID) assay. The appropriate cutoff point for Brix and STP refractometers and the transmission infrared (TIR) spectroscopy method was determined for low IgG concentrations (< 10 g/L). Serum samples were collected from alpacas (N = 169) and tested by both refractometers. The correlation between Brix % and STP was high [correlation coefficient (r) = 0.99]. However, the correlation coefficients between Brix % and STP with serum RID-IgG concentrations were only 0.56 and 0.55, respectively. Twenty-one (12.4%) of 169 alpaca serum samples had IgG concentrations of < 10 g/L. Using receiver operator characteristic curve (ROC) analysis, the optimal cutoff points for the TIR assay, digital Brix, and optical STP refractometers for assessing low IgG (RID < 10 g/L) were 13 g/L, 8.8%, and 50 g/L, respectively. The TIR assay showed higher sensitivity (Se = 95.2%) and specificity (Sp = 96.8%) than either the digital Brix (Se = 90.5% and Sp = 65.5%) or optical STP (Se = 81% and Sp = 73.7%) refractometers for assessing alpacas with low IgG. In conclusion, the Brix and STP refractometers lack accuracy in measuring alpaca IgG concentrations, but may be useful for screening animals for low serum IgG. However, the TIR assay with a cutoff point of 13 g/L was more appropriate for identifying low IgG than either refractometer. Another study that focuses on neonatal crias is recommended in order to evaluate the usefulness of these assays for field diagnosing of failure of transfer of passive immunity (FTPI).

Deployment of a Reverse Transcription Loop-Mediated Isothermal Amplification Test for Ebola Virus Surveillance in Remote Areas in Guinea.

PubMed

Kurosaki, Yohei; Magassouba, N'Faly; Bah, Hadja Aïssatou; Soropogui, Barré; Doré, Amadou; Kourouma, Fodé; Cherif, Mahamoud Sama; Keita, Sakoba; Yasuda, Jiro

2016-10-15

To strengthen the laboratory diagnostic capacity for Ebola virus disease (EVD) in the remote areas of Guinea, we deployed a mobile field laboratory and implemented reverse transcription loop-mediated isothermal amplification (RT-LAMP) for postmortem testing. We tested 896 oral swab specimens and 21 serum samples, using both RT-LAMP and reverse transcription-polymerase chain reaction (RT-PCR). Neither test yielded a positive result, and the results from RT-LAMP and RT-PCR were consistent. More than 95% of the samples were tested within 2 days of sample collection. These results highlight the usefulness of the RT-LAMP assay as an EVD diagnostic testing method in the field or remote areas. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Volatile Organic Compound (VOC) Analysis For Disease Detection: Proof Of Principle For Field Studies Detecting Paratuberculosis And Brucellosis

NASA Astrophysics Data System (ADS)

Knobloch, Henri; Köhler, Heike; Nicola, Commander; Reinhold, Petra; Turner, Claire; Chambers, Mark

2009-05-01

A proof of concept investigation was performed to demonstrate that two independent infectious diseases of cattle result in different patterns of volatile organic compounds (VOC) in the headspace of serum samples detectable using an electronic nose (e-nose). A total of 117 sera from cattle naturally infected with Mycobacterium avium subsp. paratuberculosis (paraTB, n = 43) or Brucella sp. (n = 26) and sera from corresponding control animals (n = 48) were randomly and analysed blind to infection status using a ST214 e-nose (Scensive Ltd, Leeds, UK). Samples were collected under non-standardised conditions on different farms from the UK (brucellosis) and Germany (paraTB). The e-nose could differentiate the sera from brucellosis infected, paraTB infected and healthy animals at the population level, but the technology used was not suitable for determination of the disease status of individual animals. Nevertheless, the data indicate that there are differences in the sensor responses depending on the disease status, and therefore, it shows the potential of VOC analysis from serum headspace samples for disease detection.

Determining bovine viral diarrhea and infectious bovine rhinotracheitis infections in dairy cattle using precolostral blood.

PubMed

Baillargeon, Paul; Arango-Sabogal, Juan C; Wellemans, Vincent; Fecteau, Gilles

2017-04-01

The objective of this study was to determine if precolostral blood samples are useful to detect apparent fetal infections with bovine viral diarrhea (BVD) and infectious bovine rhinotracheitis (IBR) viruses. A convenience sample of 317 sera from 50 Canadian herds was used in the study. Antibody level was measured using 2 commercial IBR and BVD ELISA kits. Precolostral status of sera was confirmed on 304 samples using serum gamma-glutamyl transferase activity. Postcolostral serum samples yielded a higher proportion of positive results to IBR (OR = 86; 95% CI: 17.8 to 415.7) and BVD (OR = 199.3; 95% CI: 41.7 to 952.3) than did precolostral samples. All positive precolostral serum samples ( n = 7 of 304) originated from calves born to vaccinated cows. Postcolostral positive serum samples ( n = 11 of 13) originated mostly (60%) from calves born to non-vaccinated cows. Precolostral serum sampling can detect apparent fetal infections in a herd.

Drug residues in serum of dogs receiving anticancer chemotherapy.

PubMed

Knobloch, A; Mohring, S A I; Eberle, N; Nolte, I; Hamscher, G; Simon, D

2010-01-01

The presence of drug residues in blood samples can represent an occupational hazard. However, studies on cytotoxic drug residues in serum of dogs are lacking in veterinary oncology. To evaluate possible occupational hazards associated with handling of blood samples from dogs receiving oncolytic drugs 7 days after treatment. Twenty-seven client-owned dogs treated for lymphoma or mast cell tumors with vincristine, vinblastine, cyclophosphamide, or doxorubicin. Prospective, observational study. Serum samples were either taken 7 days after administration of vincristine, cyclophosphamide, doxorubicin (lymphoma), and vinblastine (mast cell tumor), or 1-2 days after the last concurrent oral administration of cyclophosphamide (mast cell tumor). Additionally, serum was collected within 5 minutes of treatment. Measurement of drug residues in serum was performed by liquid chromatography tandem mass spectrometry (LC/MS/MS). In 33 samples collected within 5 minute of treatment, the median serum concentrations were vincristine: 37 microg/L (range: 11-87 microg/L), vinblastine: 13 microg/L (range: 13-35 microg/L), cyclophosphamide: 2,484 microg/L (range: 1,209-2,778 microg/L), doxorubicin: 404 microg/L (range: 234-528 microg/L). In 81 serum samples collected 7 days after treatment vinblastine (7 microg/L) was detected in 1 sample, and cyclophosphamide (7 and 9 microg/L) in 2 samples collected 1-2 days after oral administration of cyclophosphamide. Medications were not detected in any of the other samples. Handling of blood samples from dogs receiving oncolytic chemotherapy 7 days after treatment with vincristine, vinblastine, cyclophosphamide, and doxorubicin should not present a health hazard.

A rapid field test for the measurement of bovine serum immunoglobulin G using attenuated total reflectance infrared spectroscopy.

PubMed

Elsohaby, Ibrahim; Hou, Siyuan; McClure, J Trenton; Riley, Christopher B; Shaw, R Anthony; Keefe, Gregory P

2015-08-20

Following the recent development of a new approach to quantitative analysis of IgG concentrations in bovine serum using transmission infrared spectroscopy, the potential to measure IgG levels using technology and a device better designed for field use was investigated. A method using attenuated total reflectance infrared (ATR) spectroscopy in combination with partial least squares (PLS) regression was developed to measure bovine serum IgG concentrations. ATR spectroscopy has a distinct ease-of-use advantage that may open the door to routine point-of-care testing. Serum samples were collected from calves and adult cows, tested by a reference RID method, and ATR spectra acquired. The spectra were linked to the RID-IgG concentrations and then randomly split into two sets: calibration and prediction. The calibration set was used to build a calibration model, while the prediction set was used to assess the predictive performance and accuracy of the final model. The procedure was repeated for various spectral data preprocessing approaches. For the prediction set, the Pearson's and concordance correlation coefficients between the IgG measured by RID and predicted by ATR spectroscopy were both 0.93. The Bland Altman plot revealed no obvious systematic bias between the two methods. ATR spectroscopy showed a sensitivity for detection of failure of transfer of passive immunity (FTPI) of 88 %, specificity of 100 % and accuracy of 94 % (with IgG <1000 mg/dL as the FTPI cut-off value). ATR spectroscopy in combination with multivariate data analysis shows potential as an alternative approach for rapid quantification of IgG concentrations in bovine serum and the diagnosis of FTPI in calves.

Evaluation of enzyme-linked immunosorbent assays performed on milk and serum samples for detection of neosporosis and leukosis in lactating dairy cows

PubMed Central

Walsh, Robert B.; Kelton, David F.; Hietala, Sharon K.; Duffield, Todd F.

2013-01-01

Serum and milk samples from 1229 cows on 22 Ontario dairy farms were individually tested for antibodies specific for bovine leukosis virus (BLV) and Neospora caninum by enzyme-linked immunosorbent assay (ELISA). Antibodies against BLV were present in 361 serum samples (29.4%) and 369 milk samples (30.0%). Comparing the 2 tests, agreement was almost perfect (k = 0.86; 95% CI = 0.83 to 0.90) and the proportions of samples positive were not significantly different (P = 0.56). Both tests identified the same 3 herds free of bovine leukosis virus. Antibodies against N. caninum were detected in 138 serum samples (11.2%), and 111 milk samples (9.0%). Agreement between the 2 tests was moderate (k = 0.52; 95% CI = 0.43 to 0.59). Four herds were free of neosporosis by the serum test, while 10 herds were negative by the milk test. The ELISA on milk samples facilitates sample collection to classify herds free of BLV; the milk N. caninum ELISA was less reliable in predicting herd-level infection. PMID:24082160

Comparison of serum and plasma measurements of Müllerian inhibiting substance.

PubMed

Merhi, Zaher O; Messerlian, Geralyn M; Minkoff, Howard; Eklund, Elizabeth E; Macura, Jerzy; Feldman, Joseph; Rodriguez, Carlos; Seifer, David B

2008-06-01

The authors sought to determine whether measurement of plasma Müllerian inhibiting substance (MIS) is a suitable substitute for measurement of serum MIS. Eighteen samples of serum and plasma were examined that were drawn simultaneously. Levels of MIS were measured with an ELISA kit, and plasma levels were studied in parallel to serum samples. A 98.5% correlation was found between serum and plasma MIS values.

Evaluation of pharmacokinetics and the stability of daptomycin in serum at various temperatures.

PubMed

Ogami, Chika; Tsuji, Yasuhiro; Kasai, Hidefumi; Hiraki, Yoichi; Yamamoto, Yoshihiro; Matsunaga, Kazuhisa; Karube, Yoshiharu; To, Hideto

2017-04-01

Daptomycin exhibits concentration-dependent antibacterial activity. By monitoring daptomycin serum concentrations, clinicians may be able to predict the effectiveness of treatments for infections more accurately. However, it has been reported that daptomycin concentrations in plasma samples stored at -20°C decrease approximately 25% after 4 weeks. The aim of this study was to evaluate the stability of daptomycin in serum at various temperatures. Daptomycin serum samples were prepared and stored at different temperatures. The stability of daptomycin under various conditions was evaluated by sequential measurements of concentration. Although the loss of concentration of daptomycin in serum samples stored in freezers (-80°C and -20°C) was less than 10% after 168days (6 months), the concentrations in samples stored in a refrigerator (4°C) decreased by more than 70% over the same period. Furthermore, daptomycin concentrations in serum samples stored at close to body temperature (35°C, 37°C, and 39°C) decreased by more than 50% after only 24h. The results of the present study demonstrate that the measurement of serum concentrations of daptomycin needs to be performed rapidly. Furthermore, the degradation of daptomycin in serum may be involved in its elimination from the living body. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Evaluation of PCR methods for detection of Brucella strains from culture and tissues.

PubMed

Çiftci, Alper; İça, Tuba; Savaşan, Serap; Sareyyüpoğlu, Barış; Akan, Mehmet; Diker, Kadir Serdar

2017-04-01

The genus Brucella causes significant economic losses due to infertility, abortion, stillbirth or weak calves, and neonatal mortality in livestock. Brucellosis is still a zoonosis of public health importance worldwide. The study was aimed to optimize and evaluate PCR assays used for the diagnosis of Brucella infections. For this aim, several primers and PCR protocols were performed and compared with Brucella cultures and biological material inoculated with Brucella. In PCR assays, genus- or species-specific oligonucleotide primers derived from 16S rRNA sequences (F4/R2, Ba148/928, IS711, BruP6-P7) and OMPs (JPF/JPR, 31ter/sd) of Brucella were used. All primers except for BruP6-P7 detected the DNA from reference Brucella strains and field isolates. In spiked blood, milk, and semen samples, F4-R2 primer-oriented PCR assays detected minimal numbers of Brucella. In spiked serum and fetal stomach content, Ba148/928 primer-oriented PCR assays detected minimal numbers of Brucella. Field samples collected from sheep and cattle were examined by bacteriological methods and optimized PCR assays. Overall, sensitivity of PCR assays was found superior to conventional bacteriological isolation. Brucella DNA was detected in 35.1, 1.1, 24.8, 5.0, and 8.0% of aborted fetus, blood, milk, semen, and serum samples by PCR assays, respectively. In conclusion, PCR assay in optimized conditions was found to be valuable in sensitive and specific detection of Brucella infections of animals.

Effect of Repeated Freezing and Thawing on 18 Clinical Chemistry Analytes in Rat Serum

PubMed Central

Kale, Vijay P; Patel, Sweta G; Gunjal, Prashant S; Wakchaure, Santosh U; Sundar, Rajesh S; Ranvir, Ramchandra K; Jain, Mukul R

2012-01-01

In a preclinical research laboratory, using serum samples that have been frozen and thawed repeatedly is sometimes unavoidable when needing to confirm previous results or perform additional analysis. Here we determined the effects of multiple cycles of refrigeration or freezing and thawing of rat serum at 3 temperature conditions for different storage times on clinical chemistry analytes. Serum samples obtained from adult Wistar rats were stored at 2 to 8 °C and −10 to −20 °C for as long as 72 h and at −70 °C for as long as 30 d. At different time points (24, 48, and 72 h for samples stored at 2 to 8 °C or −10 to −20 °C and 1, 7, and 30 d for samples stored at −70 °C), the samples were brought to room temperature, analyzed, and then stored again at the designated temperature. The results obtained after each storage cycle were compared with those obtained from the initial analysis of fresh samples. Of the 18 serum analytes evaluated, 14 were stable without significant changes, even after 3 freeze–thaw cycles at the tested temperature ranges. Results from this study will help researchers working with rat serum to interpret the biochemical data obtained from serum samples that have been frozen and thawed repeatedly. PMID:23043814

Measurement of rivaroxaban and apixaban in serum samples of patients

PubMed Central

Harenberg, Job; Krämer, Sandra; Du, Shanshan; Zolfaghari, Shabnam; Schulze, Astrid; Krämer, Roland; Weiss, Christel; Wehling, Martin; Lip, Gregory Y H

2014-01-01

Background The determination of rivaroxaban and apixaban from serum samples of patients may be beneficial in specific clinical situations when additional blood sampling for plasma and thus the determination of factor Xa activity is not feasible or results are not plausible. Materials and methods The primary aim of this study was to compare the concentrations of rivaroxaban and apixaban in serum with those measured in plasma. Secondary aims were the performance of three different chromogenic methods and concentrations in patients on treatment with rivaroxaban 10 mg od (n = 124) or 20 mg od (n = 94) or apixaban 5 mg bid (n = 52) measured at different time. Results Concentrations of rivaroxaban and apixaban in serum were about 20–25% higher compared with plasma samples with a high correlation (r = 0·79775–0·94662) using all assays (all P < 0·0001). The intraclass correlation coefficients were about 0·90 for rivaroxaban and 0·55 for apixaban. Mean rivaroxaban concentrations were higher at 2 and 3 h compared with 1 and 12 h after administration measured from plasma and serum samples (all P-values < 0·05) and were not different between 1 vs. 12 h (plasma and serum). Conclusions The results indicate that rivaroxaban and apixaban concentrations can be determined specifically from serum samples. PMID:24931429

Chemometric resolution of fully overlapped CE peaks: quantitation of carbamazepine in human serum in the presence of several interferences.

PubMed

Vera-Candioti, Luciana; Culzoni, María J; Olivieri, Alejandro C; Goicoechea, Héctor C

2008-11-01

Drug monitoring in serum samples was performed using second-order data generated by CE-DAD, processed with a suitable chemometric strategy. Carbamazepine could be accurately quantitated in the presence of its main metabolite (carbamazepine epoxide), other therapeutic drugs (lamotrigine, phenobarbital, phenytoin, phenylephrine, ibuprofen, acetaminophen, theophylline, caffeine, acetyl salicylic acid), and additional serum endogenous components. The analytical strategy consisted of the following steps: (i) serum sample clean-up to remove matrix interferences, (ii) data pre-processing, in order to reduce the background and to correct for electrophoretic time shifts, and (iii) resolution of fully overlapped CE peaks (corresponding to carbamazepine, its metabolite, lamotrigine and unexpected serum components) by the well-known multivariate curve resolution-alternating least squares algorithm, which extracts quantitative information that can be uniquely ascribed to the analyte of interest. The analyte concentration in serum samples ranged from 2.00 to 8.00 mg/L. Mean recoveries were 102.6% (s=7.7) for binary samples, and 94.8% (s=13.5) for spiked serum samples, while CV (%)=4.0 was computed for five replicate, indicative of the acceptable accuracy and precision of the proposed method.

Veterinary extension on sampling techniques related to heartwater research.

PubMed

Steyn, H C; McCrindle, C M E; Du Toit, D

2010-09-01

Heartwater, a tick-borne disease caused by Ehrlichia ruminantium, is considered to be a significant cause of mortality amongst domestic and wild ruminants in South Africa. The main vector is Amblyomma hebraeum and although previous epidemiological studies have outlined endemic areas based on mortalities, these have been limited by diagnostic methods which relied mainly on positive brain smears. The indirect fluorescent antibody test (IFA) has a low specificity for heartwater organisms as it cross-reacts with some other species. Since the advent of biotechnology and genomics, molecular epidemiology has evolved using the methodology of traditional epidemiology coupled with the new molecular techniques. A new quantitative real-time polymerase chain reaction (qPCR) test has been developed for rapid and accurate diagnosis of heartwater in the live animal. This method can also be used to survey populations of A. hebraeum ticks for heartwater. Sampling whole blood and ticks for this qPCR differs from routine serum sampling, which is used for many serological tests. Veterinary field staff, particularly animal health technicians, are involved in surveillance and monitoring of controlled and other diseases of animals in South Africa. However, it was found that the sampling of whole blood was not done correctly, probably because it is a new sampling technique specific for new technology, where the heartwater organism is much more labile than the serum antibodies required for other tests. This qPCR technique is highly sensitive and can diagnose heartwater in the living animal within 2 hours, in time to treat it. Poor sampling techniques that decrease the sensitivity of the test will, however, result in a false negative diagnosis. This paper describes the development of a skills training programme for para-veterinary field staff, to facilitate research into the molecular epidemiology of heartwater in ruminants and eliminate any sampling bias due to collection errors. Humane handling techniques were also included in the training, in line with the current focus on improved livestock welfare.

Serological evidence for the presence of influenza D virus in small ruminants.

PubMed

Quast, Megan; Sreenivasan, Chithra; Sexton, Gabriel; Nedland, Hunter; Singrey, Aaron; Fawcett, Linda; Miller, Grant; Lauer, Dale; Voss, Shauna; Pollock, Stacy; Cunha, Cristina W; Christopher-Hennings, Jane; Nelson, Eric; Li, Feng

2015-11-18

Influenza D virus (FLUDV) was isolated from diseased pigs with respiratory disease symptoms in 2011, and since then the new virus has also been spread to cattle. Little is known about the susceptibility of other agricultural animals and poultry to FLUDV. This study was designed to determine if other farm animals such as goats, sheep, chickens, and turkey are possible hosts to this newly emerging influenza virus. 648 goat and sheep serum samples and 250 chicken and turkey serum samples were collected from 141 small ruminant and 25 poultry farms from different geographical locations in the United States and Canada. Serum samples were examined using the hemagglutination inhibition (HI) assay and the sheep and goat samples were further analyzed using the serum neutralization assay. Results of this study showed FLUDV antibodies were detected in 13.5% (17/126) of the sampled sheep farms, and 5.2% (29/557) of tested sheep serum samples were positive for FLUDV antibodies. For the goat results, the FLUDV antibodies were detected in 13.3% (2/15) of the sampled farms, and 8.8% (8/91) of the tested goat serum samples were positive for FLUDV antibodies. Furthermore, all tested poultry serum samples were negative for FLUDV antibodies. Our data demonstrated that sheep and goat are susceptible to FLUDV virus and multiple states in U.S. have this virus infection already in these two species. This new finding highlights a need for future surveillance of FLUDV virus in small ruminants toward better understanding both the origin and natural reservoir of this new virus. Copyright © 2015 Elsevier B.V. All rights reserved.

Optimizing of MALDI-ToF-based low-molecular-weight serum proteome pattern analysis in detection of breast cancer patients; the effect of albumin removal on classification performance.

PubMed

Pietrowska, M; Marczak, L; Polanska, J; Nowicka, E; Behrent, K; Tarnawski, R; Stobiecki, M; Polanski, A; Widlak, P

2010-01-01

Mass spectrometry-based analysis of the serum proteome allows identifying multi-peptide patterns/signatures specific for blood of cancer patients, thus having high potential value for cancer diagnostics. However, because of problems with optimization and standardization of experimental and computational design, none of identified proteome patterns/signatures was approved for diagnostics in clinical practice as yet. Here we compared two methods of serum sample preparation for mass spectrometry-based proteome pattern analysis aimed to identify biomarkers that could be used in early detection of breast cancer patients. Blood samples were collected in a group of 92 patients diagnosed at early (I and II) stages of the disease before the start of therapy, and in a group of age-matched healthy controls (104 women). Serum specimens were purified and analyzed using MALDI-ToF spectrometry, either directly or after membrane filtration (50 kDa cut-off) to remove albumin and other large serum proteins. Mass spectra of the low-molecular-weight fraction (2-10 kDa) of the serum proteome were resolved using the Gaussian mixture decomposition, and identified spectral components were used to build classifiers that differentiated samples from breast cancer patients and healthy persons. Mass spectra of complete serum and membrane-filtered albumin-depleted samples have apparently different structure and peaks specific for both types of samples could be identified. The optimal classifier built for the complete serum specimens consisted of 8 spectral components, and had 81% specificity and 72% sensitivity, while that built for the membrane-filtered samples consisted of 4 components, and had 80% specificity and 81% sensitivity. We concluded that pre-processing of samples to remove albumin might be recommended before MALDI-ToF mass spectrometric analysis of the low-molecular-weight components of human serum Keywords: albumin removal; breast cancer; clinical proteomics; mass spectrometry; pattern analysis; serum proteome.

Comparison of oral fluid collection methods for the molecular detection of hepatitis B virus.

PubMed

Portilho, M M; Mendonça, Acf; Marques, V A; Nabuco, L C; Villela-Nogueira, C A; Ivantes, Cap; Lewis-Ximenez, L L; Lampe, E; Villar, L M

2017-11-01

This study aims to compare the efficiency of four oral fluid collection methods (Salivette, FTA Card, spitting and DNA-Sal) to detect HBV DNA by qualitative PCR. Seventy-four individuals (32 HBV reactive and 42 with no HBV markers) donated serum and oral fluid. In-house qualitative PCR to detect HBV was used for both samples and commercial quantitative PCR for serum. HBV DNA was detected in all serum samples from HBV-infected individuals, and it was not detected in control group. HBV DNA from HBV group was detected in 17 samples collected with Salivette device, 16 samples collected by FTA Card device, 16 samples collected from spitting and 13 samples collected by DNA-Sal device. Samples that corresponded to a higher viral load in their paired serum sample could be detected using all oral fluid collection methods, but Salivette collection device yielded the largest numbers of positive samples and had a wide range of viral load that was detected. It was possible to detect HBV DNA using all devices tested, but higher number of positive samples was observed when samples were collected using Salivette device, which shows high concordance to viral load observed in the paired serum samples. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd. All rights reserved.

MicroRNAs in Serum and Bile of Patients with Primary Sclerosing Cholangitis and/or Cholangiocarcinoma.

PubMed

Voigtländer, Torsten; Gupta, Shashi K; Thum, Sabrina; Fendrich, Jasmin; Manns, Michael P; Lankisch, Tim O; Thum, Thomas

2015-01-01

Patients with primary sclerosing cholangitis (PSC) are at high risk for the development of cholangiocarcinoma (CC). Analysis of micro ribonucleic acid (MiRNA) patterns is an evolving research field in biliary pathophysiology with potential value in diagnosis and therapy. Our aim was to evaluate miRNA patterns in serum and bile of patients with PSC and/or CC. Serum and bile from consecutive patients with PSC (n = 40 (serum), n = 52 (bile)), CC (n = 31 (serum), n = 19 (bile)) and patients with CC complicating PSC (PSC/CC) (n = 12 (bile)) were analyzed in a cross-sectional study between 2009 and 2012. As additional control serum samples from healthy individuals were analyzed (n = 12). The miRNA levels in serum and bile were determined with global miRNA profiling and subsequent miRNA-specific polymerase chain reaction-mediated validation. Serum analysis revealed significant differences for miR-1281 (p = 0.001), miR-126 (p = 0.001), miR-26a (p = 0.001), miR-30b (p = 0.001) and miR-122 (p = 0.034) between patients with PSC and patients with CC. All validated miRNAs were significantly lower in healthy individuals. MiR-412 (p = 0.001), miR-640 (p = 0.001), miR-1537 (p = 0.003) and miR-3189 (p = 0.001) were significantly different between patients with PSC and PSC/CC in bile. Patients with PSC and/or CC have distinct miRNA profiles in serum and bile. Furthermore, miRNA concentrations are different in bile of patients with CC on top of PSC indicating the potential diagnostic value of these miRNAs.

Optimization and validation of indirect ELISA using truncated TssB protein for the serodiagnosis of glanders amongst equines.

PubMed

Singha, Harisankar; Malik, Praveen; Goyal, Sachin K; Khurana, Sandip K; Mukhopadhyay, Chiranjay; Eshwara, Vandana K; Singh, Raj K

2014-01-01

To express truncated TssB protein of Burkholderia mallei and to evaluate its diagnostic efficacy for serological detection of glanders among equines. In an attempt to develop recombinant protein based enzyme-linked immunosorbent assay (ELISA), N-terminal 200 amino acid sequences of B. mallei TssB protein-a type 6 secretory effector protein--were expressed in prokaryotic expression system. Diagnostic potential of recombinant TssB protein was evaluated in indirect ELISA using a panel of glanders positive (n = 49), negative (n = 30), and field serum samples (n = 1811). Cross-reactivity of the assay was assessed with equine disease control serum and human melioidosis positive serum. In comparison to CFT, diagnostic sensitivity and specificity of ELISA were 99.7% and 100%, respectively. The indirect ELISA method using the truncated TssB offered safer and more rapid and efficient means of serodiagnosis of glanders in equines. These data highlight the use of TssB as potential diagnostic antigen for serological diagnosis of glanders.

Brominated flame retardants in the hair and serum samples from an e-waste recycling area in southeastern China: the possibility of using hair for biomonitoring.

PubMed

Liang, Si; Xu, Feng; Tang, Weibiao; Zhang, Zheng; Zhang, Wei; Liu, Lili; Wang, Junxia; Lin, Kuangfei

2016-08-01

Hair samples and paired serum samples were collected from e-waste and urban areas in Wenling of Zhejiang Province, China. The PBDE and DBDPE concentrations in hair and serum samples from e-waste workers were significantly higher than those of non-occupational residents and urban residents. BDE209 was the dominating BFRs in hair and serum samples from the e-waste area, while DBDPE was the major BFRs from the urban area. Statistically significant correlations were observed between hair level and serum level for some substances (BDE209, DBDPE, BDE99, BDE47, BDE28, and BDE17), although the PBDE congener profiles in hair were different from those in the serum. A statistically significant positive correlation between the PBDE concentrations and the working age, as well as gender difference, was observed in e-waste workers. Different sources of PBDEs and DBDPE in three groups were identified by principal component analysis and spearman correlation coefficient. Hair is suggested to be a useful matrix for biomonitoring the PBDE exposure in humans.

Soluble Endoglin (CD105) Serum Level as a Potential Marker in the Management of Head and Neck Paragangliomas.

PubMed

Litwiniuk, Małgorzata; Niemczyk, Kazimierz; Niderla-Bielińska, Justyna; Łukawska-Popieluch, Izabela; Grzela, Tomasz

2017-10-01

To assess the expression of endoglin in head and neck paragangliomas and the soluble endoglin level in serum of paraganglioma patients. Seven tumor samples of patients operated for cervical paraganglioma were assessed, as well as serum samples collected preoperatively, on days 4 and 28 postoperation. Serum level of endoglin in healthy controls was also determined. Tumor samples were subjected to immunofluorescent staining and examined with confocal microscope. The level of soluble endoglin in serum samples was examined using the immunoenzymatic assay (ELISA). Endoglin was highly expressed in all tumor samples. The level of soluble endoglin was significantly higher in paraganglioma patients compared to healthy controls and correlated with the tumor size. The serum level of s-endoglin was reduced after surgical excision of the tumor and remained stable after 4 weeks in all patients with complete resection of the tumor. Endoglin is an important factor in the pathophysiology of head and neck paragangliomas and may be a potential diagnostic and prognostic marker in these types of tumors.

Comparing identified and statistically significant lipids and polar metabolites in 15-year old serum and dried blood spot samples for longitudinal studies: Comparing lipids and metabolites in serum and DBS samples

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kyle, Jennifer E.; Casey, Cameron P.; Stratton, Kelly G.

The use of dried blood spots (DBS) has many advantages over traditional plasma and serum samples such as smaller blood volume required, storage at room temperature, and ability for sampling in remote locations. However, understanding the robustness of different analytes in DBS samples is essential, especially in older samples collected for longitudinal studies. Here we analyzed DBS samples collected in 2000-2001 and stored at room temperature and compared them to matched serum samples stored at -80°C to determine if they could be effectively used as specific time points in a longitudinal study following metabolic disease. Four hundred small molecules weremore » identified in both the serum and DBS samples using gas chromatograph-mass spectrometry (GC-MS), liquid chromatography-MS (LC-MS) and LC-ion mobility spectrometry-MS (LC-IMS-MS). The identified polar metabolites overlapped well between the sample types, though only one statistically significant polar metabolite in a case-control study was conserved, indicating degradation occurs in the DBS samples affecting quantitation. Differences in the lipid identifications indicated that some oxidation occurs in the DBS samples. However, thirty-six statistically significant lipids correlated in both sample types indicating that lipid quantitation was more stable across the sample types.« less

Low-level 14C methane oxidation rate measurements modified for remote field settings

NASA Astrophysics Data System (ADS)

Pack, M. A.; Pohlman, J.; Ruppel, C. D.; Xu, X.

2012-12-01

Aerobic methane oxidation limits atmospheric methane emissions from degraded subsea permafrost and dissociated methane hydrates in high latitude oceans. Methane oxidation rate measurements are a crucial tool for investigating the efficacy of this process, but are logistically challenging when working on small research vessels in remote settings. We modified a low-level 14C-CH4 oxidation rate measurement for use in the Beaufort Sea above hydrate bearing sediments during August 2012. Application of the more common 3H-CH4 rate measurement that uses 106 times more radioactivity was not practical because the R/V Ukpik cannot accommodate a radiation van. The low-level 14C measurement does not require a radiation van, but careful isolation of the 14C-label is essential to avoid contaminating natural abundance 14C measurements. We used 14C-CH4 with a total activity of 1.1 μCi, which is far below the 100 μCi permitting level. In addition, we modified field procedures to simplify and shorten sample processing. The original low-level 14C-CH4 method requires 6 steps in the field: (1) collect water samples in glass serum bottles, (2) inject 14C-CH4 into bottles, (3) incubate for 24 hours, (4) filter to separate the methanotrophic bacterial cells from the aqueous sample, (5) kill the filtrate with sodium hydroxide (NaOH), and (6) purge with nitrogen to remove unused 14C-CH4. Onshore, the 14C-CH4 respired to carbon dioxide or incorporated into cell material by methanotrophic bacteria during incubation is quantified by accelerator mass spectrometry (AMS). We conducted an experiment to test the possibility of storing samples for purging and filtering back onshore (steps 4 and 6). We subjected a series of water samples to steps 1-3 & 5, and preserved with mercuric chloride (HgCl2) instead of NaOH because HgCl2 is less likely to break down cell material during storage. The 14C-content of the carbon dioxide in samples preserved with HgCl2 and stored for up to 2 weeks was stable, showing that oxidation of 14C-CH4 did not occur during storage. On the other hand, the 14C-content of the cell material decreased during storage, and the total carbon content of the filtered biomass decreased by 20% in the first 3 days. These results show that: [1] step 6 can be performed onshore because HgCl2 is an effective preservative even when unused 14C-CH4 is not removed, and [2] cell material is not stable in HgCl2 preserved samples, so filtering (step 4) must take place in the field. Next, we attempted to simplify the field filtering procedure by incubating samples in plastic syringes, and then filtering through a luer lock filter holder connected to the syringe. However, syringe incubated samples yielded oxidation rates up to 27 times slower than those in glass bottles, and demonstrated that plastic syringes are not suitable for incubating samples. Thus, we devised a technique to filter directly from sample bottles, through a syringe filter holder, and into an evacuated glass serum bottle in one step. Overall, we were able to simplify the field protocol to work on small vessels in remote field settings without compromising data quality.

Trends in serum relaxin concentration among elite collegiate female athletes

PubMed Central

Dragoo, Jason L; Castillo, Tiffany N; Korotkova, Tatiana A; Kennedy, Ashleigh C; Kim, Hyeon Joo; Stewart, Dennis R

2011-01-01

Purpose: This study was designed to investigate the relationship between serum relaxin concentration (SRC) and menstrual history and hormonal contraceptive use among elite collegiate female athletes. Evaluation of SRC in athletes is necessary, because relaxin has been associated with increased knee joint laxity and decreased anterior cruciate ligament (ACL) strength in animal models. Methods: National Collegiate Athletic Association Division I female athletes participating in sports at high risk for ACL tears – basketball, field hockey, gymnastics, lacrosse, soccer, and volleyball – were invited to participate. All participants completed a questionnaire about their menstrual history and hormonal contraceptive use. Venipuncture was performed to obtain samples of serum progesterone and relaxin. Samples were obtained during the mid-luteal phase from ovulating participants, and between the actual or projected cycle days 21 to 24, from anovulatory participants. Serum concentration of relaxin and progesterone was determined by ELISA and the data were analyzed using SPSS statistical software with significance set at P = 0.05. Results: 169 female athletes participated. The mean SRC among all participants was 3.08 ± 6.66 pg/mL). The mean SRC differed significantly between those participants using hormonal contraceptives (1.41 pg/mL) and those not using hormonal contraceptives (3.08 pg/mL, P = 0.002). Mean SRC was lowest among amenorrheic participants (1.02 pg/mL) and highest among oligomenorrheic participants (3.71 pg/mL) and eumenorrheic participants (3.06 pg/mL); these differences were not significant (P = 0.53). Mean serum progesterone concentration (SPC) differed significantly between those participants using hormonal contraceptives (2.80 ng/mL), and those not using hormonal contraceptives (6.99 ng/mL, P < 0.0001). Conclusions: There is a positive correlation between serum progesterone and SRC and an attenuation of SRC with hormonal contraceptive use. Our results underscore the significant role that hormonal contraceptives can play in decreasing relaxin levels, if future investigations establish a link between relaxin levels and ligamentous injury among female athletes. PMID:21339934

Determination of antifungal activities in serum samples from mice treated with different antifungal drugs allows detection of an active metabolite of itraconazole.

PubMed

Maki, Katsuyuki; Watabe, Etsuko; Iguchi, Yumi; Nakamura, Hideko; Tomishima, Masaki; Ohki, Hidenori; Yamada, Akira; Matsumoto, Satoru; Ikeda, Fumiaki; Tawara, Shuichi; Mutoh, Seitaro

2006-01-01

To establish an in vitro method of predicting in vivo efficacy of antifungal drugs against Candida albicans and Aspergillus fumigatus, the antifungal activities of fluconazole, itraconazole, and amphotericin B were determined in mouse serum. The minimum inhibitory concentration (MIC) of each drug was measured using mouse serum as a diluent. For C. albicans, the assay endpoint of azoles was defined as inhibition of mycelial extension (mMIC) and for A. fumigatus, as no growth (MIC). The MICs of amphotericin B for both pathogens were defined as the MIC at which no mycelial growth occurred. Serum MIC or mMIC determinations were then used to estimate the concentration of the drugs in serum of mice treated with antifungal drugs by multiplying the antifungal titer of the serum samples by the serum (m)MIC. The serum drug concentrations were also determined by HPLC. The serum concentrations estimated microbiologically showed good agreement with those determined by HPLC, except for itraconazole. Analysis of the serum samples from itraconazole-treated mice by a sensitive bioautography revealed the presence of additional spots, not seen in control samples of itraconazole. The bioautography assay demonstrated that the additional material detected in serum from mice treated with itraconazole was an active metabolite of itraconazole. The data showed that the apparent reduction in the itraconazole serum concentration as determined by HPLC was the result of the formation of an active metabolite, and that the use of a microbiological method to measure serum concentrations of drugs can provide a method for prediction of in vivo efficacy of antifungal drugs.

Evaluation of the use of pooled serum, pooled muscle tissue fluid (meat juice) and pooled faeces for monitoring pig herds for Salmonella.

PubMed

Davies, R H; Heath, P J; Coxon, S M; Sayers, A R

2003-01-01

Monitoring for Salmonella in slaughter pigs is important to enable targeted control measures to be applied on problem farms and at the abattoir. The aim of this study was to determine whether pooled serum and meat juice could be used to identify finishing pig herds with a high prevalence of infection. Samples of meat juice, serum, caecal contents, carcase swabs and pooled faeces from pig pens were taken from 20 commercial pig finishing farms and comparisons were made between the results of Salmonella culture, individual ELISA tests on serum and meat juice and pooled samples of serum and meat juice. Salmonella was isolated from samples from 19 of 20 farms. None of the ELISA tests showed a statistically significant correlation with caecal carriage of Salmonella or contamination of carcases. Serum mean optical density (O.D.) from pools of five, 10 or 20 sera showed a significant correlation with the Salmonella status of farm pen faeces. All pooled serum O.D. and sample/positive control ratio results correlated significantly with the results of the conventional individual sample ELISA. There was a statistically significant correlation between the incidence of Salmonella in farm pen pooled faeces and the prevalence of Salmonella in caeca of slaughter pigs. The results show a generally poor correlation between serological and bacteriological results but pooled serum or meat juice samples could be used as a cheaper substitute for serological screening of farms for Salmonella than individual samples. The availability of a cheaper test should allow the costs of Salmonella monitoring of pig farms to be reduced or allow more regular testing to enhance the designation of farm Salmonella risk status.

Detection of Serum microRNAs From Department of Defense Serum Repository

PubMed Central

Woeller, Collynn F.; Thatcher, Thomas H.; Van Twisk, Daniel; Pollock, Stephen J.; Croasdell, Amanda; Kim, Nina; Hopke, Philip K.; Xia, Xiaoyan; Thakar, Juilee; Mallon, COL Timothy M.; Utell, Mark J.; Phipps, Richard P.

2017-01-01

Objective The aim of this study was to investigate whether serum samples from the Department of Defense Serum Repository (DoDSR) are of sufficient quality to detect microRNAs (miRNAs), cytokines, immunoglobulin E (IgE), and polycyclic aromatic hydrocarbons (PAHs). Methods MiRNAs were isolated and quantified by polymerase chain reaction (PCR) array. Cytokines and chemokines related to inflammation were measured using multiplex immunoassays. Cotinine and IgE were detected by enzyme-linked immunoassay (ELISA) and PAHs were detected by Liquid Chromatography/Mass Spectroscopy. Results We detected miRNAs, cytokines, IgE, and PAHs with high sensitivity. Eleven of 30 samples tested positive for cotinine suggesting tobacco exposure. Significant associations between serum cotinine, cytokine, IgE, PAHs, and miRNA were discovered. Conclusion We successfully quantified over 200 potential biomarkers of occupational exposure from DoDSR samples. The stored serum samples were not affected by hemolysis and represent a powerful tool for biomarker discovery and analysis in retrospective studies. PMID:27501106

Chemometric techniques on the analysis of Raman spectra of serum blood samples of breast cancer patients

NASA Astrophysics Data System (ADS)

Rocha-Osornio, L. N.; Pichardo-Molina, J. L.; Barbosa-Garcia, O.; Frausto-Reyes, C.; Araujo-Andrade, C.; Huerta-Franco, R.; Gutiérrez-Juárez, G.

2008-02-01

Raman spectroscopy and Multivariate methods were used to study serum blood samples of control and breast cancer patients. Blood samples were obtained from 11 patients and 12 controls from the central region of Mexico. Our results show that principal component analysis is able to discriminate serum sample of breast cancer patients from those of control group, also the loading vectors of PCA plotted as a function of Raman shift shown which bands permitted to make the maximum discrimination between both groups of samples.

Mycobacterium paratuberculosis detection in cow's milk in Argentina by immunomagnetic separation-PCR.

PubMed

Gilardoni, Liliana Rosa; Fernández, Bárbara; Morsella, Claudia; Mendez, Laura; Jar, Ana María; Paolicchi, Fernando Alberto; Mundo, Silvia Leonor

2016-01-01

The aim of this study was to standardize a diagnosis procedure to detect Mycobacterium avium subsp. paratuberculosis (Map) DNA in raw cow milk samples under field conditions. A procedure that combines both immunomagnetic separation and IS900-PCR detection (IMS-IS1 PCR) was employed on milk samples from 265 lactating Holstein cows from Map infected and uninfected herds in Argentina. IMS-IS1 PCR results were analyzed and compared with those obtained from milk and fecal culture and serum ELISA. The extent of agreement between both tests was determined by the Kappa test. IMS-IS1 PCR showed a detection limit of 10(1) CFU of Map/mL of milk, when 50:50 mix of monoclonal and polyclonal antibodies were used to coat magnetic beads. All of the 118 samples from the Map uninfected herds were negative for the set of the tests. In Map infected herds, 80 out of 147 cows tested positive by milk IMS-IS1 PCR (55%), of which 2 (1.4%) were also positive by milk culture, 15 (10%) by fecal culture, and 20 (14%) by serum ELISA. Kappa statistics (95% CI) showed a slight agreement between the different tests (<0.20), and the proportions of agreement were ≤0.55. The IMS-IS1 PCR method detected Map in milk of the cows that were not positive in other techniques. This is the first report dealing with the application of IMS-IS1 PCR in the detection of Map in raw milk samples under field conditions in Argentina. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Systematic Evaluation of the Use of Human Plasma and Serum for Mass-Spectrometry-Based Shotgun Proteomics.

PubMed

Lan, Jiayi; Núñez Galindo, Antonio; Doecke, James; Fowler, Christopher; Martins, Ralph N; Rainey-Smith, Stephanie R; Cominetti, Ornella; Dayon, Loïc

2018-04-06

Over the last two decades, EDTA-plasma has been used as the preferred sample matrix for human blood proteomic profiling. Serum has also been employed widely. Only a few studies have assessed the difference and relevance of the proteome profiles obtained from plasma samples, such as EDTA-plasma or lithium-heparin-plasma, and serum. A more complete evaluation of the use of EDTA-plasma, heparin-plasma, and serum would greatly expand the comprehensiveness of shotgun proteomics of blood samples. In this study, we evaluated the use of heparin-plasma with respect to EDTA-plasma and serum to profile blood proteomes using a scalable automated proteomic pipeline (ASAP 2 ). The use of plasma and serum for mass-spectrometry-based shotgun proteomics was first tested with commercial pooled samples. The proteome coverage consistency and the quantitative performance were compared. Furthermore, protein measurements in EDTA-plasma and heparin-plasma samples were comparatively studied using matched sample pairs from 20 individuals from the Australian Imaging, Biomarkers and Lifestyle (AIBL) Study. We identified 442 proteins in common between EDTA-plasma and heparin-plasma samples. Overall agreement of the relative protein quantification between the sample pairs demonstrated that shotgun proteomics using workflows such as the ASAP 2 is suitable in analyzing heparin-plasma and that such sample type may be considered in large-scale clinical research studies. Moreover, the partial proteome coverage overlaps (e.g., ∼70%) showed that measures from heparin-plasma could be complementary to those obtained from EDTA-plasma.

Quantitation of 25-hydroxyvitamin D in dried blood spots by 2D LC-MS/MS without derivatization and correlation with serum in adult and pediatric studies.

PubMed

Jensen, Berit P; Saraf, Rajneeta; Ma, Jing; Berry, Sarah; Grant, Cameron C; Camargo, Carlos A; Sies, Christiaan W

2018-06-01

Demand for measurement of 25-hydroxyvitamin D (25OHD) is growing and dried blood spot (DBS) sampling is attractive as samples are easier to collect, transport and store. A 2D LC-MS/MS assay without derivatization was developed. DBS punches (3.2 mm) were ultrasonicated with d 6 -25OHD 3 in 70% methanol followed by hexane extraction, dry-down and reconstitution. The assay was validated and applied to two studies comparing whole blood adult DBS with serum samples (n = 40) and neonatal whole blood DBS with cord serum samples (n = 80). The assay was validated in whole blood DBS over the range 13-106 nmol/L 25OHD 3 and 11-91 nmol/L 25OHD 2 with a limit of detection of 3 nmol/L. Intra- and inter-day imprecision was <13% CV and bias <12%. The assay had high recovery and minimal matrix effects. Triplicate DBS study samples had a mean CV of ≤13% for 25OHD 3. No 25OHD 2 was detected. DBS calculated serum 25OHD 3 concentrations correlated strongly with serum concentrations in the adult DBS/serum study (r = 0.94) and moderately in the neonatal DBS/cord serum study (r = 0.69). Direct quantitation of 25OHD in DBS by 2D LC-MS/MS without derivatization was found to be an alternative to serum quantitation applicable to clinical research studies on adult DBS samples. Copyright © 2018 Elsevier B.V. All rights reserved.

Evaluation of a commercial rubella IgM assay for use on oral fluid samples for diagnosis and surveillance of congenital rubella syndrome and postnatal rubella.

PubMed

Vijaylakshmi, P; Muthukkaruppan, V R; Rajasundari, A; Korukluoglu, G; Nigatu, W; L A Warrener; Samuel, D; Brown, D W G

2006-12-01

Clinical diagnosis (surveillance) of rubella is unreliable and laboratory confirmation is essential. Detection of virus specific IgM in serum is the most commonly used method. However, the use of serum necessitates the drawing of blood, either through venipuncture or finger/heel prick, which can be difficult in young babies. Oral fluid samples have proved useful as an alternative, less invasive sample for virus specific IgM detection however until recently no commercial rubella IgM tests were available, restricting the usefulness of this approach. To evaluate the performance of the Microimmune Rubella IgM capture EIA using oral fluid samples from outbreaks as well as in cases of suspected congenital rubella syndrome (CRS). Paired serum and oral fluids were collected from cases during a rubella outbreak in three provinces in Turkey. Matched serum and oral fluid samples were collected from children with suspected CRS in an active surveillance programme at the Aravind Eye Hospital in South India. Serum samples were collected as part of the measles surveillance programme in Ethiopia. On serum samples the sensitivity and specificity of the Microimmune Rubella IgM capture EIA compared to Behring Enzygnost rubella IgM test was 96.9% (62/64; 95% CI 94.2-100%) and 100% (53/53; 95% CI 93.2-100%). On oral fluids compared to matched Behring results on serum the sensitivity was 95.5% (42/44; 95% CI 84.5-99.4%). The sensitivity and specificity of Microimmune Rubella IgM capture EIA on oral fluids from suspected CRS cases compared to serum results using Behring Enzygnost IgM assay was 100% (95% CI 84.5-100%) and 100% (95% CI 95.8-100.0%) respectively. Microimmune Rubella IgM capture EIA has adequate performance for diagnosis and surveillance of rubella in outbreak using either serum or oral fluid specimens.

Comparison of Bovine coronavirus-specific and Bovine respiratory syncytial virus-specific antibodies in serum versus milk samples detected by enzyme-linked immunosorbent assay.

PubMed

Ohlson, Anna; Blanco-Penedo, Isabel; Fall, Nils

2014-01-01

Bovine coronavirus (BCV; Betacoronavirus 1) and Bovine respiratory syncytial virus (BRSV) are significant causes of enteric and respiratory disease in beef and dairy cattle throughout the world. Indirect enzyme-linked immunosorbent assays are widely used to detect serum antibodies for herd monitoring and prevalence studies. In dairy herds, milk is more readily collected than serum. Hence, in order to investigate the test agreement between serum and milk, both serum and milk samples from 105 cows in 27 dairy herds were analyzed in parallel for presence of immunoglobulin G antibodies to BCV and BRSV. The Bland-Altman analyses of data demonstrated good agreement between serum and milk antibody titers for both viruses. The results indicate milk samples are sufficient for surveillance of antibodies to BCV and BRSV.

Comparison of serum serotonin and serum 5-HIAA LC-MS/MS assays in the diagnosis of serotonin producing neuroendocrine neoplasms: A pilot study.

PubMed

Lindström, Mikael; Tohmola, Niina; Renkonen, Risto; Hämäläinen, Esa; Schalin-Jäntti, Camilla; Itkonen, Outi

2018-07-01

Serotonin (5-hydroxytyramine) is a mediator of gastrointestinal smooth muscle contraction, and is secreted by neuroendocrine neoplasms (NENs). We developed a liquid chromatography tandem mass spectrometry (LC-MS/MS) assay for serum serotonin to be used in NEN diagnostics and follow-up. We used serum samples from healthy volunteers (n = 31) and patients suspected or monitored for NEN (n = 98). Serotonin-D 4 internal standard was added to samples before solid phase extraction (SPE) and quantification by LC-MS/MS. The effects of sample handling and preparation on serotonin stability were studied. Finally, we established a provisional reference range for serum serotonin and compared our assay with serum 5-hydroxyindoleacetic acid (5-HIAA) for detection of NENs. Our assay is sensitive and has a wide linear range (10-10,000 nmol/l). Serum serotonin is stable for 7 days at room temperature and for 3 months at -20 °C. Sampling temperature is not critical. Normal range for serum serotonin was 270-1490 nmol/l. We found that serum serotonin and 5-HIAA performed equally well as diagnostic tests for NENs. Our LC-MS/MS assay for serum serotonin is well suited for clinical research and patient diagnostics. Our results confirm that it can complement 5-HIAA in diagnosis of NENs. Copyright © 2018 Elsevier B.V. All rights reserved.

Occurrence of Salmonella serotype Typhimurium DT104 on a commercial swine farm before, during, and after depopulation and repopulation.

PubMed

Erdman, Matthew M; Harris, Isabel T; Torremorell, Montserrat; Wilt, Vincil M; Harris, D L Hank

2005-08-01

To determine whether depopulation-repopulation could be used to eradicate Salmonella serotype Typhimurium DT104 from a commercial swine farm in the midwestern United States. Observational study A commercial swine farm undergoing depopulation-repopulation to eliminate porcine reproductive and respiratory syndrome virus and Mycoplasma hyopneumoniae. Pooled fecal samples, tissue samples, and serum samples were collected from pigs on the farm before and after depopulation-repopulation. When there were no pigs on the farm, environmental swab specimens were collected for bacterial culture. Serum was analyzed for anti-Salmonella antibodies with an indirect ELISA. Salmonella isolates obtained by bacterial culture of fecal, tissue, and environmental samples were characterized by means of serotyping, phage typing, pulsed-field gel electrophoresis (PFGE), and antimicrobial susceptibility testing. 167 Salmonella isolates representing 9 serotypes were recovered from the farm. Results of PFGE and antimicrobial susceptibility testing suggested that S. Typhimurium DT104 strain was not eradicated from the farm. However, seroprevalence of anti-Salmonella antibodies and the percentage of pooled fecal samples positive for Salmonella spp were significantly decreased following repopulation. Results suggested that depopulation-repopulation in conjunction with stringent cleaning and disinfection, attention to biosecurity procedures, control of other diseases, and changes in feed management may reduce the occurrence of, but likely will not eliminate, Salmonella spp in commercial swine herds.

Innovative approaches to vitamin A assessment.

PubMed

Craft, N E

2001-05-01

The health and sight of millions of children are compromised each year as a consequence of vitamin A (VA) deficiency. Serum retinol is the most commonly used indicator of VA status. Unfortunately, its use is impractical for national surveys because it involves collection of venous blood, centrifugation and frozen storage before analysis. To make VA assessment more practical, we have developed approaches incorporating dried blood spots (DBS) or portable instrumentation. DBS have been used as a sample matrix to screen neonates for many biochemical compounds. Until recently, it was not thought that VA was stable in DBS. However, we demonstrated that the measure of DBS retinol correlates well with serum retinol in both healthy adults (r(2) = 0.88-0.90) and compromised populations (r(2) = 0.73-0.84). Compared with serum retinol, the sensitivity and specificity of detecting VA deficiency by DBS retinol range from 73 to 93% and from 90 to 100%, respectively. Although few data are available, retinol binding protein (RBP) can also be measured in DBS. RBP has been used as a surrogate marker for serum retinol. Correlations coefficients (r(2)) between serum RBP and serum retinol range from 0.4 to 0.8. In addition, work has been done to develop portable instrumentation to measure VA status in the field. A fluorometer has been optimized for VA fluorescence and is linear into the deficient range for the direct fluorimetric measurement of serum holo-RBP. Progress is being made to use the instrument to directly measure holo-RBP in a drop of whole blood.

Simultaneous Detection of Antibodies against Apx Toxins ApxI, ApxII, ApxIII, and ApxIV in Pigs with Known and Unknown Actinobacillus pleuropneumoniae Exposure Using a Multiplexing Liquid Array Platform

PubMed Central

Giménez-Lirola, Luis G.; Jiang, Yong-Hou; Sun, Dong; Hoang, Hai; Yoon, Kyoung-Jin; Halbur, Patrick G.

2014-01-01

Surveillance for the presence of Actinobacillus pleuropneumoniae infection in a population plays a central role in controlling the disease. In this study, a 4-plex fluorescent microbead-based immunoassay (FMIA), developed for the simultaneous detection of IgG antibodies to repeat-in-toxin (RTX) toxins (ApxI, ApxII, ApxIII, and ApxIV) of A. pleuropneumoniae, was evaluated using (i) blood serum samples from pigs experimentally infected with each of the 15 known A. pleuropneumoniae serovars or with Actinobacillus suis, (ii) blood serum samples from pigs vaccinated with a bacterin containing A. pleuropneumoniae serovar 1, 3, 5, or 7, and (iii) blood serum samples from pigs with an unknown A. pleuropneumoniae exposure status. The results were compared to those obtained in a previous study where a dual-plate complement fixation test (CFT) and three commercially available enzyme-linked immunosorbent assays (ELISAs) were conducted on the same sample set. On samples from experimentally infected pigs, the 4-plex Apx FMIA detected specific seroconversion to Apx toxins as early as 7 days postinfection in a total of 29 pigs inoculated with 14 of the 15 A. pleuropneumoniae serovars. Seroconversion to ApxII and ApxIII was detected by FMIA in pigs inoculated with A. suis. The vaccinated pigs showed poor humoral responses against ApxI, ApxII, ApxIII, and ApxIV. In the field samples, the humoral response to ApxIV and the A. pleuropneumoniae seroprevalence increased with age. This novel FMIA (with a sensitivity of 82.7% and a specificity of 100% for the anti-ApxIV antibody) was found to be more sensitive and accurate than current tests (sensitivities, 9.5 to 56%; specificity, 100%) and is potentially an improved tool for the surveillance of disease and for monitoring vaccination compliance. PMID:24226091

Comparative evaluation of serum, FTA filter-dried blood and oral fluid as sample material for PRRSV diagnostics by RT-qPCR in a small-scale experimental study.

PubMed

Steinrigl, Adolf; Revilla-Fernández, Sandra; Wodak, Eveline; Schmoll, Friedrich; Sattler, Tatjana

2014-01-01

Recently, research into alternative sample materials, such as oral fluid or filter-dried blood has been intensified, in order to facilitate cost-effective and animal-friendly sampling of individuals or groups of pigs for diagnostic purposes. The objective of this study was to compare the sensitivity of porcine reproductive and respiratory syndrome virus (PRRSV)-RNA detection by reverse transcription quantitative real-time PCR (RT-qPCR) in serum, FTA filter-dried blood and oral fluid sampled from individual pigs. Ten PRRSV negative pigs were injected with an EU-type PRRSV live vaccine. Blood and oral fluid samples were taken from each pig before, and 4, 7, 14 and 21 days after vaccination. All samples were then analyzed by PRRSV RT-qPCR. In serum, eight often pigs tested RT-qPCR positive at different time points post infection. Absolute quantification showed low serum PRRSV-RNA loads in most samples. In comparison to serum, sensitivity of PRRSV-RNA detection was strongly reduced in matched FTA filter-dried blood and in oral fluid from the same pigs. These results indicate that with low PRRSV-RNA loads the diagnostic sensitivity of PRRSV-RNA detection by RT-qPCR achieved with serum is currently unmatched by either FTA filter-dried blood or oral fluid.

Quantitative bioanalysis of strontium in human serum by inductively coupled plasma-mass spectrometry

PubMed Central

Somarouthu, Srikanth; Ohh, Jayoung; Shaked, Jonathan; Cunico, Robert L; Yakatan, Gerald; Corritori, Suzana; Tami, Joe; Foehr, Erik D

2015-01-01

Aim: A bioanalytical method using inductively-coupled plasma-mass spectrometry to measure endogenous levels of strontium in human serum was developed and validated. Results & methodology: This article details the experimental procedures used for the method development and validation thus demonstrating the application of the inductively-coupled plasma-mass spectrometry method for quantification of strontium in human serum samples. The assay was validated for specificity, linearity, accuracy, precision, recovery and stability. Significant endogenous levels of strontium are present in human serum samples ranging from 19 to 96 ng/ml with a mean of 34.6 ± 15.2 ng/ml (SD). Discussion & conclusion: Calibration procedures and sample pretreatment were simplified for high throughput analysis. The validation demonstrates that the method was sensitive, selective for quantification of strontium (88Sr) and is suitable for routine clinical testing of strontium in human serum samples. PMID:28031925

Evaluation of a portable automated serum chemistry analyzer for field assessment of harlequin ducks, Histrionicus histrionicus

USGS Publications Warehouse

Stoskopf, Michael K.; Mulcahy, Daniel M.; Esler, Daniel N.

2010-01-01

A portable analytical chemistry analyzer was used to make field assessments of wild harlequin ducks (Histrionicus histrionicus) in association with telemetry studies of winter survival in Prince William Sound, Alaska. We compared serum chemistry results obtained on-site with results from a traditional laboratory. Particular attention was paid to serum glucose and potassium concentrations as potential indicators of high-risk surgical candidates based on evaluation of the field data. The median differential for glucose values ( = 8 2) between methods was 0.6 mmol/L (quartiles 0.3 and 0.9 mmol/L) with the median value higher when assayed on site. Analysis of potassium on site returned a median of 2.7 mmol/L ( = 8 8 ; quartiles 2.4 and 3.0 mmol/L). Serum potassium values were too low for quantitation by the traditional laboratory. Changes in several serum chemistry values following a three-day storm during the study support the value of on site evaluation of serum potassium to identify presurgical patients with increased anesthetic risk.

Continuous improvement of medical test reliability using reference methods and matrix-corrected target values in proficiency testing schemes: application to glucose assay.

PubMed

Delatour, Vincent; Lalere, Beatrice; Saint-Albin, Karène; Peignaux, Maryline; Hattchouel, Jean-Marc; Dumont, Gilles; De Graeve, Jacques; Vaslin-Reimann, Sophie; Gillery, Philippe

2012-11-20

The reliability of biological tests is a major issue for patient care in terms of public health that involves high economic stakes. Reference methods, as well as regular external quality assessment schemes (EQAS), are needed to monitor the analytical performance of field methods. However, control material commutability is a major concern to assess method accuracy. To overcome material non-commutability, we investigated the possibility of using lyophilized serum samples together with a limited number of frozen serum samples to assign matrix-corrected target values, taking the example of glucose assays. Trueness of the current glucose assays was first measured against a primary reference method by using human frozen sera. Methods using hexokinase and glucose oxidase with spectroreflectometric detection proved very accurate, with bias ranging between -2.2% and +2.3%. Bias of methods using glucose oxidase with spectrophotometric detection was +4.5%. Matrix-related bias of the lyophilized materials was then determined and ranged from +2.5% to -14.4%. Matrix-corrected target values were assigned and used to assess trueness of 22 sub-peer groups. We demonstrated that matrix-corrected target values can be a valuable tool to assess field method accuracy in large scale surveys where commutable materials are not available in sufficient amount with acceptable costs. Copyright © 2012 Elsevier B.V. All rights reserved.

A sensitive one-step method for quantitative detection of α-amylase in serum and urine using a personal glucose meter.

PubMed

Wang, Qing; Wang, Hui; Yang, Xiaohai; Wang, Kemin; Liu, Rongjuan; Li, Qing; Ou, Jinqing

2015-02-21

Assays of α-amylase (AMS) activity in serum and urine constitute the important indicator for the diagnosis of acute pancreatitis, mumps, renal disease and abdominal disorders. Since these diseases confer a heavy financial burden on the health care system, AMS detection in point-of-care is fundamental. Here, a one-step assay for direct determination of the AMS activity was developed using a portable personal glucose meter (PGM). In this assay, maltopentaose was used as a substrate for sensitive detection of AMS with the assistance of α-glucosidase. In the presence of AMS, maltopentaose can be readily hydrolyzed to form maltotriose and maltose quickly. With the enzymatic hydrolysis of α-glucosidase, maltotriose and maltose can be turned into glucose rapidly, which can be quantitatively measured using a portable PGM. This assay did not require any cumbersome and time consuming operations, such as surface modification, synthesis of invertase conjugate, washing and centrifugation. Detection of AMS can be achieved using only a one-step mixture, and the limit of detection was 20 U L(-1) which was lower than the clinical cutoff for AMS. More importantly, this sensitive and selective assay was also used for the detection of AMS in human serum/urine samples. The results showed that the recovery of AMS from human serum/urine samples was 91-107%. The rapid and easy-to-operate assay may have potential application in the fields of point-of-care (POC) clinical diagnosis, particularly in rural and remote areas where lab equipment may be limited.

Feasibility study of feces for noninvasive biomonitoring of brominated flame retardants in toddlers.

PubMed

Sahlström, Leena M O; Sellström, Ulla; de Wit, Cynthia A; Lignell, Sanna; Darnerud, Per Ola

2015-01-06

This study investigated the feasibility of using feces as a noninvasive matrix to estimate serum concentrations of brominated flame retardants (BFRs) in toddlers for biomonitoring purposes. Tri- to decabrominated diphenyl ethers (tri-decaBDEs), isomer-specific hexabromocyclododecanes, and 16 emerging BFRs were determined in feces from 22 toddlers (11-15 months of age), and results were compared to previously analyzed matched serum samples. BDE-47, -153, -196, -197, -203, -206, -207, -208, and -209 were detected in the feces creating a matched data set (feces-serum, n = 21). Tetra-octaBDE concentrations were significantly higher (Student's paired comparisons t test, α = 0.05) in serum versus feces with BDE-153 having the highest mean difference between the sample matrices. BDE-209 was found in significantly higher concentrations in feces compared to serum. Significant correlations (Pearson's, α = 0.05) between congener-specific concentrations in feces and serum were found for all BDEs except BDE-197 and -203. The feces-serum associations found can be used to estimate serum concentrations of tetra-decaBDEs from feces concentrations and enable a noninvasive sampling method for biomonitoring BDEs in toddlers.

Effective surveillance for early classical swine fever virus detection will utilize both virus and antibody detection capabilities.

PubMed

Panyasing, Yaowalak; Kedkovid, Roongtham; Thanawongnuwech, Roongroje; Kittawornrat, Apisit; Ji, Ju; Giménez-Lirola, Luis; Zimmerman, Jeffrey

2018-03-01

Early recognition and rapid elimination of infected animals is key to controlling incursions of classical swine fever virus (CSFV). In this study, the diagnostic characteristics of 10 CSFV assays were evaluated using individual serum (n = 601) and/or oral fluid (n = 1417) samples collected from -14 to 28 days post inoculation (DPI). Serum samples were assayed by virus isolation (VI), 2 commercial antigen-capture enzyme-linked immunosorbent assays (ELISA), virus neutralization (VN), and 3 antibody ELISAs. Both serum and oral fluid samples were tested with 3 commercial real-time reverse transcription-polymerase chain reaction (rRT-PCR) assays. One or more serum samples was positive by VI from DPIs 3 to 21 and by antigen-capture ELISAs from DPIs 6 to 17. VN-positive serum samples were observed at DPIs ≥ 7 and by antibody ELISAs at DPIs ≥ 10. CSFV RNA was detected in serum samples from DPIs 2 to 28 and in oral fluid samples from DPIs 4 to 28. Significant differences in assay performance were detected, but most importantly, no single combination of sample and assay was able to dependably identify CSFV-inoculated pigs throughout the 4-week course of the study. The results show that effective surveillance for CSFV, especially low virulence strains, will require the use of PCR-based assays for the detection of early infections (<14 days) and antibody-based assays, thereafter. Copyright © 2018 Elsevier B.V. All rights reserved.

Preparation of serum and plasma samples for determination of tricyclic antidepressants: effects of blood collection tubes and storage.

PubMed

Nyberg, G; Mårtensson, E

1986-01-01

The effects were tested of eight common types of blood collection tubes and two types of "plasma separators" on the stability of the tricyclic antidepressants amitriptyline, imipramine, clomipramine, and their monodemethylated metabolites in venous blood samples. Although EDTA-containing Venoject lavender and Vacutainer lavender tubes seemed to give the most stable plasma samples, and Venoject red the most stable serum samples, the differences were too small to have practical consequences. Vacutainer royal blue collection tubes gave significant losses of greater than 20% of some of the substances. The tubes with serum separator gel or filter proved unsuitable, since they were responsible for losses of greater than 40%. The losses were not caused by redistribution between blood cells and plasma but occurred mainly as a result of contact between the contents and the caps of the tubes. Experiments with freezing, thawing, and storage of samples showed that freshly sampled blood could be stored at room temperature for 24 h in Venoject green tubes without significant losses. Serum samples could be stored at refrigerator temperature for 4 weeks without important losses. Freezing, thawing, and storage at -20 degrees C did not influence the serum or plasma concentrations.

Transplacental transfer of polycyclic aromatic hydrocarbons in paired samples of maternal serum, umbilical cord serum, and placenta in Shanghai, China.

PubMed

Zhang, Xiaolan; Li, Xiaojing; Jing, Ye; Fang, Xiangming; Zhang, Xinyu; Lei, Bingli; Yu, Yingxin

2017-03-01

Prenatal exposure to polycyclic aromatic hydrocarbons (PAHs) is a high-priority public health concern. However, maternal to fetal transplacental transfer of PAHs has not been systematically studied. To investigate the transplacental transfer of PAHs from mother to fetus and determine the influence of lipophilicity (octanol-water partition coefficient, K OW ) on transfer process, in the present study, we measured the concentrations of 15 PAHs in 95 paired maternal and umbilical cord serum, and placenta samples (in total 285 samples) collected in Shanghai, China. The average concentration of total PAHs was the highest in maternal serums (1290 ng g -1 lipid), followed by umbilical cord serums (1150 ng g -1 lipid). The value was the lowest in placenta samples (673 ng g -1 lipid). Low molecular weight PAHs were the predominant compounds in the three matrices. Increases in fish and meat consumption did not lead to increases in maternal PAH levels, and no obvious gender differences in umbilical cord serums were observed. The widespread presence of PAHs in umbilical cord serums indicated the occurrence of transplacental transfer. The ratios of PAH concentrations in umbilical cord serum to those in maternal serum (F/M) and the concentrations in placenta to those in maternal serum (P/M) of paired samples were analyzed to characterize the transfer process of individual PAHs. Most F/M ratios on lipid basis were close to one (range: 0.79 to 1.36), which suggested that passive diffusion may control the transplacental transfer of PAHs from maternal serum to the fetal circulation. The P/M and F/M values calculated on lipid basis showed that PAHs with lower K OW were more likely to transfer from mother to fetus via the placenta. Copyright © 2016 Elsevier Ltd. All rights reserved.

Dried venous blood samples for the detection and quantification of measles IgG using a commercial enzyme immunoassay.

PubMed Central

Riddell, Michaela A.; Byrnes, Graham B.; Leydon, Jennie A.; Kelly, Heath A.

2003-01-01

OBJECTIVES: To determine whether samples of dried venous blood (DVB) were an acceptable alternative to serum for detecting measles-specific IgG in a commercial enzyme immunoassay. METHODS: Paired samples of serum and DVB were collected from 98 suspected cases of measles and 1153 schoolchildren in Victoria, Australia. All samples were tested using the Dade Behring Enzygnost Anti-Measles-Virus/IgG immunoassay. DVB samples were eluted using either the sample buffer provided with the kit or 5% dry milk powder in phosphate-buffered saline-Tween 20. FINDINGS: DVB samples eluted by sample buffer showed significantly better linear correlation to the serum samples than did DVB samples eluted in 5% dry milk in phosphate-buffered saline-Tween 20. To improve the comparability of serum and DVB samples an adjustment factor of 1.28 was applied to the optical density (OD) values of DVB. This adjustment also enabled quantification of the titre of measles IgG in mIU/ml directly from the OD value using the alpha calculation as specified by the kit protocol. For DVB samples stored for less than six months at 4 degrees C, the assay showed an overall sensitivity of 98.4% and a specificity of 97.2% compared with the results of serum testing. CONCLUSION: These results illustrate the potential for DVB samples to be widely used with the Dade Behring enzyme immunoassay system for determining the immunity of the individual and the population to the measles virus. PMID:14758429

Organohalogenated contaminants (OHCs) in the serum and hair of pet cats and dogs: biosentinels of indoor pollution.

PubMed

Ali, Nadeem; Malik, Riffat Naseem; Mehdi, Toufeer; Eqani, Syed Ali Musstjab Akber Shah; Javeed, Aqeel; Neels, Hugo; Covaci, Adrian

2013-04-01

Concentrations of different classes of organohalogenated contaminants (OHCs) viz., polybrominated diphenyl ethers (PBDEs), novel brominated flame retardants (NBFRs), bromophenols (BPs), polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs) and their metabolites were determined in cat and dog serum and hair samples from Pakistan. The major DDT metabolite, p,p'-DDE, was the major OHC in cat serum (N=20) and ranged between 1 and 2150 ng/g lipid weight (lw). p,p'-DDE was not detected in dog serum (N=16). In contrary to other OHCs, levels of ∑HO-PCBs were significantly higher in dog serum (median=6.0 ng/g lw) than cat serum (median=2.2 ng/g lw). Levels of most OHCs were significantly higher (p<0.05) in cat serum than those found in human serum from the same region, in particular for ∑PBDEs (ranged 1-1280 ng/g lw). Significantly lower levels of OCPs (p<0.05) were detected in dog serum than in human serum. The concentrations of ∑BPs were seven times higher in cat serum (median 112 ng/g lw) than dog serum (median 16 ng/g lw). To the best of our knowledge, this is the first time that NBFRs, e.g. 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE), decabromodiphenylethane (DBDPE), and bis(2-ethylhexyl)-3,4,5,6-tetrabromophthalate (TBPH), were detected in cat and dog's hair. BTBPE had the highest detection frequency (30%) in the serum samples. In cat and dog hair samples, the order of importance of OHCs was ∑OCPs>∑NBFRs>∑PBDEs>∑PCB, with the highest concentrations being around 38 ng/g hair. In paired hair-serum cat samples (N=12), ∑DDTs (r=0.65, p=0.001) were significantly correlated, while for all other OHCs no significant correlations (p<0.001) were observed in both cats and dogs. Our findings on both hair and serum samples suggested that pet dogs do not bioaccumulate DDTs. Our results are also in agreement with the hypothesis that pets may serve as biosentinels for indoor pollution. This is the first study to document the presence of OHCs in pets from Pakistan and provides baseline information for future monitoring of OHCs in pets. Copyright © 2013 Elsevier B.V. All rights reserved.

Sensitivity and Specificity of an Operon Immunochromatographic Test in Serum and Whole-Blood Samples for the Diagnosis of Trypanosoma cruzi Infection in Spain, an Area of Nonendemicity

PubMed Central

Flores-Chavez, María; Cruz, Israel; Nieto, Javier; Gárate, Teresa; Navarro, Miriam; Pérez-Ayala, Ana; López-Vélez, Rogelio

2012-01-01

Trypanosoma cruzi infection is an imported parasitic disease in Spain, and the majority of infected individuals are in the chronic phase of the disease. This study evaluated the sensitivity and specificity of the Operon immunochromatographic test (ICT-Operon; Simple Stick Chagas and Simple Chagas WB [whole blood]; Operon S.A., Spain) for different biological samples. Well-characterized serum samples were obtained from chagasic patients (n = 63), nonchagasic individuals (n = 95), visceral leishmaniasis patients (n = 38), and malaria patients (n = 55). Noncharacterized specimens were obtained from Latin American immigrants and individuals at risk with a clinical and/or epidemiological background: these specimens were recovered serum or plasma samples (n = 450), whole peripheral blood (n = 94), and capillary blood (n = 282). The concordance of the results by enzyme-linked immunosorbent assay and indirect immunofluorescence test was considered to be the “gold standard” for diagnosis. Serum and plasma samples were analyzed by Stick Chagas, and whole blood was analyzed by Simple Chagas WB. The sensitivity and specificity of the ICT-Operon in well-characterized samples were 100% and 97.9%, respectively. No cross-reactivity was found with samples obtained from visceral leishmaniasis patients. In contrast, a false-positive result was obtained in 27.3% of samples from malaria patients. The sensitivities of the rapid test in noncharacterized serum or plasma, peripheral blood, and capillary blood samples were 100%, 92.1%, and 86.4%, respectively, while the specificities were 91.6%, 93.6%, and 95% in each case. ICT-Operon showed variable sensitivity, depending on the kind of sample, performing better when serum or plasma samples were used. It could therefore be used for serological screening combined with any other conventional test. PMID:22761296

Pilot Metabolome-Wide Association Study of Benzo(a)pyrene in Serum from Military Personnel

PubMed Central

Walker, Douglas I.; Pennell, Kurt D.; Uppal, Karan; Xia, Xiaoyan; Hopke, Philip K.; Utell, Mark J.; Phipps, Richard P.; Sime, Patricia J.; Rohrbeck, Patricia; Mallon, COL Timothy M.; Jones, Dean P.

2016-01-01

Objective A pilot study was conducted to test the feasibility of using Department of Defense Serum Repository (DoDSR) samples to study health and exposure-related effects. Methods Thirty unidentified human serum samples were obtained from the DoDSR and analyzed for normal serum metabolites with high-resolution mass spectrometry and serum levels of free benzo(a)pyrene (BaP) by gas chromatography-mass spectrometry. Metabolic associations with BaP were determined using a metabolome wide association study (MWAS) and metabolic pathway enrichment. Results The serum analysis detected normal ranges of glucose, selected amino acids, fatty acids, and creatinine. Free BaP was detected in a broad concentration range. MWAS of BaP showed associations with lipids, fatty acids, and sulfur amino acid metabolic pathways. Conclusion The results show the DoDSR samples are of sufficient quality for chemical profiling of DoD personnel. PMID:27501104

Pilot Metabolome-Wide Association Study of Benzo(a)pyrene in Serum From Military Personnel.

PubMed

Walker, Douglas I; Pennell, Kurt D; Uppal, Karan; Xia, Xiaoyan; Hopke, Philip K; Utell, Mark J; Phipps, Richard P; Sime, Patricia J; Rohrbeck, Patricia; Mallon, Col Timothy M; Jones, Dean P

2016-08-01

A pilot study was conducted to test the feasibility of using Department of Defense Serum Repository (DoDSR) samples to study health and exposure-related effects. Thirty unidentified human serum samples were obtained from the DoDSR and analyzed for normal serum metabolites with high-resolution mass spectrometry and serum levels of free benzo(a)pyrene (BaP) by gas chromatography-mass spectrometry. Metabolic associations with BaP were determined using a metabolome-wide association study (MWAS) and metabolic pathway enrichment. The serum analysis detected normal ranges of glucose, selected amino acids, fatty acids, and creatinine. Free BaP was detected in a broad concentration range. MWAS of BaP showed associations with lipids, fatty acids, and sulfur amino acid metabolic pathways. The results show that the DoDSR samples are of sufficient quality for chemical profiling of DoD personnel.

Relationship of Salmonella infection and inflammatory intestinal response with hematological and serum biochemical values in laying hens.

PubMed

Soria, Mario Alberto; Bonnet, María Agustina; Bueno, Dante Javier

2015-06-15

There are few studies about the blood serum of laying hens infected with Salmonella. The differential leukocyte count and blood chemistry values are an important aid in the diagnosis of human diseases, but blood parameters in the avian species are not well known. On the other hand, invasive forms of bacterial gastroenteritis, like Salmonella, often cause intestinal inflammation so this study was undertaken to find a biomarker of Salmonella infection and inflammatory intestinal response in the hematological or serum biochemical parameters in laying hens. Furthermore, we evaluated the association of some farm characteristics with Salmonella infection and fecal leukocytes (FL). A fecal sample with at least one fecal leukocyte per field was considered positive for inflammatory intestinal response. False positive serum reactions for Salmonella infection, by serum plate agglutination (SPA) test, were reduced by heating the sample to 56°C for 30 min and then diluting it 5-fold. The range of hematological and biochemical parameter values was very wide, in addition, there was a poor agreement between the SPA and FL results. Comparison of the positive and negative samples in SPA and FL showed that 1.3% and 79.8% of the laying hens were positive and negative in both tests, respectively. Hens with a positive SPA result showed a higher percentage of monocytes than those with a negative SPA result. Hens with a positive FL test had a higher percentage of heterophils, ratio of heterophils to lymphocytes and aspartate aminotransferase values, while the percentage of lymphocytes was significantly lower (P < 0.05) than those with a negative FL test. The risk of Salmonella infection increased when the age of laying hens and the number of hens per poultry house was greater than or equal to 18 months old and 10,000 laying hens, compared to less than 18 months old and 10,000 laying hens, respectively. On the other hand, the risk of inflammatory intestinal response was higher in laying hens ≥ 18 months old than in hens < 18 months old. Despite the fact that we did not find any specific biomarker of Salmonella infection, this is the first report about the change of Salmonella infection and inflammatory response in hematological/serum biochemical values for laying hens. Copyright © 2015 Elsevier B.V. All rights reserved.

Diagnostic performance of a seven-marker serum protein biosignature for the diagnosis of active TB disease in African primary healthcare clinic attendees with signs and symptoms suggestive of TB.

PubMed

Chegou, Novel N; Sutherland, Jayne S; Malherbe, Stephanus; Crampin, Amelia C; Corstjens, Paul L A M; Geluk, Annemieke; Mayanja-Kizza, Harriet; Loxton, Andre G; van der Spuy, Gian; Stanley, Kim; Kotzé, Leigh A; van der Vyver, Marieta; Rosenkrands, Ida; Kidd, Martin; van Helden, Paul D; Dockrell, Hazel M; Ottenhoff, Tom H M; Kaufmann, Stefan H E; Walzl, Gerhard

2016-09-01

User-friendly, rapid, inexpensive yet accurate TB diagnostic tools are urgently needed at points of care in resource-limited settings. We investigated host biomarkers detected in serum samples obtained from adults with signs and symptoms suggestive of TB at primary healthcare clinics in five African countries (Malawi, Namibia, South Africa, The Gambia and Uganda), for the diagnosis of TB disease. We prospectively enrolled individuals presenting with symptoms warranting investigation for pulmonary TB, prior to assessment for TB disease. We evaluated 22 host protein biomarkers in stored serum samples using a multiplex cytokine platform. Using a pre-established diagnostic algorithm comprising of laboratory, clinical and radiological findings, participants were classified as either definite TB, probable TB, questionable TB status or non-pulmonary TB. Of the 716 participants enrolled, 185 were definite and 29 were probable TB cases, 6 had questionable TB disease status, whereas 487 had no evidence of TB. A seven-marker biosignature of C reactive protein, transthyretin, IFN-γ, complement factor H, apolipoprotein-A1, inducible protein 10 and serum amyloid A identified on a training sample set (n=491), diagnosed TB disease in the test set (n=210) with sensitivity of 93.8% (95% CI 84.0% to 98.0%), specificity of 73.3% (95% CI 65.2% to 80.1%), and positive and negative predictive values of 60.6% (95% CI 50.3% to 70.1%) and 96.4% (95% CI 90.5% to 98.8%), respectively, regardless of HIV infection status or study site. We have identified a seven-marker host serum protein biosignature for the diagnosis of TB disease irrespective of HIV infection status or ethnicity in Africa. These results hold promise for the development of a field-friendly point-of-care screening test for pulmonary TB. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

Mimivirus Circulation among Wild and Domestic Mammals, Amazon Region, Brazil

PubMed Central

Dornas, Fábio P.; Rodrigues, Felipe P.; Boratto, Paulo V.M.; Silva, Lorena C.F.; Ferreira, Paulo C.P.; Bonjardim, Cláudio A.; Trindade, Giliane S.; Kroon, Erna G.; La Scola, Bernard

2014-01-01

To investigate circulation of mimiviruses in the Amazon Region of Brazil, we surveyed 513 serum samples from domestic and wild mammals. Neutralizing antibodies were detected in 15 sample pools, and mimivirus DNA was detected in 9 pools of serum from capuchin monkeys and in 16 pools of serum from cattle. PMID:24564967

Identification of a Proteinaceous Component in the Leaf of Moringa Oleifera lam. with Effects on High Serum Creatinine

PubMed Central

Sahoo, S.; Raghavendra, K. M.; Biswas, S.

2014-01-01

Moringa oleifera Lam. has been an important plant in the history of mankind, both for its nutritional and medicinal uses. Apart from bactericidal effects, the parts of this plant have been effectively used in the treatment of circulatory, respiratory, endocrine, digestive as well as neural disorders. Till date, though, there has been no reported activity of the involvement of any proteinaceous extract from M. oleifera on high levels of serum creatinine. To address this issue, blood samples with high levels of serum creatinine (2 mg/dl and above) were treated with leaf extract from M. oleifera. The crude extract was partially purified initially and eventually purified to completion as well. All these proteinaceous fractions were used to treat samples with high levels of serum creatinine as mentioned above. While the treatment of serum sample having high creatinine with crude extract and partially purified protein fractions showed a decrease of approximately 20% in the levels of serum creatinine over a period of 24 h, the samples treated with purified protein fraction reduced the serum creatinine level by 50%. In light of the fact that increased level of serum creatinine levels have adverse downstream effects on the heart, lungs and other organs, this communication assumes significance because it suggests a way of reducing the level of serum creatinine as an emergency measure. Further, the identification and characterisation of this proteinaceous component and possible in vivo experiments would provide a major tool for the treatment of downstream complications associated with increased serum creatinine via a new sources, albeit a natural one. PMID:24799742

Development of dried serum spot sampling techniques for the assessment of trace elements in serum samples by LA-ICP-MS.

PubMed

Chantada-Vázquez, María Pilar; Moreda-Piñeiro, Jorge; Cantarero-Roldán, Alicia; Bermejo-Barrera, Pilar; Moreda-Piñeiro, Antonio

2018-08-15

A novel approach for serum analysis by dried matrix spot (DMS) technique is proposed. The methodology consists of sampling filter paper discs (2.7 mm in diameter) containing the large amount of serum retained after a single spotting. Several oxidizers (sodium chlorate, sodium azide, acetic acid, formic acid, 1-butyl-3-methylimidazoliumm chloride/bromide) were tested (oxidizers premixed with the sample before spotting, and papers previously soaked in concentrated additive/oxidizer solutions). Direct multi-element determination (Al, Be, Ca, Cu, Fe, K, Li, Mg, Mn, Mo, Na, P, Rb, Se, V, and Zn) in dried serum spots at very low levels was therefore assessed by laser ablation (LA) coupled with inductively coupled plasma - mass spectrometry (ICP-MS). Laser ablation was performed using a focused Nd: YAG laser beam in lineal scan mode (wavelength 213 nm, laser fluency 2.2 J cm -2 , repetition rate 20 Hz, laser spot diameter 90 µm, depth 0 µm, scanning speed 12 µm s -1 ). Matrix-matched calibration mode and 13 C as internal standard (for signal intensities normalization) was used throughout the work. Limits of quantification were found to be from 21 µg L -1 to 221 mg L -1 . Repeatability (seven ablations of the same dried serum spot) and reproducibility (two ablations of seven dried serum spot from the same material) offered RSDs below 12% for all analytes, which seems satisfactory for clinical purposes. The method was validated by analyzing several certified reference materials (Seronorm™ level I and II trace elements in serum), and it was applied to several DMS from serum samples from healthy adults. Copyright © 2018 Elsevier B.V. All rights reserved.

Conjugated bile acids in gallbladder bile and serum as potential biomarkers for cholesterol polyps and adenomatous polyps.

PubMed

Zhao, Mei-Fen; Huang, Peng; Ge, Chun-Lin; Sun, Tao; Ma, Zhi-Gang; Ye, Fei-Fei

2016-02-28

To identify conjugated bile acids in gallbladder bile and serum as possible biomarkers for cholesterol polyps (CPs) and adenomatous polyps (APs). Gallbladder bile samples and serum samples were collected from 18 patients with CPs (CP group), 9 patients with APs (AP group), and 20 patients with gallstones (control group) from March to November, 2013. High performance liquid chromatography (HPLC) assay with ultraviolent detection was used to detect the concentration of 8 conjugated bile acids (glycocholic acid, GCA; taurocholic acid, TCA; glycochenodeoxycholic acid, GCDCA; taurochenodeoxycholic acid, TCDCA; glycodeoxycholic acid, GDCA; taurodeoxycholic acid, TDCA; taurolithocholic acid, TLCA; tauroursodeoxycholic acid, TUDCA) in bile samples and serum samples. The diagnostic efficacy of serum GCA, GCDCA and TCDCA was evaluated. These 8 conjugated bile acids in gallbladder bile and serum were completely identified within 10 minutes with good linearity (correlation coefficient: R>0.9900; linearity range: 3.91-500 µg/mL). Among these conjugated bile acids, the levels of gallbladder bile GCDCA and TCDCA in the CP group were significantly higher than those in the AP group (p<0.05). Furthermore, serum GCDCA and TCDCA as well as GCA were significantly higher in the AP group than the CP group (p<0.05). Serum GCDCA alone (≤12 µg/mL) had relatively better diagnostic efficacy than the other conjugated bile acids. The levels of serum GCA, GCDCA and TCDCA may be valuable for differentiation of APs and CPs.

Polyfluoroalkyl substance exposure in the Mid-Ohio River Valley, 1991-2012.

PubMed

Herrick, Robert L; Buckholz, Jeanette; Biro, Frank M; Calafat, Antonia M; Ye, Xiaoyun; Xie, Changchun; Pinney, Susan M

2017-09-01

Industrial discharges of perfluorooctanoic acid (PFOA) to the Ohio River, contaminating water systems near Parkersburg, WV, were previously associated with nearby residents' serum PFOA concentrations above US general population medians. Ohio River PFOA concentrations downstream are elevated, suggesting Mid-Ohio River Valley residents are exposed through drinking water. Quantify PFOA and 10 other per- and polyfluoroalkyl substances (PFAS) in Mid-Ohio River Valley resident sera collected between 1991 and 2013 and determine whether the Ohio River and Ohio River Aquifer are exposure sources. We measured eleven PFAS in 1608 sera from 931 participants. Serum PFOA concentration and water source associations were assessed using linear mixed-effects models. We estimated between-sample serum PFOA using one-compartment pharmacokinetics for participants with multiple samples. In serum samples collected as early as 1991, PFOA (median = 7.6 ng/mL) was detected in 99.9% of sera; 47% had concentrations greater than US population 95th percentiles. Five other PFAS were detected in greater than 82% of samples; median other PFAS concentrations were similar to the US general population. Serum PFOA was significantly associated with water source, sampling year, age at sampling, tap water consumption, pregnancy, gravidity and breastfeeding. Serum PFOA was 40-60% lower with granular activated carbon (GAC) use. Repeated measurements and pharmacokinetics suggest serum PFOA peaked 2000-2006 for participants using water without GAC treatment; where GAC was used, serum PFOA concentrations decreased from 1991 to 2012. Mid-Ohio River Valley residents appear to have PFOA, but not other PFAS, serum concentrations above US population levels. Drinking water from the Ohio River and Ohio River Aquifer, primarily contaminated by industrial discharges 209-666 km upstream, is likely the primary exposure source. GAC treatment of drinking water mitigates, but does not eliminate, PFOA exposure. Copyright © 2017 Elsevier Ltd. All rights reserved.

A method for the routine determination of aluminium in serum and water by flameless atomic absorption spectrometry.

PubMed

Parkinson, I S; Ward, M K; Kerr, D N

1982-10-27

A simple but reliable method for the routine determination of aluminium in serum and water by flameless atomic absorption spectrometry is described. No preparatory procedures are required for water samples, although serum is mixed with a wetting agent (Triton X-100) to allow complete combustion of the samples and to improve analytical precision. Precautions to prevent contamination during sample handling are discussed and instrumental parameters are defined. The method has a sensitivity of 35.5 pg and detection limits of 2.3 micrograms Al/l for serum and 1.3 micrograms Al/l for water. The method was used to determine the aluminium concentration in serum of 46 normal subjects. The mean aluminium content was 7.3 micrograms/l (range 2--15 micrograms/l.

Bead-based microfluidic immunoassay for diagnosis of Johne's disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wadhwa, Ashutosh; Foote, Robert; Shaw, Robert W

2012-01-01

Microfluidics technology offers a platform for development of point-of-care diagnostic devices for various infectious diseases. In this study, we examined whether serodiagnosis of Johne s disease (JD) can be conducted in a bead-based microfluidic assay system. Magnetic micro-beads were coated with antigens of the causative agent of JD, Mycobacterium avium subsp. paratuberculosis. The antigen-coated beads were incubated with serum samples of JD-positive or negative serum samples and then with a fluorescently-labeled secondary antibody (SAB). To confirm binding of serum antibodies to the antigen, the beads were subjected to flow cytometric analysis. Different conditions (dilutions of serum and SAB, types ofmore » SAB, and types of magnetic beads) were optimized for a great degree of differentiation between the JD-negative and JD-positive samples. Using the optimized conditions, we tested a well-classified set of 155 serum samples from JD negative and JD-positive cattle by using the bead-based flow cytometric assay. Of 105 JD-positive samples, 63 samples (60%) showed higher antibody binding levels than a cut-off value determined by using antibody binding levels of JD-negative samples. In contrast, only 43-49 JD-positive samples showed higher antibody binding levels than the cut-off value when the samples were tested by commercially-available immunoassays. Microfluidic assays were performed by magnetically immobilizing a number of beads within a microchannel of a glass microchip and detecting antibody on the collected beads by laser-induced fluorescence. Antigen-coated magnetic beads treated with bovine serum sample and fluorescently-labeled SAB were loaded into a microchannel to measure the fluorescence (reflecting level of antibody binding) on the beads in the microfluidic system. When the results of five bovine serum samples obtained with the system were compared to those obtained with the flow cytometer, a high level of correlation (linear regression, r2 = 0.994) was observed. In a further experiment, we magnetically immobilized antigen-coated beads in a microchannel, reacted the beads with serum and SAB in the channel, and detected antibody binding to the beads in the microfluidic system. A strong antibody binding in JD-positive serum was detected, whereas there was only negligible binding in negative control experiments. Our data suggest that the bead-based microfluidic system may form a basis for development of an on-site serodiagnosis of JD. Key Words: Mycobacterium avium ssp. paratuberculosis, Johne s disease, microfluidics, lab-on-a-chip.« less

Serum factors and clinical characteristics associated with serum E-Screen activity

PubMed Central

Wang, Jue; Trentham-Dietz, Amy; Hemming, Jocelyn D. C.; Hedman, Curtis J.; Sprague, Brian L.

2013-01-01

Background The E-Screen bioassay can measure the mitogenicity of human serum and thus may be useful as a biomarker in epidemiologic studies of breast cancer. While the assay’s MCF-7 cells are known to proliferate in response to estrogen, the specific determinants of variation in E-Screen activity in human serum samples are poorly understood. We sought to identify serum molecules and patient characteristics associated with serum E-Screen activity among postmenopausal women. Methods Postmenopausal women (N=219) aged 55–70 with no history of postmenopausal hormone use or breast cancer completed a questionnaire and provided a blood sample. Serum was analyzed for E-Screen activity and a variety of molecules including sex hormones, growth factors, and environmental chemicals. Stepwise selection procedures were used to identify correlates of E-Screen activity. Results Serum samples from all women had detectable E-Screen activity, with a median estradiol equivalents value of 0.027 ng/mL and interquartile range of 0.018–0.036 ng/mL. In the final multivariable-adjusted model, serum E-Screen activity was positively associated with serum estradiol, estrone, IGFBP-3, and testosterone levels (p<0.05), as well as body mass index (p=0.03). Serum E-Screen activity was lower among women with higher SHBG (p<0.0001) and progesterone levels (p=0.03). Conclusion Serum E-Screen activity varies according to levels of endogenous estrogens and other serum molecules. Obesity appears to confer additional serum mitogenicity beyond its impact on the measured hormones and growth factors. Impact By capturing mitogenicity due to a variety of patient and serum factors, the E-Screen may provide advantages for use as a biomarker in breast cancer studies. PMID:23588007

Correlation of serum IgG concentration in foals and refractometry index of the dam's pre- and post-parturient colostrums: an assessment for failure of passive transfer in foals.

PubMed

Korosue, Kenji; Murase, Harutaka; Sato, Fumio; Ishimaru, Mutsuki; Kotoyori, Yasumitsu; Nambo, Yasuo

2012-11-01

The object of this study was to evaluate the usefulness of measuring the differences in the values of the serum total protein (DVSTP) concentration of foals and the refractometry index (DVRI) of the milk of dams before and after nursing of the colostrum for assessing failure of passive transfer (FPT) in foals. Serum samples from 31 foals were collected before the first nursing and other 1 to 6 times between 4 and 24 hr after birth. Paired colostrum and milk samples were collected from 14 of their dams at the same time. Serum samples were analyzed for IgG concentration using a single radial immunodiffusion (SRID) test (98 samples) and total protein concentration using a temperature-compensating refractometer (98 samples). Colostrum and milk samples were analyzed for refractometry index (RI) using a Brix refractometer (71 samples). DVSTP concentration and DVRI were significantly correlated with serum IgG concentration. The negative predictive values (NPVs) of DVSTP concentration for detecting serum IgG concentrations<400 mg/dl and<800 mg/dl were 98.2% and 91.3% when the cutoff value is set to 0.4 mg/dl and 0.8 mg/dl, respectively. Furthermore, the NPVs of DVRI for detecting serum IgG concentrations<400 mg/dl and<800 mg/dl were 97.3% and 96.3% when the cutoff value is set to 6% and 10%, respectively. The results suggest that measurement of DVRI is useful in assessing FPT as an initial "stall-side" screening test, because it is easy, inexpensive to perform and allows for rapid interpretation.

Influence of the collection tube on metabolomic changes in serum and plasma.

PubMed

López-Bascón, M A; Priego-Capote, F; Peralbo-Molina, A; Calderón-Santiago, M; Luque de Castro, M D

2016-04-01

Major threats in metabolomics clinical research are biases in sampling and preparation of biological samples. Bias in sample collection is a frequently forgotten aspect responsible for uncontrolled errors in metabolomics analysis. There is a great diversity of blood collection tubes for sampling serum or plasma, which are widely used in metabolomics analysis. Most of the existing studies dealing with the influence of blood collection on metabolomics analysis have been restricted to comparison between plasma and serum. However, polymeric gel tubes, which are frequently proposed to accelerate the separation of serum and plasma, have not been studied. In the present research, samples of serum or plasma collected in polymeric gel tubes were compared with those taken in conventional tubes from a metabolomics perspective using an untargeted GC-TOF/MS approach. The main differences between serum and plasma collected in conventional tubes affected to critical pathways such as the citric acid cycle, metabolism of amino acids, fructose and mannose metabolism and that of glycerolipids, and pentose and glucuronate interconversion. On the other hand, the polymeric gel only promoted differences at the metabolite level in serum since no critical differences were observed between plasma collected with EDTA tubes and polymeric gel tubes. Thus, the main changes were attributable to serum collected in gel and affected to the metabolism of amino acids such as alanine, proline and threonine, the glycerolipids metabolism, and two primary metabolites such as aconitic acid and lactic acid. Therefore, these metabolite changes should be taken into account in planning an experimental protocol for metabolomics analysis. Copyright © 2016 Elsevier B.V. All rights reserved.

Performance of polymerase chain reaction for the diagnosis of cystic echinococcosis using serum, urine, and cyst fluid samples

PubMed Central

Chaya, DR; Parija, Subhash Chandra

2014-01-01

Introduction: Cystic echinococcosis (CE) is a chronic zoonosis which presents with variable clinical manifestations. Currently the diagnosis of this disease is based on radiological findings and serological tests which lack specificity. Although antigen detection from the cyst fluid is the most specific, it is seldom done due to the complications involved. Detecting the presence of Echinococcus granulosus specific deoxyribonucleic acid (DNA) by the polymerase chain reaction (PCR) could provide a definitive diagnosis of CE. Materials and Methods: An in-house PCR assay was devised to detect E. granulosus specific DNA in serum, urine and hydatid cyst fluid. The ability of the PCR to detect E. granulosus in the above mentioned samples were observed in comparison with other antigen and antibody detection tests. Results: Serum samples from surgically confirmed patients of CE with ruptured cysts contained the corresponding DNA while the in the majority of cases who had an intact cyst had no DNA of E. granulosus in their serum. DNA of E. granulosus was not found to be excreted in urine. PCR performed equal to antigen detection ELISA while testing hydatid cyst fluid samples. Conclusions: Serum and urine might not serve as useful samples for the molecular diagnosis of cystic echinococcosis. However, PCR can be useful on serum samples to detect ruptured hydatid cysts and on hydatid cyst fluid to confirm the parasitic diagnosis. PMID:24754027

Performance of polymerase chain reaction for the diagnosis of cystic echinococcosis using serum, urine, and cyst fluid samples.

PubMed

Chaya, Dr; Parija, Subhash Chandra

2014-01-01

Cystic echinococcosis (CE) is a chronic zoonosis which presents with variable clinical manifestations. Currently the diagnosis of this disease is based on radiological findings and serological tests which lack specificity. Although antigen detection from the cyst fluid is the most specific, it is seldom done due to the complications involved. Detecting the presence of Echinococcus granulosus specific deoxyribonucleic acid (DNA) by the polymerase chain reaction (PCR) could provide a definitive diagnosis of CE. An in-house PCR assay was devised to detect E. granulosus specific DNA in serum, urine and hydatid cyst fluid. The ability of the PCR to detect E. granulosus in the above mentioned samples were observed in comparison with other antigen and antibody detection tests. Serum samples from surgically confirmed patients of CE with ruptured cysts contained the corresponding DNA while the in the majority of cases who had an intact cyst had no DNA of E. granulosus in their serum. DNA of E. granulosus was not found to be excreted in urine. PCR performed equal to antigen detection ELISA while testing hydatid cyst fluid samples. Serum and urine might not serve as useful samples for the molecular diagnosis of cystic echinococcosis. However, PCR can be useful on serum samples to detect ruptured hydatid cysts and on hydatid cyst fluid to confirm the parasitic diagnosis.

Effect of storage duration on cytokine stability in human serum and plasma.

PubMed

Vincent, Fabien B; Nim, Hieu T; Lee, Jacinta P W; Morand, Eric F; Harris, James

2018-06-14

Quantification of analytes such as cytokines in serum samples is intrinsic to translational research in immune diseases. Optimising pre-analytical conditions is critical for ensuring study quality, including evaluation of cytokine stability. We aimed to evaluate the effect on cytokine stability of storage duration prior to freezing of serum, and compare to plasma samples obtained from patients with systemic lupus erythematosus (SLE). Protein stability was analysed by simultaneously quantifying 18 analytes using a custom multi-analyte profile in SLE patient serum and plasma samples that had been prospectively stored at 4 °C for pre-determined periods between 0 and 30 days, prior to freezing. Six analytes were excluded from analysis, because most tested samples were above or below the limit of detection. Amongst the 12 analysed proteins, 11 did not show significant signal degradation. Significant signal degradation was observed from the fourth day of storage for a single analyte, CCL19. Proteins levels were more stable in unseparated serum compared to plasma for most analytes, with the exception of IL-37 which appears slightly more stable in plasma. Based on this, a maximum 3 days of storage at 4 °C for unseparated serum samples is recommended for biobanked samples intended for cytokine analysis in studies of human immune disease. Copyright © 2018 Elsevier Ltd. All rights reserved.

Pre-analytical effects of blood sampling and handling in quantitative immunoassays for rheumatoid arthritis.

PubMed

Zhao, Xiaoyan; Qureshi, Ferhan; Eastman, P Scott; Manning, William C; Alexander, Claire; Robinson, William H; Hesterberg, Lyndal K

2012-04-30

Variability in pre-analytical blood sampling and handling can significantly impact results obtained in quantitative immunoassays. Understanding the impact of these variables is critical for accurate quantification and validation of biomarker measurements. Particularly, in the design and execution of large clinical trials, even small differences in sample processing and handling can have dramatic effects in analytical reliability, results interpretation, trial management and outcome. The effects of two common blood sampling methods (serum vs. plasma) and two widely-used serum handling methods (on the clot with ambient temperature shipping, "traditional", vs. centrifuged with cold chain shipping, "protocol") on protein and autoantibody concentrations were examined. Matched serum and plasma samples were collected from 32 rheumatoid arthritis (RA) patients representing a wide range of disease activity status. Additionally, a set of matched serum samples with two sample handling methods was collected. One tube was processed per manufacturer's instructions and shipped overnight on cold packs (protocol). The matched tube, without prior centrifugation, was simultaneously shipped overnight at ambient temperatures (traditional). Upon delivery, the traditional tube was centrifuged. All samples were subsequently aliquoted and frozen prior to analysis of protein and autoantibody biomarkers. Median correlation between paired serum and plasma across all autoantibody assays was 0.99 (0.98-1.00) with a median % difference of -3.3 (-7.5 to 6.0). In contrast, observed protein biomarker concentrations were significantly affected by sample types, with median correlation of 0.99 (0.33-1.00) and a median % difference of -10 (-55 to 23). When the two serum collection/handling methods were compared, the median correlation between paired samples for autoantibodies was 0.99 (0.91-1.00) with a median difference of 4%. In contrast, significant increases were observed in protein biomarker concentrations among certain biomarkers in samples processed with the 'traditional' method. Autoantibody quantification appears robust to both sample type (plasma vs. serum) and pre-analytical sample collection/handling methods (protocol vs. traditional). In contrast, for non-antibody protein biomarker concentrations, sample type had a significant impact; plasma samples generally exhibit decreased protein biomarker concentrations relative to serum. Similarly, sample handling significantly impacted the variability of protein biomarker concentrations. When biomarker concentrations are combined algorithmically into a single test score such as a multi-biomarker disease activity test for rheumatoid arthritis (MBDA), changes in protein biomarker concentrations may result in a bias of the score. These results illustrate the importance of characterizing pre-analytical methodology, sample type, sample processing and handling procedures for clinical testing in order to ensure test accuracy. Copyright © 2012 Elsevier B.V. All rights reserved.

Serosurvey of selected viruses in captive giant pandas (Ailuropoda melanoleuca) in China.

PubMed

Qin, Qin; Li, Desheng; Zhang, Hemin; Hou, Rong; Zhang, Zhihe; Zhang, Chenglin; Zhang, Jinguo; Wei, Fuwen

2010-05-19

Serum samples from 92 giant pandas in three captive facilities were tested for antibodies against five viruses of carnivores. Antibody titers against canine distemper virus (CDV) in two facilities in which giant pandas were vaccinated were variable. The canine adenovirus (CAV-1) and canine parvovirus (CPV) titers in vaccinated group were both positive, but titers were not high and varied among individual except one vaccinated panda had extremely high CAV-1 titer, indicating infection with the field virus following vaccination. Our results suggest that the vaccines used for these giant pandas do not elicit consistent antibody titers. Antibody titers against CDV, CPV and CAV-1 in unvaccinated giant pandas were highly variable, especially CPV titer. Almost half of sera were CPV antibody positive, and CPV titers were high enough to suggest infection with the virus. Canine coronavirus (CCV) and canine parainfluenza virus (CPIV) titers were not detected in all serum samples. The results of this study emphasize the need for research on infectious diseases of giant pandas and development of suitable vaccines for the species. Copyright 2009 Elsevier B.V. All rights reserved.

A rapid field test for sylvatic plague exposure in wild animals.

PubMed

Abbott, Rachel C; Hudak, Robert; Mondesire, Roy; Baeten, Laurie A; Russell, Robin E; Rocke, Tonie E

2014-04-01

Plague surveillance is routinely conducted to predict future epizootics in wildlife and exposure risk for humans. The most common surveillance method for sylvatic plague is detection of antibodies to Yersinia pestis F1 capsular antigen in sentinel animals, such as coyotes (Canis latrans). Current serologic tests for Y. pestis, hemagglutination (HA) test and enzyme-linked immunosorbent assay (ELISA), are expensive and labor intensive. To address this need, we developed a complete lateral flow device for the detection of specific antibodies to Y. pestis F1 and V antigens. Our test detected anti-F1 and anti-V antibodies in serum and Nobuto filter paper samples from coyotes, and in serum samples from prairie dogs (Cynomys ludovicianus), lynx (Lynx canadensis), and black-footed ferrets (Mustela nigripes). Comparison of cassette results for anti-F1 and anti-V antibodies with results of ELISA or HA tests showed correlations ranging from 0.68 to 0.98. This device provides an affordable, user-friendly tool that may be useful in plague surveillance programs and as a research tool.

A rapid field test for sylvatic plague exposure in wild animals

USGS Publications Warehouse

Abbott, Rachel C.; Hudak, Robert; Mondesire, Roy; Baeten, Laurie A.; Russell, Robin E.; Rocke, Tonie E.

2014-01-01

Plague surveillance is routinely conducted to predict future epizootics in wildlife and exposure risk for humans. The most common surveillance method for sylvatic plague is detection of antibodies to Yersinia pestis F1 capsular antigen in sentinel animals, such as coyotes (Canis latrans). Current serologic tests for Y. pestis, hemagglutination (HA) test and enzyme-linked immunosorbent assay (ELISA), are expensive and labor intensive. To address this need, we developed a complete lateral flow device for the detection of specific antibodies to Y. pestis F1 and V antigens. Our test detected anti-F1 and anti-V antibodies in serum and Nobuto filter paper samples from coyotes, and in serum samples from prairie dogs (Cynomys ludovicianus), lynx (Lynx canadensis), and black-footed ferrets (Mustela nigripes). Comparison of cassette results for anti-F1 and anti-V antibodies with results of ELISA or HA tests showed correlations ranging from 0.68 to 0.98. This device provides an affordable, user-friendly tool that may be useful in plague surveillance programs and as a research tool.

Hand-held optical sensor using denatured antibody coated electro-active polymer for ultra-trace detection of copper in blood serum and environmental samples.

PubMed

Chandra, Sutapa; Dhawangale, Arvind; Mukherji, Soumyo

2018-07-01

An optimum copper concentration in environment is highly desired for all forms of life. We have developed an ultrasensitive copper sensor which functions from femto to micro molar concentration accurately (R 2 = 0.98). The sensor is based on denatured antibody immunoglobulin G (IgG), immobilized on polyaniline (PAni) which in turn is the coating on the core of an optical fiber. The sensing relies on changes in evanescent wave absorbance in the presence of the analyte. The sensor showed excellent selectivity towards Cu (II) ions over all other metal ions. The sensor was tested with lake and marine water samples to determine unknown concentrations of copper ions and the recovery results were within 90-115%, indicating reasonable accuracy. We further integrated the fiber-optic sensor with a miniaturized hand-held instrumentation platform to develop an accurate and field deployable device which can broadly be applicable to determine Cu (II) concentration in a wide range of systems - natural water bodies, soil as well as blood serum. Copyright © 2018 Elsevier B.V. All rights reserved.

Coronavirus antibody detection in cats by computer-assisted kinetics-based enzyme-linked immunosorbent assay (KELA): field studies.

PubMed

Barlough, J E; Jacobson, R H; Sorresso, G P; Lynch, T J; Scott, F W

1986-07-01

A total of 2238 feline serum samples submitted to the New York State Diagnostic Laboratory over a 1-year period were tested for the presence of coronavirus antibodies, using the computer-assisted, kinetics-based enzyme-linked immunosorbent assay (KELA). Cats from which sera were obtained were categorized by sex, age, breed, and disease status, and variations in mean antibody titers for different sub-classifications within each category were analyzed by computerized statistical analysis. As expected, higher mean antibody titers were recorded for cats with feline infectious peritonitis, and for cats with a recent history of possible coronavirus exposure. However, an unexpected inverse relationship between coronavirus antibody titer and age was also found. Certain cattery-oriented pure breeds appeared to have higher mean antibody titers, because their sample populations contained a higher percentage of younger cats and cats of unknown age-groups which, over-all, had higher mean titers. Taken together, the data substantiated the efficacy of the computer-assisted KELA for routine detection of serum coronavirus antibodies in cats.

An enzyme-linked immunosorbent assay for the evaluation of thrombocytopenia induced by heparin.

PubMed

Howe, S E; Lynch, D M

1985-05-01

Five patients with heparin-associated thrombocytopenia (HAT) were evaluated by platelet aggregation and quantitation of immunoglobulin binding to intact target platelets in both the presence and absence of heparin. These patients developed thrombocytopenia (12,000 to 70,000 platelets/microliter) 7 to 15 days and embolic and hemorrhagic complications 9 to 15 days after the initiation of heparin therapy. Platelet aggregation after the addition of heparin was demonstrated in two of four HAT serum samples, whereas normal serum samples showed no significant platelet aggregation. The five HAT serum samples showed normal to elevated baseline serum platelet-bindable immunoglobulin (SPBIg) with a range of 4.3 to 11.4 fg/platelet (normal less than or equal to 1.0 to 6.5 fg/platelet). When HAT sera were incubated with target platelets and heparin (5 U/ml), the SPBIg increased to 8.5 to 37.5 fg/platelet, a mean increase of 148% in the presence of heparin. Normal and control serum samples (from 10 normal laboratory volunteers, nine patients without thrombocytopenia receiving heparin, nine patients with autoimmune thrombocytopenic purpura, and nine patients with nonimmune thrombocytopenia not receiving heparin) showed only a slight increase in SPBIg of 0 to 2.8 fg/platelet above baseline, a mean increase of 15% after heparin incubation with the serum samples. The measurement of SPBIg of washed platelets incubated with test serum samples in the presence and absence of heparin is potentially a specific and sensitive in vitro test for the diagnosis of HAT and may prove more sensitive than platelet aggregation studies with heparin.

METHOD FOR MICRORNA ISOLATION FROM CLINICAL SERUM SAMPLES

PubMed Central

Li, Yu; Kowdley, Kris V.

2012-01-01

MicroRNAs are a group of intracellular non-coding RNA molecules that have been implicated in a variety of human diseases. Due to their high stability in blood, microRNAs released into circulation could be potentially utilized as non-invasive biomarkers for diagnosis or prognosis. Current microRNA isolation protocols are specifically designed for solid tissues and are impractical for biomarker development utilizing small-volume serum samples on a large scale. Thus, a protocol for microRNA isolation from serum is needed to accommodate these conditions in biomarker development. To establish such a protocol, we developed a simplified approach to normalize sample input by using single synthetic spike-in microRNA. We evaluated three commonly used commercial microRNA isolation kits for the best performance by comparing RNA quality and yield. The manufacturer’s protocol was further modified to improve the microRNA yield from 200 μL of human serum. MicroRNAs isolated from a large set of clinical serum samples were tested on the miRCURY LNA real-time PCR panel and confirmed to be suitable for high-throughput microRNA profiling. In conclusion, we have established a proven method for microRNA isolation from clinical serum samples suitable for microRNA biomarker development. PMID:22982505

Genotyping of Single Nucleotide Polymorphisms in DNA Isolated from Serum Using Sequenom MassARRAY Technology.

PubMed

Clendenen, Tess V; Rendleman, Justin; Ge, Wenzhen; Koenig, Karen L; Wirgin, Isaac; Currie, Diane; Shore, Roy E; Kirchhoff, Tomas; Zeleniuch-Jacquotte, Anne

2015-01-01

Large epidemiologic studies have the potential to make valuable contributions to the assessment of gene-environment interactions because they prospectively collected detailed exposure data. Some of these studies, however, have only serum or plasma samples as a low quantity source of DNA. We examined whether DNA isolated from serum can be used to reliably and accurately genotype single nucleotide polymorphisms (SNPs) using Sequenom multiplex SNP genotyping technology. We genotyped 81 SNPs using samples from 158 participants in the NYU Women's Health Study. Each participant had DNA from serum and at least one paired DNA sample isolated from a high quality source of DNA, i.e. clots and/or cell precipitates, for comparison. We observed that 60 of the 81 SNPs (74%) had high call frequencies (≥95%) using DNA from serum, only slightly lower than the 85% of SNPs with high call frequencies in DNA from clots or cell precipitates. Of the 57 SNPs with high call frequencies for serum, clot, and cell precipitate DNA, 54 (95%) had highly concordant (>98%) genotype calls across all three sample types. High purity was not a critical factor to successful genotyping. Our results suggest that this multiplex SNP genotyping method can be used reliably on DNA from serum in large-scale epidemiologic studies.

Exploring the concurrent presence of hepatitis A virus genome in serum, stool, saliva, and urine samples of hepatitis A patients.

PubMed

Joshi, Madhuri S; Bhalla, Shilpa; Kalrao, Vijay R; Dhongade, Ramchandra K; Chitambar, Shobha D

2014-04-01

The use of saliva and urine as an alternative to serum samples for detection of anti-hepatitis A virus (HAV) IgM antibodies has been documented. However, these samples remain underreported or unexplored for shedding of HAV. To address this issue, paired serum, stool, saliva, and urine samples collected from hepatitis A patients were screened by reverse transcription polymerase chain reaction for detection of HAV RNA. HAV RNA was detected in 67.6% (44/65), 52.3% (34/65), 8.7% (5/57), and 12.3% (8/65) of the serum, stool, saliva, and urine samples, respectively. Phylogenetic analysis of nucleotide sequences obtained for partial RNA polymerase region grouped HAV strains from all of the clinical samples of the study in subgenotype IIIA. Low frequency of HAV nucleic acid in saliva and urine samples indicates limited utility of these samples in genomic studies on HAV but suggests its potential for transmission and infection of hepatitis A. Copyright © 2014 Elsevier Inc. All rights reserved.

A Schistosoma haematobium-specific real-time PCR for diagnosis of urogenital schistosomiasis in serum samples of international travelers and migrants.

PubMed

Cnops, Lieselotte; Soentjens, Patrick; Clerinx, Jan; Van Esbroeck, Marjan

2013-01-01

Diagnosis of urogenital schistosomiasis by microscopy and serological tests may be elusive in travelers due to low egg load and the absence of seroconversion upon arrival. There is need for a more sensitive diagnostic test. Therefore, we developed a real-time PCR targeting the Schistosoma haematobium-specific Dra1 sequence. The PCR was evaluated on urine (n = 111), stool (n = 84) and serum samples (n = 135), and one biopsy from travelers and migrants with confirmed or suspected schistosomiasis. PCR revealed a positive result in 7/7 urine samples, 11/11 stool samples and 1/1 biopsy containing S. haematobium eggs as demonstrated by microscopy and in 22/23 serum samples from patients with a parasitological confirmed S. haematobium infection. S. haematobium DNA was additionally detected by PCR in 7 urine, 3 stool and 5 serum samples of patients suspected of having schistosomiasis without egg excretion in urine and feces. None of these suspected patients demonstrated other parasitic infections except one with Blastocystis hominis and Entamoeba cyst in a fecal sample. The PCR was negative in all stool samples containing S. mansoni eggs (n = 21) and in all serum samples of patients with a microscopically confirmed S. mansoni (n = 22), Ascaris lumbricoides (n = 1), Ancylostomidae (n = 1), Strongyloides stercoralis (n = 1) or Trichuris trichuria infection (n = 1). The PCR demonstrated a high specificity, reproducibility and analytical sensitivity (0.5 eggs per gram of feces). The real-time PCR targeting the Dra1 sequence for S. haematobium-specific detection in urine, feces, and particularly serum, is a promising tool to confirm the diagnosis, also during the acute phase of urogenital schistosomiasis.

Expression of proteins in serum, synovial fluid, synovial membrane, and articular cartilage samples obtained from dogs with stifle joint osteoarthritis secondary to cranial cruciate ligament disease and dogs without stifle joint arthritis.

PubMed

Garner, Bridget C; Kuroki, Keiichi; Stoker, Aaron M; Cook, Cristi R; Cook, James L

2013-03-01

To identify proteins with differential expression between healthy dogs and dogs with stifle joint osteoarthritis secondary to cranial cruciate ligament (CCL) disease. Serum and synovial fluid samples obtained from dogs with stifle joint osteoarthritis before (n = 10) and after (8) surgery and control dogs without osteoarthritis (9) and archived synovial membrane and articular cartilage samples obtained from dogs with stifle joint osteoarthritis (5) and dogs without arthritis (5). Serum and synovial fluid samples were analyzed via liquid chromatography-tandem mass spectrometry; results were compared against a nonredundant protein database. Expression of complement component 3 in archived tissue samples was determined via immunohistochemical methods. No proteins had significantly different expression between serum samples of control dogs versus those of dogs with stifle joint osteoarthritis. Eleven proteins (complement component 3 precursor, complement factor I precursor, apolipoprotein B-100 precursor, serum paraoxonase and arylesterase 1, zinc-alpha-2-glycoprotein precursor, serum amyloid A, transthyretin precursor, retinol-binding protein 4 precursor, alpha-2-macroglobulin precursor, angiotensinogen precursor, and fibronectin 1 isoform 1 preproprotein) had significantly different expression (> 2.0-fold) between synovial fluid samples obtained before surgery from dogs with stifle joint osteoarthritis versus those obtained from control dogs. Complement component 3 was strongly expressed in all (5/5) synovial membrane samples of dogs with stifle joint osteoarthritis and weakly expressed in 3 of 5 synovial membrane samples of dogs without stifle joint arthritis. Findings suggested that the complement system and proteins involved in lipid and cholesterol metabolism may have a role in stifle joint osteoarthritis, CCL disease, or both.

Plasma and Serum Metabolite Association Networks: Comparability within and between Studies Using NMR and MS Profiling.

PubMed

Suarez-Diez, Maria; Adam, Jonathan; Adamski, Jerzy; Chasapi, Styliani A; Luchinat, Claudio; Peters, Annette; Prehn, Cornelia; Santucci, Claudio; Spyridonidis, Alexandros; Spyroulias, Georgios A; Tenori, Leonardo; Wang-Sattler, Rui; Saccenti, Edoardo

2017-07-07

Blood is one of the most used biofluids in metabolomics studies, and the serum and plasma fractions are routinely used as a proxy for blood itself. Here we investigated the association networks of an array of 29 metabolites identified and quantified via NMR in the plasma and serum samples of two cohorts of ∼1000 healthy blood donors each. A second study of 377 individuals was used to extract plasma and serum samples from the same individual on which a set of 122 metabolites were detected and quantified using FIA-MS/MS. Four different inference algorithms (ARANCE, CLR, CORR, and PCLRC) were used to obtain consensus networks. The plasma and serum networks obtained from different studies showed different topological properties with the serum network being more connected than the plasma network. On a global level, metabolite association networks from plasma and serum fractions obtained from the same blood sample of healthy people show similar topologies, and at a local level, some differences arise like in the case of amino acids.

Plasma and Serum Metabolite Association Networks: Comparability within and between Studies Using NMR and MS Profiling

PubMed Central

2017-01-01

Blood is one of the most used biofluids in metabolomics studies, and the serum and plasma fractions are routinely used as a proxy for blood itself. Here we investigated the association networks of an array of 29 metabolites identified and quantified via NMR in the plasma and serum samples of two cohorts of ∼1000 healthy blood donors each. A second study of 377 individuals was used to extract plasma and serum samples from the same individual on which a set of 122 metabolites were detected and quantified using FIA–MS/MS. Four different inference algorithms (ARANCE, CLR, CORR, and PCLRC) were used to obtain consensus networks. The plasma and serum networks obtained from different studies showed different topological properties with the serum network being more connected than the plasma network. On a global level, metabolite association networks from plasma and serum fractions obtained from the same blood sample of healthy people show similar topologies, and at a local level, some differences arise like in the case of amino acids. PMID:28517934

Organic cattle products: Authenticating production origin by analysis of serum mineral content.

PubMed

Rodríguez-Bermúdez, Ruth; Herrero-Latorre, Carlos; López-Alonso, Marta; Losada, David E; Iglesias, Roberto; Miranda, Marta

2018-10-30

An authentication procedure for differentiating between organic and non-organic cattle production on the basis of analysis of serum samples has been developed. For this purpose, the concentrations of fourteen mineral elements (As, Cd, Co, Cr, Cu, Fe, Hg, I, Mn, Mo, Ni, Pb, Se and Zn) in 522 serum samples from cows (341 from organic farms and 181 from non-organic farms), determined by inductively coupled plasma spectrometry, were used. The chemical information provided by serum analysis was employed to construct different pattern recognition classification models that predict the origin of each sample: organic or non-organic class. Among all classification procedures considered, the best results were obtained with the decision tree C5.0, Random Forest and AdaBoost neural networks, with hit levels close to 90% for both production types. The proposed method, involving analysis of serum samples, provided rapid, accurate in vivo classification of cattle according to organic and non-organic production type. Copyright © 2018 Elsevier Ltd. All rights reserved.

[Development of a SPA-ELISA method for detecting anti-coronavirus IgG antibodies in serum samples from fulvous fruit bats].

PubMed

Zhou, Jie; Liao, Yu-xue; Chen, Zhong; Li, Yu-chun; Gao, Lu-Lu; Chen, Yi-xiong; Cai, Lian-gong; Chen, Qing; Yu, Shou-yi

2008-05-01

To develop an simple and sensitive method for detecting anti-coronavirus IgG antibodies in bat sera based on enzyme-linked immunosorbent assay (ELISA). A commercial ELISA kit for detecting SARS-CoV antibody was modified for detecting coronavirus antibodies in bat serum samples. The second antibody in the kit was replaced with horseradish peroxidase-conjugated protein-A (HRP-SPA) based on the characteristics of binding between Staphylococcus aureus protein A (SPA) and mammal IgG Fc fragment. The sera of 55 fulvous fruit bats (Rousettus dasymallus) were tested using the SPA-ELISA. The test results of the positive and negative controls in the kit and the serum samples from convalescent ;patient were consistent with expectation. Coronavirus antibody was detected in 2 out of the 55 bat serum samples. Serum neutralization test confirmed the validity of the SPA-ELISA method. This SPA-ELISA method is applicable for detecting coronavirus antibody in bat sera.

Core lipid, surface lipid and apolipoprotein composition analysis of lipoprotein particles as a function of particle size in one workflow integrating asymmetric flow field-flow fractionation and liquid chromatography-tandem mass spectrometry

PubMed Central

Jones, Jeffery I.; Gardner, Michael S.; Schieltz, David M.; Parks, Bryan A.; Toth, Christopher A.; Rees, Jon C.; Andrews, Michael L.; Carter, Kayla; Lehtikoski, Antony K.; McWilliams, Lisa G.; Williamson, Yulanda M.; Bierbaum, Kevin P.; Pirkle, James L.; Barr, John R.

2018-01-01

Lipoproteins are complex molecular assemblies that are key participants in the intricate cascade of extracellular lipid metabolism with important consequences in the formation of atherosclerotic lesions and the development of cardiovascular disease. Multiplexed mass spectrometry (MS) techniques have substantially improved the ability to characterize the composition of lipoproteins. However, these advanced MS techniques are limited by traditional pre-analytical fractionation techniques that compromise the structural integrity of lipoprotein particles during separation from serum or plasma. In this work, we applied a highly effective and gentle hydrodynamic size based fractionation technique, asymmetric flow field-flow fractionation (AF4), and integrated it into a comprehensive tandem mass spectrometry based workflow that was used for the measurement of apolipoproteins (apos A-I, A-II, A-IV, B, C-I, C-II, C-III and E), free cholesterol (FC), cholesterol esters (CE), triglycerides (TG), and phospholipids (PL) (phosphatidylcholine (PC), sphingomyelin (SM), phosphatidylethanolamine (PE), phosphatidylinositol (PI) and lysophosphatidylcholine (LPC)). Hydrodynamic size in each of 40 size fractions separated by AF4 was measured by dynamic light scattering. Measuring all major lipids and apolipoproteins in each size fraction and in the whole serum, using total of 0.1 ml, allowed the volumetric calculation of lipoprotein particle numbers and expression of composition in molar analyte per particle number ratios. Measurements in 110 serum samples showed substantive differences between size fractions of HDL and LDL. Lipoprotein composition within size fractions was expressed in molar ratios of analytes (A-I/A-II, C-II/C-I, C-II/C-III. E/C-III, FC/PL, SM/PL, PE/PL, and PI/PL), showing differences in sample categories with combinations of normal and high levels of Total-C and/or Total-TG. The agreement with previous studies indirectly validates the AF4-LC-MS/MS approach and demonstrates the potential of this workflow for characterization of lipoprotein composition in clinical studies using small volumes of archived frozen samples. PMID:29634782

Optimization of the radioimmunoassays for measuring fentanyl and alfentanil in human serum

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schuettler, J.; White, P.F.

Measurement of serum fentanyl and alfentanil concentrations by radioimmunoassay (RIA) may result in significant errors and high variability when the technique described in the available fentanyl and alfentanil RIA kits is used. The authors found a 29-94% overestimation of measured fentanyl and alfentanil serum levels when 3H-fentanyl or 3H-alfentanil was added lastly to the mixture of antiserum and sample. This finding is related to a reduction in binding sites for the labeled compounds after preincubation of sample and antiserum. If this sequence is used, it becomes necessary to extend the incubation period up to 6 h for fentanyl and upmore » to 10 h for alfentanil in order to achieve equilibration between unlabeled and labeled drug with respect to antiserum binding. However, when antiserum is added lastly to the mixture of sample and labeled drug, measurement accuracy and precision for fentanyl and alfentanil serum concentrations are enhanced markedly. In addition, it is important to perform the calibration curves and sample measurements using the same medium (i.e., serum alone or a serum/buffer dilution). In summary, to optimize the RIA for fentanyl and alfentanil, the authors recommend the following: 1) adding the antiserum lastly to the mixture of sample and labeled drug; 2) performing calibration curves using patient's blank serum when possible; 3) carefully examining and standardizing each step of the RIA procedure to reduce variability, and, finally; 4) comparing results with those of other established RIA laboratories.« less

Steroid Profiling by Gas Chromatography–Mass Spectrometry and High Performance Liquid Chromatography–Mass Spectrometry for Adrenal Diseases

PubMed Central

McDonald, Jeffrey G.; Matthew, Susan

2012-01-01

The ability to measure steroid hormone concentrations in blood and urine specimens is central to the diagnosis and proper treatment of adrenal diseases. The traditional approach has been to assay each steroid hormone, precursor, or metabolite using individual aliquots of serum, each with a separate immunoassay. For complex diseases, such as congenital adrenal hyperplasia and adrenocortical cancer, in which the assay of several steroids is essential for management, this approach is time consuming and costly, in addition to using large amounts of serum. Gas chromatography/mass spectrometry profiling of steroid metabolites in urine has been employed for many years but only in a small number of specialized laboratories and suffers from slow throughput. The advent of commercial high-performance liquid chromatography instruments coupled to tandem mass spectrometers offers the potential for medium- to high-throughput profiling of serum steroids using small quantities of sample. Here, we review the physical principles of mass spectrometry, the instrumentation used for these techniques, the terminology used in this field and applications to steroid analysis. PMID:22170384

Surface imprinted beads for the recognition of human serum albumin.

PubMed

Bonini, Francesca; Piletsky, Sergey; Turner, Anthony P F; Speghini, Adolfo; Bossi, Alessandra

2007-04-15

The synthesis of poly-aminophenylboronic acid (ABPA) imprinted beads for the recognition of the protein human serum albumin (HSA) is reported. In order to create homogeneous recognition sites, covalent immobilisation of the template HSA was exploited. The resulting imprinted beads were selective for HSA. The indirect imprinting factor (IF) calculated from supernatant was 1.6 and the direct IF, evaluated from the protein recovered from the beads, was 1.9. The binding capacity was 1.4 mg/g, which is comparable to commercially available affinity materials. The specificity of the HSA recognition was evaluated with competitive experiments, indicating a molar ratio 4.5/1 of competitor was necessary to displace half of the bound HSA. The recognition and binding of the imprinted beads was also tested with a complex sample, human serum and targeted removal of HSA without a loss of the other protein components was demonstrated. The easy preparation protocol of derivatised beads and a good protein recognition properties make the approach an attractive solution to analytical and bio-analytical problems in the field of biotechnology.

Quantitating Iron in Serum Ferritin by Use of ICP-MS

NASA Technical Reports Server (NTRS)

Smith, Scott M.; Gillman, Patricia L.

2003-01-01

A laboratory method has been devised to enable measurement of the concentration of iron bound in ferritin from small samples of blood (serum). Derived partly from a prior method that depends on large samples of blood, this method involves the use of an inductively-coupled-plasma mass spectrometer (ICP-MS). Ferritin is a complex of iron with the protein apoferritin. Heretofore, measurements of the concentration of serum ferritin (as distinguished from direct measurements of the concentration of iron in serum ferritin) have been used to assess iron stores in humans. Low levels of serum ferritin could indicate the first stage of iron depletion. High levels of serum ferritin could indicate high levels of iron (for example, in connection with hereditary hemochromatosis an iron-overload illness that is characterized by progressive organ damage and can be fatal). However, the picture is complicated: A high level of serum ferritin could also indicate stress and/or inflammation instead of (or in addition to) iron overload, and low serum iron concentration could indicate inflammation rather than iron deficiency. Only when concentrations of both serum iron and serum ferritin increase and decrease together can the patient s iron status be assessed accurately. Hence, in enabling accurate measurement of the iron content of serum ferritin, the present method can improve the diagnosis of the patient s iron status. The prior method of measuring the concentration of iron involves the use of an atomic-absorption spectrophotometer with a graphite furnace. The present method incorporates a modified version of the sample- preparation process of the prior method. First, ferritin is isolated; more specifically, it is immobilized by immunoprecipitation with rabbit antihuman polyclonal antibody bound to agarose beads. The ferritin is then separated from other iron-containing proteins and free iron by a series of centrifugation and wash steps. Next, the ferritin is digested with nitric acid to extract its iron content. Finally, a micronebulizer is used to inject the sample of the product of the digestion into the ICPMS for analysis of its iron content. The sensitivity of the ICP-MS is high enough to enable it to characterize samples smaller than those required in the prior method (samples can be 0.15 to 0.60 mL).

Serial Sampling of Serum Protein Biomarkers for Monitoring Human Traumatic Brain Injury Dynamics: A Systematic Review.

PubMed

Thelin, Eric Peter; Zeiler, Frederick Adam; Ercole, Ari; Mondello, Stefania; Büki, András; Bellander, Bo-Michael; Helmy, Adel; Menon, David K; Nelson, David W

2017-01-01

The proteins S100B, neuron-specific enolase (NSE), glial fibrillary acidic protein (GFAP), ubiquitin carboxy-terminal hydrolase L1 (UCH-L1), and neurofilament light (NF-L) have been serially sampled in serum of patients suffering from traumatic brain injury (TBI) in order to assess injury severity and tissue fate. We review the current literature of serum level dynamics of these proteins following TBI and used the term "effective half-life" ( t 1/2 ) in order to describe the "fall" rate in serum. Through searches on EMBASE, Medline, and Scopus, we looked for articles where these proteins had been serially sampled in serum in human TBI. We excluded animal studies, studies with only one presented sample and studies without neuroradiological examinations. Following screening (10,389 papers), n  = 122 papers were included. The proteins S100B ( n  = 66) and NSE ( n  = 27) were the two most frequent biomarkers that were serially sampled. For S100B in severe TBI, a majority of studies indicate a t 1/2 of about 24 h, even if very early sampling in these patients reveals rapid decreases (1-2 h) though possibly of non-cerebral origin. In contrast, the t 1/2 for NSE is comparably longer, ranging from 48 to 72 h in severe TBI cases. The protein GFAP ( n  = 18) appears to have t 1/2 of about 24-48 h in severe TBI. The protein UCH-L1 ( n  = 9) presents a t 1/2 around 7 h in mild TBI and about 10 h in severe. Frequent sampling of these proteins revealed different trajectories with persisting high serum levels, or secondary peaks, in patients with unfavorable outcome or in patients developing secondary detrimental events. Finally, NF-L ( n  = 2) only increased in the few studies available, suggesting a serum availability of >10 days. To date, automated assays are available for S100B and NSE making them faster and more practical to use. Serial sampling of brain-specific proteins in serum reveals different temporal trajectories that should be acknowledged. Proteins with shorter serum availability, like S100B, may be superior to proteins such as NF-L in detection of secondary harmful events when monitoring patients with TBI.

Single exosome detection in serum using microtoroid optical resonators (Conference Presentation)

NASA Astrophysics Data System (ADS)

Su, Judith

2016-03-01

Recently exosomes have attracted interest due to their potential as cancer biomarkers. We report the real time, label-free sensing of single exosomes in serum using microtoroid optical resonators. We use this approach to assay the progression of tumors implanted in mice by specifically detecting low concentrations of tumor-derived exosomes. Our approach measures the adsorption of individual exosomes onto a functionalized silica microtoroid by tracking changes in the optical resonant frequency of the microtoroid. When exosomes land on the microtoroid, they perturb its refractive index in the evanescent field and thus shift its resonance frequency. Through digital frequency locking, we are able to rapidly track these shifts with accuracies of better than 10 attometers (one part in 10^11). Samples taken from tumor-implanted mice from later weeks generated larger frequency shifts than those from earlier weeks. Control samples taken from a mouse with no tumor generated no such increase in signal between subsequent weeks. Analysis of shifts from tumor-implanted mouse samples show a distribution of unitary steps, with the maximum step having a height of ~1.2 fm, corresponding to an exosome size of 44 ± 4.8 nm. This size range corresponds to that found by performing nanoparticle tracking analysis on the same samples. Our results demonstrate development towards a minimally-invasive tumor "biopsy" that eliminates the need to find and access a tumor.

Comparative analyses of semen and endocrine characteristics of free-living versus captive jaguars (Panthera onca).

PubMed

Morato, R G; Conforti, V A; Azevedo, F C; Jacomo, A T; Silveira, L; Sana, D; Nunes, A L; Guimarães, M A; Barnabe, R C

2001-11-01

Semen and blood samples were obtained from free-living (n = 6) and captive (n = 8) jaguars (Panthera onca) to compare reproductive characteristics between the two populations. Semen samples were analysed for volume (ml), percentage of motile spermatozoa, rate of forward progression (0-5), concentration (10(6) ml(-1)), total sperm count (10(6)) and sperm morphology. Serum testosterone concentration was determined by radioimmunoassay. Although ejaculate volume was greater in captive jaguars (n = 47 samples) than in free-living jaguars (n = 7 samples) (P < 0.05), the free-living jaguars produced more total spermatozoa (59.3 +/- 12.8 versus 152.0 +/- 88.0 x 10(6), respectively; not significant) with better viability and forward progression (2.8 +/- 0.1 versus 3.5 +/- 0.2, respectively; P < 0.05) and more spermatozoa with normal morphology (73.5 +/- 3.9 versus 5.0 +/- 1.1%, respectively; P < 0.05). Serum testosterone concentrations were similar for captive and free-living male jaguars (3.1 +/- 0.7 and 2.1 +/- 0.8 ng ml(-1), respectively). In summary, the data showed that semen may be collected successfully from free-living jaguars and evaluated under field conditions to establish normative reproductive values in this species. The results also indicate that jaguars maintained in zoos show inferior seminal characteristics compared with free-living animals.

Serum fibroblast growth factor 23 concentrations in dogs with chronic kidney disease.

PubMed

Dittmer, Keren E; Perera, Kalyani C; Elder, Peter A

2017-10-01

The aim of this study was to determine if serum fibroblast growth factor (FGF23) concentrations were increased in dogs with chronic kidney disease (CKD). Serum samples submitted to a commercial laboratory were collected over a 15-month period, 14 samples were from dogs with a history of polyuria/polydipsia, azotaemia and low urine specific gravity, 20 samples were from non-azotaemic dogs. Serum FGF23, parathyroid hormone, total calcium and phosphorus, urea and creatinine were measured. Mann-Whitney test was used to determine differences between non-azotaemic and CKD groups; a one-way ANOVA with Tukey pairwise comparisons was used to determine any differences between International Renal Interest Society stages; and regression models were used to determine predictors of International Renal Interest Society stage, serum phosphorus and FGF23 concentrations. The median serum FGF23 concentration of dogs with CKD was 5194.6pg/mL, which was significantly greater (P<0.001) than the median serum FGF23 concentration of non-azotaemic dogs (259.2pg/mL). Log serum FGF23 and age were significantly associated with IRIS stage (P=0.027 and P=0.032 respectively), while log serum phosphorus concentration (P<0.001) was significantly associated with log serum FGF23 concentration. In summary, serum FGF23 concentration is increased in dogs with CKD, and is associated with serum phosphorus concentration. This phosphatonin pathway may be a useful target for the development of future treatments to control plasma phosphorus concentrations in chronic kidney disease. Copyright © 2017 Elsevier Ltd. All rights reserved.

Heterophile antibody interference in qualitative urine/serum hCG devices: Case report.

PubMed

Patel, Khushbu K; Gronowski, Ann M

2016-06-01

This case report investigates the origin of a false positive result on a serum qualitative human chorionic gonadotropin (hCG) device. A 46-year-old woman diagnosed with chronic myeloid leukemia presented with nausea and vomiting. A qualitative serum hCG test was interpreted as positive; however, a quantitative serum hCG test was negative (<5IU/L). To further investigate this discrepancy, the sample was pretreated with heterophilic blocking reagent (HBR). Additionally, the sample was tested on other qualitative hCG devices composed of antibodies from different animal sources. Blocking reagent from an automated quantitative immunoassay was also tested for its ability to inhibit the heterophile antibody interference. The qualitative test result was negative after pretreatment with heterophilic blocking reagent. Other devices composed of antibodies from different animal sources also demonstrated mixed results with the patient's sample. Blocking reagent obtained from the automated quantitative assay inhibited the heterophile antibody interference in the patient's sample. This case demonstrates that positive serum point-of-care hCG results should be interpreted with caution and confirmed with a quantitative serum hCG immunoassay when clinical suspicion is raised. Copyright © 2016 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Evaluation of an enzyme immunoassay for detection of dengue virus NS1 antigen in human serum.

PubMed

Dussart, Philippe; Labeau, Bhety; Lagathu, Gisèle; Louis, Philippe; Nunes, Marcio R T; Rodrigues, Sueli G; Storck-Herrmann, Cécile; Cesaire, Raymond; Morvan, Jacques; Flamand, Marie; Baril, Laurence

2006-11-01

We evaluated a one-step sandwich-format microplate enzyme immunoassay for detecting dengue virus NS1 antigen (Ag) in human serum by use of Platelia Dengue NS1 Ag kits (Bio-Rad Laboratories, Marnes La Coquette, France). We collected 299 serum samples from patients with dengue disease and 50 serum samples from patients not infected with dengue virus. For the 239 serum samples from patients with acute infections testing positive by reverse transcription-PCR and/or virus isolation for one of the four dengue virus serotypes, the sensitivity of the Platelia Dengue NS1 Ag kit was 88.7% (95% confidence interval, 84.0% to 92.4%). None of the serum samples from patients not infected with dengue virus tested positive with the Platelia Dengue NS1 Ag kit. A diagnostic strategy combining the Platelia Dengue NS1 Ag test for acute-phase sera and immunoglobulin M capture enzyme-linked immunosorbent assay for early-convalescent-phase sera increased sensitivity only from 88.7% to 91.9%. Thus, NS1 antigen detection with the Platelia Dengue NS1 Ag kit could be used for first-line testing for acute dengue virus infection in clinical diagnostic laboratories.

Assessment of MR-based R2* and quantitative susceptibility mapping for the quantification of liver iron concentration in a mouse model at 7T.

PubMed

Simchick, Gregory; Liu, Zhi; Nagy, Tamas; Xiong, May; Zhao, Qun

2018-03-25

To assess the feasibility of quantifying liver iron concentration (LIC) using R2* and quantitative susceptibility mapping (QSM) at a high field strength of 7 Tesla (T). Five different concentrations of Fe-dextran were injected into 12 mice to produce various degrees of liver iron overload. After mice were sacrificed, blood and liver samples were harvested. Ferritin enzyme-linked immunosorbent assay (ELISA) and inductively coupled plasma mass spectrometry were performed to quantify serum ferritin concentration and LIC. Multiecho gradient echo MRI was conducted to estimate R2* and the magnetic susceptibility of each liver sample through complex nonlinear least squares fitting and a morphology enabled dipole inversion method, respectively. Average estimates of serum ferritin concentration, LIC, R2*, and susceptibility all show good linear correlations with injected Fe-dextran concentration; however, the standard deviations in the estimates of R2* and susceptibility increase with injected Fe-dextran concentration. Both R2* and susceptibility measurements also show good linear correlations with LIC (R 2  = 0.78 and R 2  = 0.91, respectively), and a susceptibility-to-LIC conversion factor of 0.829 ppm/(mg/g wet) is derived. The feasibility of quantifying LIC using MR-based  R2* and QSM at a high field strength of 7T is demonstrated. Susceptibility quantification, which is an intrinsic property of tissues and benefits from being field-strength independent, is more robust than R2* quantification in this ex vivo study. A susceptibility-to-LIC conversion factor is presented that agrees relatively well with previously published QSM derived results obtained at 1.5T and 3T. © 2018 International Society for Magnetic Resonance in Medicine.

Assessment of adenosine deaminase (ADA) activity and oxidative stress in patients with chronic tonsillitis.

PubMed

Garca, Mehmet Fatih; Demir, Halit; Turan, Mahfuz; Bozan, Nazım; Kozan, Ahmet; Belli, Şeyda Bayel; Arslan, Ayşe; Cankaya, Hakan

2014-06-01

To emphasize the effectiveness of adenosine deaminase (ADA) enzyme, which has important roles in the differentiation of lymphoid cells, and oxidative stress in patients with chronic tonsillitis. Serum and tissue samples were obtained from 25 patients who underwent tonsillectomy due to recurrent episodes of acute tonsillitis. In the control group, which also had 25 subjects, only serum samples were taken as obtaining tissue samples would not have been ethically appropriate. ADA enzyme activity, catalase (CAT), carbonic anhydrase (CA), nitric oxide (NO) and malondialdehyde (MDA) were measured in the serum and tissue samples of patients and control group subjects. The serum values of both groups were compared. In addition, the tissue and serum values of patients were compared. Serum ADA activity and the oxidant enzymes MDA and NO values of the patient group were significantly higher than those of the control group (p < 0.001), the antioxidant enzymes CA and CAT values of the patient group were significantly lower than those of the control group (p < 0.001). In addition, while CA, CAT and NO enzyme levels were found to be significantly higher in the tonsil tissue of the patient group when compared to serum levels (p < 0.05), there was no difference between tissue and serum MDA and ADA activity (p > 0.05). Elevated ADA activity may be effective in the pathogenesis of chronic tonsillitis both by impairing tissue structure and contributing to SOR formation.

Serum and egg yolk antibody detection in chickens infected with low pathogenicity avian influenza virus.

PubMed

Sá e Silva, Mariana; Swayne, David E

2012-09-01

Surveillance for low pathogenicity avian influenza virus (LPAIV) infections has primarily relied on labor-intensive collection and serological testing of serum, but for many poultry diseases, easier-to-collect yolk samples have replaced serum for surveillance testing. A time-course LPAIV infection study in layers was performed to evaluate the utility of antibody detection in serum vs. egg yolk samples. Layers inoculated with the LPAIV A/Bobwhite Quail/Pennsylvania/20304/98 (H7N2) were tested for antibody levels in the serum and egg yolk by using the agar gel immunodiffusion test (AGID), hemagglutination-inhibition test (HI), and a commercially available enzyme-linked immunosorbent assay (ELISA). Anti-influenza specific antibodies were detected in the serum as early as 7 days postinoculation (DPI), and the majority of the hens remained positive until 42 DPI. Antibodies in the egg yolk were first detected by AGID at 7 DPI, which was also the first day of detection in serum. However, the majority of the eggs were positive by all techniques at 11 DPI and remained positive until 42 DPI, at which time the number of AGID+ and HI+ samples declined slightly as compared to ELISA+ samples. These results suggest that egg yolk can be an alternative to serum for flock serological surveillance against LPAIV infections, and the three methods (AGID, HI, and ELISA) will give similar results for first 42 days after infection, although AGID may give earlier positive response.

Serum levels of perfluoroalkyl compounds in human maternal and umbilical cord blood samples

DOE Office of Scientific and Technical Information (OSTI.GOV)

Monroy, Rocio; Morrison, Katherine; Teo, Koon

2008-09-15

Perfluoroalkyl compounds (PFCs) are end-stage metabolic products from industrial flourochemicals used in the manufacture of plastics, textiles, and electronics that are widely distributed in the environment. The objective of the present study was to quantify exposure to perfluorooctane sulfonate (PFOS), perfluorooctanoate (PFOA), perfluorodecanoic acid (PFDeA), perfluorohexane sulfonate (PFHxS), perfluoroheptanoic acid (PFHpA), and perfluorononanoic acid (PFNA) in serum samples collected from pregnant women and the umbilical cord at delivery. Pregnant women (n=101) presenting for second trimester ultrasound were recruited and PFC residue levels were quantified in maternal serum at 24-28 weeks of pregnancy, at delivery, and in umbilical cord blood (UCB;more » n=105) by liquid chromatography-mass spectrometry. Paired t-test and multiple regression analysis were performed to determine the relationship between the concentrations of each analyte at different sample collection time points. PFOA and PFOS were detectable in all serum samples analyzed including the UCB. PFOS serum levels (mean{+-}S.D.) were significantly higher (p<0.001) in second trimester maternal serum (18.1{+-}10.9 ng/mL) than maternal serum levels at delivery (16.2{+-}10.4 ng/mL), which were higher than the levels found in UCB (7.3{+-}5.8 ng/mL; p<0.001). PFHxS was quantifiable in 46/101 (45.5%) maternal and 21/105 (20%) UCB samples with a mean concentration of 4.05{+-}12.3 and 5.05{+-}12.9 ng/mL, respectively. There was no association between serum PFCs at any time point studied and birth weight. Taken together our data demonstrate that although there is widespread exposure to PFCs during development, these exposures do not affect birth weight.« less

Chemistry Testing on Plasma Versus Serum Samples in Dialysis Patients: Clinical and Quality Improvement Implications.

PubMed

Carey, Roger Neill; Jani, Chinu; Johnson, Curtis; Pearce, Jim; Hui-Ng, Patricia; Lacson, Eduardo

2016-09-07

Plasma samples collected in tubes containing separator gels have replaced serum samples for most chemistry tests in many hospital and commercial laboratories. Use of plasma samples for blood tests in the dialysis population eliminates delays in sample processing while waiting for clotting to complete, laboratory technical issues associated with fibrin formation, repeat sample collection, and patient care issues caused by delay of results because of incompletely clotted specimens. Additionally, a larger volume of plasma is produced than serum for the same amount of blood collected. Plasma samples are also acceptable for most chemical tests involved in the care of patients with ESRD. This information becomes very important when United States regulatory requirements for ESRD inadvertently limit the type of sample that can be used for government reporting, quality assessment, and value-based payment initiatives. In this narrative, we summarize the renal community experience and how the subsequent resolution of the acceptability of phosphorus levels measured from serum and plasma samples may have significant implications in the country's continued development of a value-based Medicare ESRD Quality Incentive Program. Copyright © 2016 by the American Society of Nephrology.

Preconcentration and Separation of Mixed-Species Samples Near a Nano-Junction in a Convergent Microchannel

PubMed Central

Chiu, Ping-Hsien; Weng, Chen-Hsun; Yang, Ruey-Jen

2015-01-01

A fluidic microchip incorporating a convergent microchannel and a Nafion-nanoporous membrane is proposed for the preconcentration and separation of multi-species samples on a single platform. In the device, sample preconcentration is achieved by means of the ion concentration polarization effect induced at the micro/nano interface under the application of an external electric field, while species separation is achieved by exploiting the different electrophoretic mobilities of the sample components. The experimental results show that the device is capable of detecting C-reactive protein (CRP) with an initial concentration as low as 9.50 × 10−6 mg/L given a sufficient preconcentration time and driving voltage. In addition, it is shown that a mixed-species sample consisting of three negatively-charged components (bovine serum albumin (BSA), tetramethylrhodamine(TAMRA) isothiocyanate-Dextran and fluorescent polymer beads) can be separated and preconcentrated within 20 min given a driving voltage of 100 V across 1 cm microchannel in length. In general, the present results confirm the feasibility of the device for the immunoassay or detection of various multi-species samples under low concentration in the biochemical and biomedical fields. The novel device can therefore improve the detection limit of traditional medical facilities. PMID:26690167

Evaluation of the COBAS AMPLICOR CMV MONITOR test for detection of viral DNA in specimens taken from patients after liver transplantation.

PubMed

Sia, I G; Wilson, J A; Espy, M J; Paya, C V; Smith, T F

2000-02-01

Detection of cytomegalovirus (CMV) DNA in blood by PCR is a sensitive method for the detection of infection in patients posttransplantation. The test, however, has low specificity for the identification of overt CMV disease. Quantitative CMV PCR has been shown to overcome this shortcoming. The COBAS AMPLICOR CMV MONITOR test was evaluated by using consecutive serum and peripheral blood mononuclear cell (PBMN) samples from liver transplant patients. Twenty-five patients had CMV viremia (by shell vial cell culture assay) and/or tissue-invasive disease (by biopsy); 20 had no active infection. A total of 262 serum and 62 PBMN specimens were tested. Of 159 serum specimens from patients with overt CMV infection, the COBAS assay detected CMV DNA in 21 patients (sensitivity, 84%). Only 1 of 103 samples from patients with no evidence of active infection had detectable CMV DNA (341 copies/ml). By comparison of 62 matching serum and PBMN samples by the same assay, 12 PBMN samples were exclusively positive, whereas only 2 serum samples were exclusively positive (P < 0.05). At the time of clinical CMV infection, viral copy numbers were higher in PBMNs than serum from four of five patients. The COBAS AMPLICOR CMV MONITOR test is a sensitive and specific test for the quantitative detection of CMV DNA in blood. Clinical applications of the assay will require further validation with samples from a larger population of transplant patients.

Evaluation of the COBAS AMPLICOR CMV MONITOR Test for Detection of Viral DNA in Specimens Taken from Patients after Liver Transplantation

PubMed Central

Sia, Irene G.; Wilson, Jennie A.; Espy, Mark J.; Paya, Carlos V.; Smith, Thomas F.

2000-01-01

Detection of cytomegalovirus (CMV) DNA in blood by PCR is a sensitive method for the detection of infection in patients posttransplantation. The test, however, has low specificity for the identification of overt CMV disease. Quantitative CMV PCR has been shown to overcome this shortcoming. The COBAS AMPLICOR CMV MONITOR test was evaluated by using consecutive serum and peripheral blood mononuclear cell (PBMN) samples from liver transplant patients. Twenty-five patients had CMV viremia (by shell vial cell culture assay) and/or tissue-invasive disease (by biopsy); 20 had no active infection. A total of 262 serum and 62 PBMN specimens were tested. Of 159 serum specimens from patients with overt CMV infection, the COBAS assay detected CMV DNA in 21 patients (sensitivity, 84%). Only 1 of 103 samples from patients with no evidence of active infection had detectable CMV DNA (341 copies/ml). By comparison of 62 matching serum and PBMN samples by the same assay, 12 PBMN samples were exclusively positive, whereas only 2 serum samples were exclusively positive (P < 0.05). At the time of clinical CMV infection, viral copy numbers were higher in PBMNs than serum from four of five patients. The COBAS AMPLICOR CMV MONITOR test is a sensitive and specific test for the quantitative detection of CMV DNA in blood. Clinical applications of the assay will require further validation with samples from a larger population of transplant patients. PMID:10655353

Interlaboratory comparison of test results for detection of Lyme disease by 516 participants in the Wisconsin State Laboratory of Hygiene/College of American Pathologists Proficiency Testing Program.

PubMed Central

Bakken, L L; Callister, S M; Wand, P J; Schell, R F

1997-01-01

In 1991, we reported that 55% of laboratories participating in the Wisconsin Proficiency Testing Program could not accurately identify serum samples from Lyme disease patients containing antibody against Borrelia burgdorferi. The purpose of this study was to determine whether the accuracy of Lyme disease test results reported by approximately 500 participants in the Wisconsin State Laboratory of Hygiene/College of American Pathologists Lyme Disease Survey had improved. From 1992 through 1994, 50 serum samples were sent to participants of the survey. Each laboratory received 28 serum samples from individuals with Lyme disease according to the case definition of the Centers for Disease Control and Prevention and 22 serum samples from healthy individuals. Unfortunately, the serodiagnosis of Lyme disease by participants had not improved. The specificity of the Lyme disease assays steadily decreased from approximately 95% to approximately 81% during the 3-year period of the survey. False-positive test results approached 55% with some of the serum samples from healthy donors. A serum sample containing antibody against Treponema pallidum was reported as positive by 70% of the participants. In addition, the sensitivity fluctuated between 93 and 75%, depending upon the conjugate used by the laboratories. These results suggest that stronger criteria must be applied for approving and continuing to approve commercially available kits for the serodiagnosis of Lyme disease. PMID:9041384

Hair mercury association with selenium, serum lipid spectrum, and gamma-glutamyl transferase activity in adults.

PubMed

Tinkov, Alexey A; Skalnaya, Margarita G; Demidov, Vasily A; Serebryansky, Eugeny P; Nikonorov, Alexandr A; Skalny, Anatoly V

2014-12-01

The primary objective of the research is to estimate the dependence between hair mercury content, hair selenium, mercury-to-selenium ratio, serum lipid spectrum, and gamma-glutamyl transferase (GGT) activity in 63 adults (40 men and 23 women). Serum triglyceride (TG) concentration in the high-mercury group significantly exceeded the values obtained for low- and medium-mercury groups by 72 and 42 %, respectively. Serum GGT activity in the examinees from high-Hg group significantly exceeded the values of the first and the second groups by 75 and 28 %, respectively. Statistical analysis of the male sample revealed similar dependences. Surprisingly, no significant changes in the parameters analyzed were detected in the female sample. In all analyzed samples, hair mercury was not associated with hair selenium concentrations. Significant correlation between hair mercury content and serum TG concentration (r = 0.531) and GGT activity (r = 0.524) in the general sample of the examinees was detected. The respective correlations were observed in the male sample. Hair mercury-to-selenium ratios significantly correlated with body weight (r = 0.310), body mass index (r = 0.250), serum TG (r = 0.389), atherogenic index (r = 0.257), and GGT activity (r = 0.393). The same correlations were observed in the male sample. Hg/Se ratio in women did not correlate with the analyzed parameters. Generally, the results of the current study show the following: (1) hair mercury is associated with serum TG concentration and GGT activity in men, (2) hair selenium content is not related to hair mercury concentration, and (3) mercury-to-selenium ratio correlates with lipid spectrum parameters and GGT activity.

Use of Fourier-transform infrared spectroscopy to quantify immunoglobulin G concentrations in alpaca serum.

PubMed

Burns, J; Hou, S; Riley, C B; Shaw, R A; Jewett, N; McClure, J T

2014-01-01

Rapid, economical, and quantitative assays for measurement of camelid serum immunoglobulin G (IgG) are limited. In camelids, failure of transfer of maternal immunoglobulins has a reported prevalence of up to 20.5%. An accurate method for quantifying serum IgG concentrations is required. To develop an infrared spectroscopy-based assay for measurement of alpaca serum IgG and compare its performance to the reference standard radial immunodiffusion (RID) assay. One hundred and seventy-five privately owned, healthy alpacas. Eighty-two serum samples were collected as convenience samples during routine herd visits whereas 93 samples were recruited from a separate study. Serum IgG concentrations were determined by RID assays and midinfrared spectra were collected for each sample. Fifty samples were set aside as the test set and the remaining 125 training samples were employed to build a calibration model using partial least squares (PLS) regression with Monte Carlo cross validation to determine the optimum number of PLS factors. The predictive performance of the calibration model was evaluated by the test set. Correlation coefficients for the IR-based assay were 0.93 and 0.87, respectively, for the entire data set and test set. Sensitivity in the diagnosis of failure of transfer of passive immunity (FTPI) ([IgG] <1,000 mg/dL) was 71.4% and specificity was 100% for the IR-based method (test set) as gauged relative to the RID reference method assay. This study indicated that infrared spectroscopy, in combination with chemometrics, is an effective method for measurement of IgG in alpaca serum. Copyright © 2014 by the American College of Veterinary Internal Medicine.

Occurrence of infectious laryngotracheitis outbreaks in commercial layer hens detected by ELISA.

PubMed

Aras, Zeki; Yavuz, Orhan; Sanioğlu Gölen, Gökçenur

2018-02-09

Infectious laryngotracheitis (ILT) is an acute respiratory disease of chickens and a cause of great economic loss in commercial layers. The aims of this study were to investigate the prevalence of ILT in the field outbreaks and to compare the characteristics of ILT-infected and free flocks of commercial layers. A total of 625 blood serum samples were collected from 25 different layer flocks. The presence of antibodies against infectious laryngotracheitis virus (ILTV) in each sample was determined by ELISA. Of the 625 serum samples, 266 (42.56%) were found to be positive for ILTV antibodies. A total of 16 (64%) flocks were detected ILT positive by ELISA method. The mortality of infected flocks was statistically higher (P < 0.05) than uninfected flocks. The egg production of positive flocks was lower than that of the free flocks, but this difference was not statistically significant. The average live weight of hens in infected flocks was lower (P > 0.05) than hens in free flocks. In conclusion, the results of this study indicated a high prevalence of ILT infection in the commercial layer flocks in Konya region, Turkey. In outbreaks, ILT significantly increased the mortality rate and decreased the average live weight in layer hens.

Diagnosis of paratuberculosis by fecal culture and ELISA on milk and serum samples in two types of Chilean dairy goat herds.

PubMed

Salgado, Miguel; Kruze, Juan; Collins, Michael T

2007-01-01

Fecal culture has been the primary method used to diagnose paratuberculosis in goats. It is laborious, slow, and expensive. Validation of enzyme-linked immunosorbent assays (ELISAs) on milk samples could make paratuberculosis testing more widely available for goat farmers. The aim of this study was to determine the accuracy of serum and milk ELISAs for paratuberculosis, relative to fecal culture, in Chilean dairy goats. Eight dairy goat herds were selected. Feces, blood, and milk samples were collected from all female goats >2 years old. Fecal samples were cultured using Herrold egg yolk medium with mycobactin J and antibiotics. Serum and milk samples were tested using a commercial ELISA kit for Mycobacterium avium subsp. paratuberculosis antibody detection. A total of 383 goats were tested by ELISA and fecal culture. The sensitivity of ELISA on serum and milk relative to fecal culture was 74.3% (95% CI: 59.8-88.8) and 60% (95% CI: 43.8-76.2), respectively. The corresponding values for ELISA specificity based on the percentage of non- M. avium subsp. paratuberculosis-infected goats testing ELISA-negative were 98.6% (95% CI: 96.6-100) and 99.3% (95% CI: 97.9-100) on serum and milk, respectively. Proportions of positive results for serum and fecal samples were significantly different, whereas the proportions of positive results for milk and fecal samples were not significantly different. The milk ELISA had a moderate level of agreement with fecal culture results (Kappa = 0.57). The paratuberculosis ELISA on goat milk samples may be a cost-effective, accurate alternative to fecal culture.

Application of laboratory and portable attenuated total reflectance infrared spectroscopic approaches for rapid quantification of alpaca serum immunoglobulin G

PubMed Central

Burns, Jennifer B.; Riley, Christopher B.; Shaw, R. Anthony; McClure, J. Trenton

2017-01-01

The objective of this study was to develop and compare the performance of laboratory grade and portable attenuated total reflectance infrared (ATR-IR) spectroscopic approaches in combination with partial least squares regression (PLSR) for the rapid quantification of alpaca serum IgG concentration, and the identification of low IgG (<1000 mg/dL), which is consistent with the diagnosis of failure of transfer of passive immunity (FTPI) in neonates. Serum samples (n = 175) collected from privately owned, healthy alpacas were tested by the reference method of radial immunodiffusion (RID) assay, and laboratory grade and portable ATR-IR spectrometers. Various pre-processing strategies were applied to the ATR-IR spectra that were linked to corresponding RID-IgG concentrations, and then randomly split into two sets: calibration (training) and test sets. PLSR was applied to the calibration set and calibration models were developed, and the test set was used to assess the accuracy of the analytical method. For the test set, the Pearson correlation coefficients between the IgG measured by RID and predicted by both laboratory grade and portable ATR-IR spectrometers was 0.91. The average differences between reference serum IgG concentrations and the two IR-based methods were 120.5 mg/dL and 71 mg/dL for the laboratory and portable ATR-IR-based assays, respectively. Adopting an IgG concentration <1000 mg/dL as the cut-point for FTPI cases, the sensitivity, specificity, and accuracy for identifying serum samples below this cut point by laboratory ATR-IR assay were 86, 100 and 98%, respectively (within the entire data set). Corresponding values for the portable ATR-IR assay were 95, 99 and 99%, respectively. These results suggest that the two different ATR-IR assays performed similarly for rapid qualitative evaluation of alpaca serum IgG and for diagnosis of IgG <1000 mg/dL, the portable ATR-IR spectrometer performed slightly better, and provides more flexibility for potential application in the field. PMID:28651006

Application of laboratory and portable attenuated total reflectance infrared spectroscopic approaches for rapid quantification of alpaca serum immunoglobulin G.

PubMed

Elsohaby, Ibrahim; Burns, Jennifer B; Riley, Christopher B; Shaw, R Anthony; McClure, J Trenton

2017-01-01

The objective of this study was to develop and compare the performance of laboratory grade and portable attenuated total reflectance infrared (ATR-IR) spectroscopic approaches in combination with partial least squares regression (PLSR) for the rapid quantification of alpaca serum IgG concentration, and the identification of low IgG (<1000 mg/dL), which is consistent with the diagnosis of failure of transfer of passive immunity (FTPI) in neonates. Serum samples (n = 175) collected from privately owned, healthy alpacas were tested by the reference method of radial immunodiffusion (RID) assay, and laboratory grade and portable ATR-IR spectrometers. Various pre-processing strategies were applied to the ATR-IR spectra that were linked to corresponding RID-IgG concentrations, and then randomly split into two sets: calibration (training) and test sets. PLSR was applied to the calibration set and calibration models were developed, and the test set was used to assess the accuracy of the analytical method. For the test set, the Pearson correlation coefficients between the IgG measured by RID and predicted by both laboratory grade and portable ATR-IR spectrometers was 0.91. The average differences between reference serum IgG concentrations and the two IR-based methods were 120.5 mg/dL and 71 mg/dL for the laboratory and portable ATR-IR-based assays, respectively. Adopting an IgG concentration <1000 mg/dL as the cut-point for FTPI cases, the sensitivity, specificity, and accuracy for identifying serum samples below this cut point by laboratory ATR-IR assay were 86, 100 and 98%, respectively (within the entire data set). Corresponding values for the portable ATR-IR assay were 95, 99 and 99%, respectively. These results suggest that the two different ATR-IR assays performed similarly for rapid qualitative evaluation of alpaca serum IgG and for diagnosis of IgG <1000 mg/dL, the portable ATR-IR spectrometer performed slightly better, and provides more flexibility for potential application in the field.

Toward Standardization of Epstein-Barr Virus DNA Load Monitoring: Unfractionated Whole Blood as Preferred Clinical Specimen

PubMed Central

Stevens, Servi J. C.; Pronk, Inge; Middeldorp, Jaap M.

2001-01-01

Epstein-Barr virus (EBV) DNA load monitoring in peripheral blood has been shown to be a useful tool for the diagnosis of aberrant EBV infections. In the present study we compared the relative diagnostic values of EBV DNA load monitoring in unfractionated whole blood and simultaneously obtained serum or plasma samples from Burkitt's lymphoma (BL) patients, transplant recipients, human immunodeficiency virus (HIV)-infected individuals, and infectious mononucleosis (IM) patients by a quantitative competitive PCR (Q-PCR). The EBV DNA load in BL patients was mainly situated in the cellular blood compartment (up to 4.5 × 106 copies/ml). EBV DNA loads in unfractionated whole blood and parallel serum samples showed no correlation. In transplant recipients, IM patients, and HIV-infected patients, the EBV burden in the circulation was almost exclusively restricted to the cellular blood compartment, because serum or plasma samples from these patients yielded negative results by Q-PCR, despite high viral loads in corresponding whole-blood samples. A 10-fold more sensitive but qualitative BamHI-W-repeat PCR occasionally revealed the presence of EBV at <2,000 copies of EBV DNA per ml of serum. Spiking of 100 copies of EBV DNA in samples with negative Q-PCR results excluded the presence of inhibitory factors in serum or plasma that influenced the Q-PCR result. Serum samples from all populations were often positive for β-globin DNA, indicating cell damage in vivo or during serum preparation. We conclude that serum is an undesirable clinical specimen for EBV DNA load monitoring because it omits the presence of cell-associated virus and uncontrolled cell lysis may give irreproducible results or overestimation of the DNA load. Unfractionated whole blood is strongly preferred since it combines all blood compartments that may harbor EBV and it best reflects the absolute viral burden in the patient's circulation. PMID:11283029

Design of Indigenous ELISA Using Tachyzoites from the RH Strain of Toxoplasma gondii and Comparison with Commercial Kits in Ahvaz, Southwest of Iran, 2015.

PubMed

Mohammadpour, Niloofar; Saki, Jasem; Rafiei, Abdollah; Khodadadi, Ali; Tavalla, Mehdi; Cheraghian, Bahman

2016-10-01

Toxoplasma gondii is one of the most common causes of latent infections in humans worldwide. Detecting anti- Toxoplasma antibodies in serum using serological tests is a common method to diagnose toxoplasmosis. In the present study, an indigenous ELISA kit was prepared using tachyzoites from the RH strain of T. gondii , and its sensitivity and specificity were compared with those of commercial kits. To produce antigens, 0.02 mL of locally isolated T. gondii RH strain parasites along with 10 9 tachyzoites were injected into the peritoneal cavities of 50 laboratory mice (BALB/C). Parasites were collected after 4 days. After filtering and washing, the concentration of protein in sonicated tachyzoites was calculated using the Lowry protein assay. The dilution of antigen, serum and alkaline phosphatase conjugate was assessed in designing an indigenous ELISA method; then ELISA was performed based on these dilutions, and its sensitivity was determined using 200 serum samples. In addition, the specificity of the assay was evaluated using 40 serum samples from patients with tuberculosis, leukemia or hydatid cyst. Indigenous ELISA was used to examine 100 serum samples containing anti- T. gondii IgG, with a sensitivity of 98% (commercial kits: 100%). Another 100 serum samples containing anti- T. gondii IgM were also tested, with a sensitivity of 99% (commercial kits: 100%). When 40 serum samples from patients with leukemia, hydatid cyst or tuberculosis were examined using anti- T. gondii IgG, the specificity was 100%, identical to commercial kits. However, the specificity of a similar test with anti- T. gondii IgM was just 28.6% for serum samples from leukemia patients, 21.4% for hydatid cyst and 16.7% for tuberculosis. We found that purified locally isolated soluble crude antigens of the RH strain of T. gondii from the peritoneal cavity of mice may be one of the most promising antigens for detection of human toxoplasmosis in routine screening.

HBV serum DNA and RNA levels in nucleos(t)ide analogue-treated or untreated patients during chronic and acute infection.

PubMed

Butler, Emily K; Gersch, Jeffrey; McNamara, Anne; Luk, Ka-Cheung; Holzmayer, Vera; de Medina, Maria; Schiff, Eugene; Kuhns, Mary; Cloherty, Gavin A

2018-05-07

Treatment of chronic hepatitis B (CHB) patients with nucleos(t)ide analogues (NA) suppresses HBV DNA synthesis but does not affect synthesis of HBV pregenomic RNA (pgRNA). HBV pgRNA is detectable in the serum during NA treatment and has been proposed as a marker of HBV covalently closed circular DNA (cccDNA) activity within the infected hepatocyte. We developed an automated assay for the quantification of serum HBV pgRNA using a dual-target qRT-PCR approach on the Abbott m2000sp/rt system. We demonstrate accurate detection and quantification of serum HBV RNA. HBV DNA was quantified using the Abbott RealTime HBV viral load assay. We further compared serum nucleic acid levels and kinetics in HBV-positive populations. Samples included: on-therapy CHB samples (N=16), samples (N=89) from 10 treatment naïve CHB subjects receiving 12-weeks of NA treatment with 8-week follow-up, HBsAg-positive blood donor samples (N=102), and 3 seroconversion series from plasmapheresis donors (N=79 samples). During NA treatment of CHB subjects, we observed low correlation of HBV DNA to pgRNA levels; pgRNA concentration was generally higher than HBV DNA concentrations. In contrast, when NA treatment was absent we observed serum pgRNA at concentrations that correlated to HBV DNA and were approximately 2 log lower than HBV DNA. Importantly, we observe this trend in untreated subject samples from both chronic infections and throughout seroconversion during acute infection. Results demonstrate that the presence of pgRNA in serum is part of the HBV lifecycle; constant relative detection of pgRNA and HBV DNA in the serum is suggestive of a linked mechanism for egress for HBV DNA or pgRNA containing virions. This article is protected by copyright. All rights reserved. © 2018 by the American Association for the Study of Liver Diseases.

Spectro-refractometry of individual microscopic objects using swept-source quantitative phase imaging.

PubMed

Jung, Jae-Hwang; Jang, Jaeduck; Park, Yongkeun

2013-11-05

We present a novel spectroscopic quantitative phase imaging technique with a wavelength swept-source, referred to as swept-source diffraction phase microscopy (ssDPM), for quantifying the optical dispersion of microscopic individual samples. Employing the swept-source and the principle of common-path interferometry, ssDPM measures the multispectral full-field quantitative phase imaging and spectroscopic microrefractometry of transparent microscopic samples in the visible spectrum with a wavelength range of 450-750 nm and a spectral resolution of less than 8 nm. With unprecedented precision and sensitivity, we demonstrate the quantitative spectroscopic microrefractometry of individual polystyrene beads, 30% bovine serum albumin solution, and healthy human red blood cells.

Magnetically responsive polycaprolactone nanoparticles for progesterone screening in biological and environmental samples using gas chromatography.

PubMed

Es'haghi, Zarrin; Nezhadali, Azizollah; Khatibi, Aram-Dokht

2016-08-01

A new Fe3O4/poly(є-caprolactone) (PCL) magnetite nanocomposite was fabricated and used as a sorbent for magnetically mediated PCL microspheres solid-phase extraction (MM-PCL-SPE) followed by gas chromatography-flame ionization detection (GC-FID) for monitoring of progesterone (PGN) hormone in biological and environmental matrices, namely blood serum, tap water, urine, and hospital wastewater. The nanomagnetite core of the sorbent was synthesized by a co-precipitation method. Magnetic nanoparticles (MNPs) were then microencapsulated with PCL microspheres using emulsion polymerization. The nanocomposite was characterized by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). The magnetite sorbent can be effectively dispersed in aqueous solution and attracted to an external magnetic field. The MM-PCL-SPE process for PGN assay involved (a) dispersion of the sorbent in the donor phase aqueous solution with sonication, (b) exposure to a magnetic field to collect sorbent that had adsorbed the analyte, and (c) solvent desorption of extracted PGN for GC-FID analysis. The work demonstrates the usefulness of MM-PCL-SPE in the rapid and sensitive monitoring of trace amounts of PGN in real samples. The limit of detection (LOD) and limit of quantification (LOQ) were 1.00 and 3.30 ng/mL, respectively. The relative recoveries in real samples were adequate. Linearity was observed over a wide range of 2.2-10,000.0 ng/mL in aqueous media and urine and 0.01-70.0 μg/mL in blood serum. Graphical Abstract In this research new Fe3O4/poly(є-caprolactone) (PCL) magnetite microspheres were developed as an efficient sorbent for solid-phase extraction of progesterone hormone in biological and environmental matrices.

Evaluation of ammonia as diluent for serum sample preparation and determination of selenium by graphite furnace atomic absorption spectrometry*1

NASA Astrophysics Data System (ADS)

Hernández-Caraballo, Edwin A.; Burguera, Marcela; Burguera, José L.

2002-12-01

A method for the determination of total selenium in serum samples by graphite furnace atomic absorption spectrometry was evaluated. The method involved direct introduction of 1:5 diluted serum samples (1% v/v NH 4OH+0.05% w/v Triton X-100 ®) into transversely heated graphite tubes, and the use of 10 μg Pd+3 μg Mg(NO 3) 2 as chemical modifier. Optimization of the modifier mass and the atomization temperature was conducted by simultaneously varying such parameters and evaluating both the integrated absorbance and the peak height/peak area ratio. The latter allowed the selection of compromise conditions rendering good sensitivity and adequate analyte peak profiles. A characteristic mass of 49 pg and a detection limit (3s) of 6 μg 1 -1 Se, corresponding to 30 μg l -1 Se in the serum sample, were obtained. The analyte addition technique was used for calibration. The accuracy was assessed by the determination of total selenium in Seronorm™ Trace Elements Serum Batch 116 (Nycomed Pharma AS). The method was applied for the determination of total selenium in ten serum samples taken from individuals with no known physical affection. The selenium concentration ranged between 79 and 147 μg l -1, with a mean value of 114±22 μg l -1.

Potential of cancer screening with serum surface-enhanced Raman spectroscopy and a support vector machine

NASA Astrophysics Data System (ADS)

Li, S. X.; Zhang, Y. J.; Zeng, Q. Y.; Li, L. F.; Guo, Z. Y.; Liu, Z. M.; Xiong, H. L.; Liu, S. H.

2014-06-01

Cancer is the most common disease to threaten human health. The ability to screen individuals with malignant tumours with only a blood sample would be greatly advantageous to early diagnosis and intervention. This study explores the possibility of discriminating between cancer patients and normal subjects with serum surface-enhanced Raman spectroscopy (SERS) and a support vector machine (SVM) through a peripheral blood sample. A total of 130 blood samples were obtained from patients with liver cancer, colonic cancer, esophageal cancer, nasopharyngeal cancer, gastric cancer, as well as 113 blood samples from normal volunteers. Several diagnostic models were built with the serum SERS spectra using SVM and principal component analysis (PCA) techniques. The results show that a diagnostic accuracy of 85.5% is acquired with a PCA algorithm, while a diagnostic accuracy of 95.8% is obtained using radial basis function (RBF), PCA-SVM methods. The results prove that a RBF kernel PCA-SVM technique is superior to PCA and conventional SVM (C-SVM) algorithms in classification serum SERS spectra. The study demonstrates that serum SERS, in combination with SVM techniques, has great potential for screening cancerous patients with any solid malignant tumour through a peripheral blood sample.

Development and application of a fluorescence protein microarray for detecting serum alpha-fetoprotein in patients with hepatocellular carcinoma.

PubMed

Zhang, Aiying; Yin, Chengzeng; Wang, Zhenshun; Zhang, Yonghong; Zhao, Yuanshun; Li, Ang; Sun, Huanqin; Lin, Dongdong; Li, Ning

2016-12-01

Objective To develop a simple, effective, time-saving and low-cost fluorescence protein microarray method for detecting serum alpha-fetoprotein (AFP) in patients with hepatocellular carcinoma (HCC). Method Non-contact piezoelectric print techniques were applied to fluorescence protein microarray to reduce the cost of prey antibody. Serum samples from patients with HCC and healthy control subjects were collected and evaluated for the presence of AFP using a novel fluorescence protein microarray. To validate the fluorescence protein microarray, serum samples were tested for AFP using an enzyme-linked immunosorbent assay (ELISA). Results A total of 110 serum samples from patients with HCC ( n = 65) and healthy control subjects ( n = 45) were analysed. When the AFP cut-off value was set at 20 ng/ml, the fluorescence protein microarray had a sensitivity of 91.67% and a specificity of 93.24% for detecting serum AFP. Serum AFP quantified via fluorescence protein microarray had a similar diagnostic performance compared with ELISA in distinguishing patients with HCC from healthy control subjects (area under receiver operating characteristic curve: 0.906 for fluorescence protein microarray; 0.880 for ELISA). Conclusion A fluorescence protein microarray method was developed for detecting serum AFP in patients with HCC.

Development and application of a fluorescence protein microarray for detecting serum alpha-fetoprotein in patients with hepatocellular carcinoma

PubMed Central

Zhang, Aiying; Yin, Chengzeng; Wang, Zhenshun; Zhang, Yonghong; Zhao, Yuanshun; Li, Ang; Sun, Huanqin; Lin, Dongdong

2016-01-01

Objective To develop a simple, effective, time-saving and low-cost fluorescence protein microarray method for detecting serum alpha-fetoprotein (AFP) in patients with hepatocellular carcinoma (HCC). Method Non-contact piezoelectric print techniques were applied to fluorescence protein microarray to reduce the cost of prey antibody. Serum samples from patients with HCC and healthy control subjects were collected and evaluated for the presence of AFP using a novel fluorescence protein microarray. To validate the fluorescence protein microarray, serum samples were tested for AFP using an enzyme-linked immunosorbent assay (ELISA). Results A total of 110 serum samples from patients with HCC (n = 65) and healthy control subjects (n = 45) were analysed. When the AFP cut-off value was set at 20 ng/ml, the fluorescence protein microarray had a sensitivity of 91.67% and a specificity of 93.24% for detecting serum AFP. Serum AFP quantified via fluorescence protein microarray had a similar diagnostic performance compared with ELISA in distinguishing patients with HCC from healthy control subjects (area under receiver operating characteristic curve: 0.906 for fluorescence protein microarray; 0.880 for ELISA). Conclusion A fluorescence protein microarray method was developed for detecting serum AFP in patients with HCC. PMID:27885040

Stability of BDNF in Human Samples Stored Up to 6 Months and Correlations of Serum and EDTA-Plasma Concentrations.

PubMed

Polyakova, Maryna; Schlögl, Haiko; Sacher, Julia; Schmidt-Kassow, Maren; Kaiser, Jochen; Stumvoll, Michael; Kratzsch, Jürgen; Schroeter, Matthias L

2017-06-03

Brain-derived neurotrophic factor (BDNF), an important neural growth factor, has gained growing interest in neuroscience, but many influencing physiological and analytical aspects still remain unclear. In this study we assessed the impact of storage time at room temperature, repeated freeze/thaw cycles, and storage at -80 °C up to 6 months on serum and ethylenediaminetetraacetic acid (EDTA)-plasma BDNF. Furthermore, we assessed correlations of serum and plasma BDNF concentrations in two independent sets of samples. Coefficients of variations (CVs) for serum BDNF concentrations were significantly lower than CVs of plasma concentrations ( n = 245, p = 0.006). Mean serum and plasma concentrations at all analyzed time points remained within the acceptable change limit of the inter-assay precision as declared by the manufacturer. Serum and plasma BDNF concentrations correlated positively in both sets of samples and at all analyzed time points of the stability assessment ( r = 0.455 to r s = 0.596; p < 0.004). In summary, when considering the acceptable change limit, BDNF was stable in serum and in EDTA-plasma up to 6 months. Due to a higher reliability, we suggest favoring serum over EDTA-plasma for future experiments assessing peripheral BDNF concentrations.

Serum and saliva cortisol relations in adolescents during pregnancy and the early postpartum period.

PubMed

Dorn, L D; Susman, E J

1993-08-15

The purpose of this investigation was to examine: (1) relations between serum and saliva cortisol in adolescents in pregnancy and early postpartum and (2) short-term consistency of serum and saliva cortisol across three samples, 20 minutes apart, as well as the long-term consistency from pregnancy to early postpartum. Pregnant adolescents (n = 40), ages 14 to 19 years, were enrolled in this study. Subjects were seen at 20 weeks gestation or earlier (T1), 34-36 weeks gestation (T2), and 2-3 weeks postpartum (T3). Blood samples were drawn at T1 and T3, at 0, 20, and 40 minutes. Saliva samples were collected across the same 40-minute period at T1, T2, and T3. Spearman rho (rs) correlation coefficients between serum and saliva ranged from 0.72 to 0.77 (T1), and 0.42 to 0.60 (T3) (p < or = 0.05). Short-term consistency between serum cortisol samples was 0.86-0.97 at T1 and 0.60-0.82 at T3. Short-term consistency for saliva cortisol samples was 0.70-0.96 at T1, 0.91-0.95 at T2, and 0.64-0.89 at T3. Long-term consistency (T1 to T3) for serum and saliva cortisol was low. Individual differences as well as dramatic changes in the endocrine environment in pregnancy and the early postpartum period may explain the more moderate serum-saliva correlations in the postpartum period.

Determination of carbamazepine in serum and saliva samples by high performance liquid chromatography with ultraviolet detection.

PubMed

Dordević, Snezana; Kilibarda, Vesna; Stojanović, Tomislav

2009-05-01

Carbamazepine is antiepileptic drug widely used for the treatment of epilepsy. Due to low therapeutic index of carbamazepine there is a need for routine measuring its concentrations in biological fluids. The aim of the study was to describe a method for concomitant determination of carbamazepine in the serum and saliva. Separation of the drug from matrix is achieved by reversed-phase chromatography on a C18 column, with a mobile phase of methanol-water-acetic acid (65:34:1) at a flow-rate of 1.0 ml/min. Detection was effected by ultra-violet absorption at 285 nm. The total run time was 5 min. Samples were prepared by alkaline extraction (pH 10) using chlorophorm. Calibration curves were in the range 0.1-5 microg/mL for serum and saliva samples. Mean recoveries of spiked serum and saliva were 97.59 and 92.30%, respectively. Limits of detection (LOD) of carbamazepine in serum and saliva were 0.166 and 0.178 microg/mL, respectively. Limits of quantification (LOQ) in the serum and saliva were 0.237 and 0.226 microg/mL, respectively. The method precision was carried out with coefficient of variation of 2.10% and 4.03% for the serum and saliva, respectively. The obtained data showed that there was a strong correlation between saliva and serum concentrations (r = 0.9481, p < 0.001). The method described here is rapid, precise, accurate and simple, and can be used for quantitative determination of carbamazepine in human serum and saliva after therapy applying. Saliva samples could be used as an alternative matrix for therapeutic drug monitoring of this antiepileptic drug.

In vitro production of GHB in blood and serum samples under various storage conditions.

PubMed

Zörntlein, S W; Kopp, A; Becker, J; Kaufmann, T J; Röhrich, J; Urban, R

2012-01-10

The in vitro production of GHB was observed in freshly collected, untreated whole blood samples using glass BD-Vacutainers and polypropylene S-monovettes. GHB concentrations were determined daily over a period of one week and after 3, 6 and 9 weeks again. Furthermore, the GHB concentration in 40 untreated random whole blood samples stored at 4°C for a longer period of time (10 samples 12 month, 10 samples 24 month and 20 samples 36 month) was also determined. For comparison, the in vitro production of GHB in freshly collected and prepared serum samples was observed. GHB serum concentrations were determined three times over a period of one week and once again after six weeks. Sample preparation was performed by means of methanolic extraction following the precipitation of whole blood and serum samples. A methanolic standard calibration was done in a low range of 0.005-0.1 μg/mL (LOD: 0.004, LLOQ: 0.013). For quantification a spiked blood bank serum with a determined GHB concentration of 0.09 μg/mL was used. Corrected calibrations in the range of 0.09-5.09 μg/mL were used (LOD: 0.08 μg/mL, LLOQ: 0.30 μg/mL), recovery: 91.3% (high level: 4.09 μg/mL) 50.5% (low level: 0.19 μg/mL). Relevant elevation of GHB was observed in all whole blood samples stored in liquid form (4°C or room temperature). In two of the 40 whole blood samples stored over a longer period of time at 4°C, GHB concentrations in the range of 13 μg/mL were even determined. These findings constitute grounds for caution. Even a GHB cut-off level of 5 μg/mL cannot be considered as "absolutely positive" proof of a case of exogenous administration, at least in untreated liquid blood samples in long time storage. However, no significant elevations of GHB were otherwise observed in any of the serum samples independently of storage temperature nor in the whole blood samples that were frozen for storage. The results suggest that the cut-off for exogenous GHB of 5 μg/mL could be lowered significantly, with the consequence of winning valuable time for the potential victim, but only if serum is collected for GHB determination or if the whole blood sample is frozen immediately after collection and the procedure well documented. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Whole Blood Reveals More Metabolic Detail of the Human Metabolome than Serum as Measured by 1H-NMR Spectroscopy: Implications for Sepsis Metabolomics

PubMed Central

Stringer, Kathleen A.; Younger, John G.; McHugh, Cora; Yeomans, Larisa; Finkel, Michael A.; Puskarich, Michael A.; Jones, Alan E.; Trexel, Julie; Karnovsky, Alla

2015-01-01

Serum is a common sample of convenience for metabolomics studies. Its processing time can be lengthy and may result in the loss of metabolites including those of red blood cells (RBC). Unlike serum, whole blood (WB) is quickly processed, minimizing the influence of variable hemolysis while including RBC metabolites. To determine differences between serum and WB metabolomes, both sample types, collected from healthy volunteers, were assayed by 1H-NMR spectroscopy. A total of 34 and 50 aqueous metabolites were quantified from serum and WB, respectively. Free hemoglobin (Hgb) levels in serum were measured and the correlation between Hgb and metabolite concentrations was determined. All metabolites detected in serum were at higher concentrations in WB with the exception of acetoacetate and propylene glycol. The 18 unique metabolites of WB included adenosine, AMP, ADP and ATP, which are associated with RBC metabolism. The use of serum results in the underrepresentation of a number of metabolic pathways including branched chain amino acid degradation and glycolysis and gluconeogenesis. The range of free Hgb in serum was 0.03-0.01 g/dL and 8 metabolites were associated (p ≤ 0.05) with free Hgb. The range of free Hgb in serum samples from 18 sepsis patients was 0.02-0.46 g/dL. WB and serum have unique aqueous metabolite profiles but the use of serum may introduce potential pathway bias. Use of WB for metabolomics may be particularly important for studies in diseases like sepsis in which RBC metabolism is altered and mechanical and sepsis-induced hemolysis contributes to variance in the metabolome. PMID:26009817

Antibodies against MERS coronavirus in dromedary camels, United Arab Emirates, 2003 and 2013.

PubMed

Meyer, Benjamin; Müller, Marcel A; Corman, Victor M; Reusken, Chantal B E M; Ritz, Daniel; Godeke, Gert-Jan; Lattwein, Erik; Kallies, Stephan; Siemens, Artem; van Beek, Janko; Drexler, Jan F; Muth, Doreen; Bosch, Berend-Jan; Wernery, Ulrich; Koopmans, Marion P G; Wernery, Renate; Drosten, Christian

2014-04-01

Middle East respiratory syndrome coronavirus (MERS-CoV) has caused an ongoing outbreak of severe acute respiratory tract infection in humans in the Arabian Peninsula since 2012. Dromedary camels have been implicated as possible viral reservoirs. We used serologic assays to analyze 651 dromedary camel serum samples from the United Arab Emirates; 151 of 651 samples were obtained in 2003, well before onset of the current epidemic, and 500 serum samples were obtained in 2013. Recombinant spike protein-specific immunofluorescence and virus neutralization tests enabled clear discrimination between MERS-CoV and bovine CoV infections. Most (632/651, 97.1%) camels had antibodies against MERS-CoV. This result included all 151 serum samples obtained in 2003. Most (389/651, 59.8%) serum samples had MERS-CoV-neutralizing antibody titers >1,280. Dromedary camels from the United Arab Emirates were infected at high rates with MERS-CoV or a closely related, probably conspecific, virus long before the first human MERS cases.

Use of a commercial ELISA for the detection of measles-specific immunoglobulin G (IgG) in dried blood spots collected from children living in low-resource settings.

PubMed

Colson, K Ellicott; Potter, Alan; Conde-Glez, Carlos; Hernandez, Bernardo; Ríos-Zertuche, Diego; Zúñiga-Brenes, Paola; Iriarte, Emma; Mokdad, Ali H

2015-09-01

Seroepidemiological monitoring of population immunity to vaccine-preventable diseases is critical to prevent future outbreaks. Dried blood spots (DBS), drops of capillary blood dried on filter paper, are an affordable, minimally invasive alternative to venipuncture for collecting blood in field settings. However, few proven methods exist to analyze DBS for the presence of protective antibodies. This study validates a novel technique for measuring measles-specific immunoglobulin G (IgG) in capillary DBS using a commercial ELISA. The predictive performance of a new method for analyzing DBS was tested by comparing matched serum and DBS samples from 50 children. The accuracy, precision, and reliability of the procedure were evaluated, and the optimal cut points to classify positive and negative samples were determined. The method was then applied to 1,588 DBS collected during a large survey of children in Mexico and Nicaragua. Measles-specific IgG in serum samples were 62% negative, 10% equivocal, and 28% positive. In comparisons with matched serum, DBS results were 100% sensitive and 96 · 8% specific, and agreed in 46 of 50 (92%) cases. The inter-assay and intra-assay coefficients of variation from kit-provided controls were greater than desired (24.8% and 8.4%, respectively). However, in predictive simulations the average misclassification was only 3.9%. Procedures were found to be acceptable to surveyors and participants. Analyzing DBS collected in low-resources settings is a feasible and accurate means of measuring population immunity to measles and should be used to generate objective measures of health status and health system performance. © 2015 Wiley Periodicals, Inc.

Determination of chlormequat in pig serum and sow milk by LC-MS/MS.

PubMed

Poulsen, M E; Christensen, H B; Sørensen, M T; Leffers, H; Andersen, J H

2007-11-01

Chlormequat is a plant growth regulator widely used on cereals, and there is general concern that it may impair human fertility. A LC-MS/MS method for the analysis of chlormequat in milk and serum was developed and validated in connection with an investigation on the effect of chlormequat on pig reproduction. Validation of the method was based on recovery tests at three spiking levels, determined as double determinations and repeated at least four times. Samples were extracted with methanol-water-acetic acid, centrifuged, filtrated and determined by LC-MS/MS. The mean recoveries were in the range 80-110%, and the LOD was 0.2 ng/g for serum and 0.3 ng/g for milk. The values for repeatability and reproducibility were within 2/3 of the limits given by the Horwitz equation. Samples of pig serum (59) and sow milk (27) were analyzed using the method. Chlormequat was determined in four milk samples in the range of 0.4 ng/g to 1.2 ng/g and in all serum samples in the range of 0.2 ng/g-4.0 ng/g.

EDRN Breast and Ovary Cancer CVC, Study 4: Phase 3 Validation of Ovarian Cancer Serum Markers in Preclinical WHI Samples — EDRN Public Portal

Cancer.gov

The WHI offers an opportunity to evaluate ovarian cancer markers and screening decision rules developed and validated in EDRN CVC Studies 2 and 3 in women who were not being screened. It is particularly well suited to validation of risk markers, since many serum samples were drawn well before clinical diagnosis of cancer in the WHI cohorts. A strategy is needed to identify from among the general population of women over the age of 50 those at high-risk for a diagnosis of ovarian/fallopian tube cancer so that they can be referred for appropriate surveillance, imaging or surgical consult. Tools to identify high-risk women will be investigated including serum markers CA125, HE4, MSLN, and MMP7 and epidemiologic risk factors. We will optimize decision rules using stored serum samples from the WHI OS and conduct a simulated prospective validation using stored serum samples from the WHI CT. Decision rules to select women for ovarian cancer screening will be investigated as well as decision rules for use in ovarian cancer screening.

Detection of antibodies against hepatitis A in blood spots dried on filter paper. Is this a reliable method for epidemiological studies?

PubMed Central

Gil, A.; González, A.; Dal-Ré, R.; Dominguez, V.; Astasio, P.; Aguilar, L.

1997-01-01

Diluted dried blood drops on filter paper were compared with serum samples as a specimen source for qualitative anti-HAV antibody determination by ELISA. A total of 298 serum samples and dried blood drops were collected from a population of healthy adolescents (15.3 +/- 1.2 years old). The prevalence of anti-HAV antibody obtained by testing serum samples was 7.7% (95% CI:4.8 10.1). Compared with serum sampling the sensitivity and specificity of diluted dried blood drops were 91.3 and 99.3%. The positive and negative predictive values were 91.3 and 99.3%, respectively, and the likelihood ratios of positive and negative results were 91 and 0.09. It is proposed that this test represents a reliable procedure for anti-HAV antibody testing. PMID:9129596

Bulk- and surface-modified combinable PDMS capillary sensor array as an easy-to-use sensing device with enhanced sensitivity to elevated concentrations of multiple serum sample components.

PubMed

Fujii, Yuji; Henares, Terence G; Kawamura, Kunio; Endo, Tatsuro; Hisamoto, Hideaki

2012-04-21

To enhance sensitivity and facilitate easy sample introduction into a combinable poly(dimethylsiloxane) (PDMS) capillary (CPC) sensor array, PDMS was modified in bulk and on its surface to prepare "black" PDMS coated with a silver layer and self-assembled monolayer (SAM). India ink, a traditional Japanese black ink, was added to the PDMS pre-polymer for bulk modification. The surface was modified by a silver mirror reaction followed by SAM formation using cysteine. These modifications enhanced the fluorescence signals by reflecting them from the surface and reducing background interference. A decrease in the water contact angle led to enhanced sensitivity and easy sample introduction. Furthermore, a CPC sensor array for multiplex detection of serum sample components was prepared that could quantify the analytes glucose, potassium, and alkaline phosphatase (ALP). When serum samples were introduced by capillary action, the CPC sensor array showed fluorescence responses for each analyte and successfully identified the components with elevated concentrations in the serum samples.

Gastrointestinal parasites of feral cats from Christmas Island.

PubMed

Adams, P J; Elliot, A D; Algar, D; Brazell, R I

2008-01-01

To investigate the gastrointestinal parasites present in feral cats on Christmas Island, with particular interest in the protozoan parasite Toxoplasma gondii. Faecal and serum samples were collected from 28 and 25 cats respectively that were trapped as part of an ongoing eradication program being run on Christmas Island by the Department of Environment and Conservation. Faecal samples were screened microscopically for helminth and protozoan parasites. Serum samples were screened for antibodies to T gondii using a commercial indirect immunofluorescence assay (IFA) and a latex agglutination test (LAT). The most common helminth parasites detected were Toxocara cati (present in 15 of 28 faecal samples), Strongyloides sp (13/28), Aelurostrongylus abstrusus, (7/28), an unidentified capillarid (6/28) and Ancylostoma sp (4/28). Based on serology, T gondii was the most common parasite detected (protozoan or otherwise) with antibodies detected in 24 serum samples by IFA and 23 serum samples by LAT. Cats on Christmas Island harbour many of the helminth and protozoan parasites reported from feral cats elsewhere in Australia. The high seroprevalence of T gondii in these cats indicates a high level of exposure to the parasite in this environment.

Integration of a highly ordered gold nanowires array with glucose oxidase for ultra-sensitive glucose detection.

PubMed

Cui, Jiewu; Adeloju, Samuel B; Wu, Yucheng

2014-01-27

A highly sensitive amperometric nanobiosensor has been developed by integration of glucose oxidase (GO(x)) with a gold nanowires array (AuNWA) by cross-linking with a mixture of glutaraldehyde (GLA) and bovine serum albumin (BSA). An initial investigation of the morphology of the synthesized AuNWA by field emission scanning electron microscopy (FESEM) and field emission transmission electron microscopy (FETEM) revealed that the nanowires array was highly ordered with rough surface, and the electrochemical features of the AuNWA with/without modification were also investigated. The integrated AuNWA-BSA-GLA-GO(x) nanobiosensor with Nafion membrane gave a very high sensitivity of 298.2 μA cm(-2) mM(-1) for amperometric detection of glucose, while also achieving a low detection limit of 0.1 μM, and a wide linear range of 5-6000 μM. Furthermore, the nanobiosensor exhibited excellent anti-interference ability towards uric acid (UA) and ascorbic acid (AA) with the aid of Nafion membrane, and the results obtained for the analysis of human blood serum indicated that the device is capable of glucose detection in real samples. Copyright © 2013 Elsevier B.V. All rights reserved.

Comparison of Polyfluoroalkyl Compound Concentrations in Maternal Serum and Amniotic Fluid: A Pilot Study

PubMed Central

Stein, Cheryl R.; Wolff, Mary S.; Calafat, Antonia M.; Kato, Kayoko; Engel, Stephanie M.

2012-01-01

The extent to which polyfluoroalkyl compounds (PFCs) are detectable in amniotic fluid is unknown. Using paired samples from 28 women, we compared the concentration of 8 PFCs measured in serum, the standard matrix for assessing human exposure, amniotic fluid from routine amniocentesis, and urine. Perfluorooctanoate (PFOA), perfluorononanoate (PFNA), perfluorooctane sulfonate (PFOS), and perfluorohexane sulfonate (PFHxS) were detected in all maternal serum samples. The number of amniotic fluid samples with detectable concentrations differed by PFC (PFOA n=24; PFNA n=10; PFOS n=9; PFHxS n=4). The correlation coefficient between maternal serum and amniotic PFC levels varied considerably by PFC (PFOA ρ=0.64, p<0.001; PFNA ρ=0.05, p=0.9; PFOS ρ=0.76, p=0.01; PFHxS ρ=0.80, p=0.2). Using linear regression, PFOA appeared to be commonly detected in amniotic fluid if the serum concentration exceeded approximately 1.5 ng/mL whereas PFOS was rarely detected in amniotic fluid until the serum concentration was about 5.5 ng/mL. No PFCs were detected in urine. PMID:22613200

Feasibility Study of an Optical Caustic Plasmonic Light Scattering Sensor for Human Serum Anti-Dengue Protein E Antibody Detection

PubMed Central

García, Antonio A.; Pirez-Gomez, Miguel A.; Pech-Pacheco, José L.; Mendez-Galvan, Jorge F.; Machain-Williams, Carlos; Talavera-Aguilar, Lourdes; Espinosa-Carrillo, José H.; Duarte-Villaseñor, Miriam M.; Be-Ortiz, Christian; Espinosa-de los Monteros, Luz E.; Castillo-Pacheco, Ariel; Garcia-Rejon, Julian E.

2017-01-01

Antibody detection and accurate diagnosis of tropical diseases is essential to help prevent the spread of disease. However, most detection methods lack cost-effectiveness and field portability, which are essential features for achieving diagnosis in a timely manner. To address this, 3D-printed oblate spheroid sample chambers were fabricated to measure green light scattering of gold nanoparticles using an optical caustic focus to detect antibodies. Scattering signals of 20–200 nm gold nanoparticles using a green laser were compared to green light emitting diode (LED) light source signals and to Mie theory. The change in signal from 60 to 120 nm decreased in the order of Mie Theory > optical caustic scattering > 90° scattering. These results suggested that conjugating 60 nm gold nanoparticles and using an optical caustic system to detect plasmonic light scattering, would result in a sensitive test for detecting human antibodies in serum. Therefore, we studied the light scattering response of conjugated gold nanoparticles exposed to different concentrations of anti-protein E antibody, and a feasibility study of 10 human serum samples using dot blot and a handheld optical caustic-based sensor device. The overall agreement between detection methods suggests that the new sensor concept shows promise to detect gold nanoparticle aggregation in a homogeneous assay. Further testing and protocol optimization is needed to draw conclusions on the positive and negative predictive values for this new testing system. PMID:28817080

Development and validation of an automated liquid-liquid extraction GC/MS method for the determination of THC, 11-OH-THC, and free THC-carboxylic acid (THC-COOH) from blood serum.

PubMed

Purschke, Kirsten; Heinl, Sonja; Lerch, Oliver; Erdmann, Freidoon; Veit, Florian

2016-06-01

The analysis of Δ(9)-tetrahydrocannabinol (THC) and its metabolites 11-hydroxy-Δ(9)-tetrahydrocannabinol (11-OH-THC), and 11-nor-9-carboxy-Δ(9)-tetrahydrocannabinol (THC-COOH) from blood serum is a routine task in forensic toxicology laboratories. For examination of consumption habits, the concentration of the phase I metabolite THC-COOH is used. Recommendations for interpretation of analysis values in medical-psychological assessments (regranting of driver's licenses, Germany) include threshold values for the free, unconjugated THC-COOH. Using a fully automated two-step liquid-liquid extraction, THC, 11-OH-THC, and free, unconjugated THC-COOH were extracted from blood serum, silylated with N-methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA), and analyzed by GC/MS. The automation was carried out by an x-y-z sample robot equipped with modules for shaking, centrifugation, and solvent evaporation. This method was based on a previously developed manual sample preparation method. Validation guidelines of the Society of Toxicological and Forensic Chemistry (GTFCh) were fulfilled for both methods, at which the focus of this article is the automated one. Limits of detection and quantification for THC were 0.3 and 0.6 μg/L, for 11-OH-THC were 0.1 and 0.8 μg/L, and for THC-COOH were 0.3 and 1.1 μg/L, when extracting only 0.5 mL of blood serum. Therefore, the required limit of quantification for THC of 1 μg/L in driving under the influence of cannabis cases in Germany (and other countries) can be reached and the method can be employed in that context. Real and external control samples were analyzed, and a round robin test was passed successfully. To date, the method is employed in the Institute of Legal Medicine in Giessen, Germany, in daily routine. Automation helps in avoiding errors during sample preparation and reduces the workload of the laboratory personnel. Due to its flexibility, the analysis system can be employed for other liquid-liquid extractions as well. To the best of our knowledge, this is the first publication on a comprehensively automated classical liquid-liquid extraction workflow in the field of forensic toxicological analysis. Graphical abstract GC/MS with MPS Dual Head at the Institute of Legal Medicine, Giessen, Germany. Modules from left to right: (quick) Mix (for LLE), wash station, tray 1 (vials for extracts), solvent reservoir, (m) VAP (for extract evaporation), Solvent Filling Station (solvent supply), cooled tray 2 (vials for serum samples), and centrifuge (for phase separation).

The caffeine breath test and caffeine urinary metabolite ratios in the Michigan cohort exposed to polybrominated biphenyls: A preliminary study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lambert, G.H.; Schoeller, D.A.; Kotake, A.N.

1990-11-01

A field biochemical epidemiology study was conducted using the Michigan cohort consisting of 51 rural residents exposed to polybrominated biphenyls (PBB). The study had three major objectives: (a) to determine the serum half-life of the major PBB congener, hexabromobiphenyl (HBB), in the human, (b) to determine if the PBB-exposed subjects had elevated cytochrome P-450I function as determined by the caffeine breath test (CBT) and the caffeine urinary metabolite ratio (CMR), and (c) to determine the applicability of the CBT and CMR in field studies. PBB serum levels were detected in 36 of the 51 PBB-exposed subjects. The serum half-life ofmore » HBB was determined by comparing the current serum HBB values to the subject's previous serum values obtained 5 to 8 years earlier. The median HBB half-life was 12 years (range 4-97 years). The CBT and CMR were elevated in the subjects exposed to PBBs as compared to the values obtained from urban nonsmokers and were similar to those found in adults who smoke. A gender effect was seen in the PBB-exposed subjects. There was a correlation between the CBT and the HBB serum values but not between CMR and HBB serum values. The CBT and CMR were easily conducted in the field and appear to be useful metabolic probes of cytochrome P-450I activity in human environmental toxicology.« less

Development and evaluation of an enzyme-labeled antibody test for the rapid detection of hog cholera antibodies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Saunders, G.C.

1977-01-01

A rapid enzyme-labeled antibody (ELA) microtechnique for the screening of swine for hog cholera antibodies was developed and evaluated with a blind study, using a 640-sample hog cholera serum bank. The total time to run a group of 22 samples was approximately 1 hour. The ELA test results correlated >99% with hog cholera serum-neutralization test results on the same serums. Test results also indicated that the ELA test shares with the hog cholera serum-neutralization test the problem of cross reactions between the antibodies of hog cholera and bovine viral diarrhea.

Determination of dapsone in serum and saliva using reversed-phase high-performance liquid chromatography with ultraviolet or electrochemical detection.

PubMed

Moncrieff, J

1994-03-18

A simple, extractionless method for the determination of dapsone in serum and saliva is described. Reversed-phase high-performance liquid chromatography is used with UV detection at 295 nm or electrochemical detection at 0.7 V. Diazoxide in buffer is the internal standard for UV detection and practolol for electrochemical detection. Sample preparation is minimal with protein precipitation of serum samples whilst saliva samples are simply diluted with addition of an internal standard. Low-level serum and saliva samples are front-cut on-line with a 3 cm laboratory-made precolumn in the loop position on a standard Valco injection valve. Isocratic separation is achieved on a 250 mm x 4.6 mm I.D. stainless-steel Spherisorb S5 ODS-1 column. The mobile phase for high levels of dapsone is acetonitrile-elution buffer (12:88, v/v) at 2 ml/min and a column temperature of 40 degrees C for both serum and saliva separations. For the low-level assays using electrochemical detection and solid-phase clean-up, the mobile phase is acetonitrile-methanol-elution buffer (9:4:87, v/v/v). The UV and electrochemical detection limits are 25 ng/ml and 200 pg/ml, respectively, in both serum and saliva. This simple method is applicable to the routine monitoring of dapsone levels in serum from leprotic patients and electrochemical detection gives a simple, reliable method for the monitoring of trough values in subjects on anti-malarial prophylaxis.

Effectiveness analyses may underestimate protection of infants after group C meningococcal immunization.

PubMed

Vu, David M; Kelly, Dominic; Heath, Paul T; McCarthy, Noel D; Pollard, Andrew J; Granoff, Dan M

2006-07-15

Group C meningococcal conjugate-vaccine effectiveness in the United Kingdom declines from ~90% in the first year to 0% between 1 and 4 years after immunization in infants immunized at 2, 3, and 4 months of age and to 61% in toddlers given a single dose. Confidence intervals are wide, and the extent of protection is uncertain. Serum samples were obtained from children 3-5 years of age who were participants in a preschool booster-vaccine trial. Serum bactericidal activity was measured with human complement. Group C anticapsular antibody concentrations were measured by a radioantigen binding assay. Passive protection was analyzed in an infant rat bacteremia model. Serum samples from UK children who had been immunized 2-3 years earlier as infants or toddlers had higher levels of radioantigen binding, bactericidal activity, and passive protection than did historical control serum samples from unimmunized children (P<.05). A higher proportion of children immunized as infants had serum bactericidal activity titers > or =1 : 4 (considered to be protective) than those immunized as toddlers (61% vs. 24%; P<.01), but there were no significant differences in the proportion of serum samples conferring passive protection (50% and 41%, respectively; P=.4). We found no evidence of lower immunity in children immunized as infants than as toddlers. On the basis of serum bactericidal activity and/or passive protection, 40%-50% of both age groups are protected at 2-3 years after immunization, which was significantly greater than in unimmunized historical controls (<5%).

Daily rhythm of salivary and serum urea concentration in sheep

PubMed Central

Piccione, Giuseppe; Foà, Augusto; Bertolucci, Cristiano; Caola, Giovanni

2006-01-01

Background In domestic animals many biochemical and physiological processes exhibit daily rhythmicity. The aim of the present study was to investigate the rhythmic pattern of salivary and serum urea concentrations in sheep. Methods Six 3-year-old female sheep kept in the same environmental conditions were used. Sheep were sampled at 4 hour intervals for 48 consecutive hours starting at 08:00 of the first day and finishing at 04:00 of the second day. Blood samples were collected via intravenous cannulae inserted into the jugular vein; saliva samples were collected through a specific tube, the "Salivette". Salivary and serum urea concentrations were assayed by means of UV spectrophotometer. ANOVA was used to determine significant differences. The single Cosinor procedure was applied to the results showing significant differences over time. Results ANOVA showed a significant effect of time on salivary and serum urea concentrations. Serum and salivary urea peaked during the light phase. In the dark phase serum and salivary urea concentrations decreased, and the diurnal trough occurred at midnight. Cosinor analysis showed diurnal acrophases for salivary and serum urea concentrations. Daily mean levels were significantly higher in the serum than in the saliva. Conclusion In sheep both salivary and serum urea concentrations showed daily fluctuations. Urea is synthesized in the liver and its production is strongly influenced by food intake. Future investigation should clarify whether daily urea rhythms in sheep are endogenous or are simply the result of the temporal administration of food. PMID:17123442

Significance of serum and bile tumor markers in the diagnostic approach of patients with malignant pancreatobiliary disease.

PubMed

Natsios, Athanasios; Vezakis, Antonios; Kaparos, Georgios; Fragulidis, Georgios; Karakostas, Nikolaos; Kouskouni, Evangelia; Logothetis, Emmanouil; Polydorou, Andreas

2015-01-01

Serum and bile tumor markers are under intense scrutiny for the diagnosis of malignant disease. The purpose of our study was to report the usefulness of serum and bile tumor markers for the discrimination between benign and malignant pancreatobiliary diseases. Between March 2010 and May 2013, 95 patients with obstructive jaundice or history of biliary obstruction, were included in the study. During ERCP, bile samples were obtained for measurement of tumor markers CEA, CA19- 9, CA125, CA72-4 and CA242. Serum samples were taken before ERCP for the same measurements. The patients were divided into two groups: patients with malignant disease and patients with benign disease. Serum tumor marker levels were significantly higher in patients with malignant disease. Serum CA242 and CA19-9 exhibited the highest diagnostic accuracy (76.8% and 73.7%, respectively). CA125 and CA72-4 levels in bile samples were significantly higher in patients with malignant disease. Bile CA125, CEA and CA72-4 achieved the best diagnostic accuracy (69, 65 and 65), respectively). The combined detection of CA19-9, CA242 in serum and CA125, CA72-4 in bile along with total bilirubin levels, showed the best diagnostic accuracy (81%). Serum and bile tumor markers, when studied alone, lack the diagnostic yield to discriminate benign from malignant pancreatobiliary diseases. In cases of diagnostic dilemmas the combination of serum and bile markers might be helpful.

Hematologic and serum biochemistry reference values in wild-caught white-footed tamarins (Saguinus leucopus) housed in captivity.

PubMed

Fox, Maureen; Brieva, Claudia; Moreno, Carlos; MacWilliams, Peter; Thomas, Chet

2008-12-01

The white-footed tamarin (Saguinus leucopus) is an endangered primate that lives in a small forest corridor in northern Colombia, South America. Hematologic and serum biochemistry reference values are important tools in evaluating the health of the white-footed tamarin and serve to promote the survival of this species. The purpose of this study was to establish diagnostically important hematologic and serum biochemistry reference values for healthy white-footed tamarins and to determine whether there was significant variation with respect to age class (juveniles, adults), gender, and housing facility. Blood samples were collected for hematologic and serum biochemistry measurements from 29 wild-caught captive tamarins during February and April 2005, housed at three different facilities in central Colombia. Hematology and serum biochemistry values were similar in adults and juveniles. Female white-footed tamarins had absolute reticulocyte counts that were lower than those of male tamarins and males had lower serum chloride concentrations than female tamarins. Mean values for hemoglobin, mean cell hemoglobin, serum total protein, albumin, glucose, and alkaline phosphatase varied among the three housing facilities. Twenty-two of the 29 tamarins sampled were microfilaria-positive and had significantly higher mean serum alkaline phosphatase concentrations. Among the 29 tamarins sampled in this study, serum values for mean alkaline phosphatase and creatine kinase concentrations were higher than reported values for other mammals. The reference intervals determined in this study were comparable to normal ranges reported for other members of the family Callitrichidae.

Serum Levels of Migration Inhibitory Factor (MIF) and In Situ Expression of MIF and Its Receptor CD74 in Lepromatous Leprosy Patients: A Preliminary Report

PubMed Central

Martinez-Guzman, Marco Alonso; Alvarado-Navarro, Anabell; Delgado-Rizo, Vidal; Garcia-Orozco, Alejandra; Mayorga-Rodríguez, Jorge Arturo; Pereira-Suarez, Ana Laura; Fafutis-Morris, Mary

2018-01-01

Leprosy is a chronic disease caused by Mycobacterium leprae that affects the skin and peripheral nerves. It may present as one of two distinct poles: the self-limiting tuberculoid leprosy and the highly infectious lepromatous leprosy (LL) characterized by M. leprae-specific absence of cellular immune response. The pro-inflammatory cytokine macrophage migration inhibitory factor (MIF) enhance the bactericide activities of macrophages after interaction with its receptor, CD74. Importantly, MIF also possesses chemoattractant properties, and it is a key factor in situ for the activation of macrophages and in blood to promote leukocytes migration. MIF-mediated activation of macrophages is a key process for the elimination of pathogens such as Mycobacterium tuberculosis; however, its participation for the clearance of M. leprae is unclear. The aim of this study was to evaluate the serum levels of MIF as well as MIF and CD74 expression in skin lesions of LL and compare it with healthy skin (HSk) taken from subjects attending to dermatological consult. Samples of serum and skin biopsies were taken from 39 LL patients and compared with 36 serum samples of healthy subjects (HS) and 10 biopsies of HSk. Serum samples were analyzed by ELISA and skin biopsies by immunohistochemistry (IHC). IHC smears were observed in 12 100× microscopic fields, in which percentage of stained cells and staining intensity were evaluated. Both variables were used to calculate a semi-quantitative expression score that ranged from 0 to 3+. We found no differences in MIF levels between LL patients and HS in sera. In addition, MIF was observed in over 75% of cells with high intensity in the skin of patients and HSk. Although we found no differences in MIF expression between the groups, a CD74 score statistically higher was found in LL skin than HSk (p < 0.001); this was the result of a higher percentage of cells positive for CD74 (p < 0.001). As a conclusion, we found that CD74-positive cells are intensely recruited to the skin with LL lesions. In this manner, MIF signaling may be enhanced in the skin of LL patients due to increased expression of its receptor, but further studies are required. PMID:29487601

Parathyroid hormone related protein concentration in human serum and CSF correlates with age.

PubMed

Kushnir, Mark M; Peterson, Lisa K; Strathmann, Frederick G

2018-02-01

Parathyroid Hormone-Related Protein (PTHrP) is involved in intracellular calcium (Ca) regulation, and has been demonstrated to participate in regulation of Ca in brain cells, activation of neurons, and modulation of pain. However, there are conflicting reports regarding the presence of PTHrP in CSF. PTHrP and Ca were quantified in paired CSF and serum samples using mass spectrometry-based methods. Associations between PTHrP and Ca concentrations with age, sex and concentrations of nine CSF diagnostic markers in a set of 140 paired serum and CSF patient samples were evaluated. The observed median PTHrP concentration in CSF was 51 times higher than in serum; the median concentration of Ca in CSF was 1.8 times lower than in serum. We observed positive correlation between concentrations of PTHrP in CSF and serum (p=0.013). Distribution of PTHrP concentrations in serum was associated with age (p=0.0068) and the concentrations were higher in women. In samples with serum calcium concentrations within the reference intervals (n=118), central 95% distribution of concentrations for Ca-CSF, PTHrP-serum and PTHrP-CSF were 5.4 (4.5-6.1) mg/dL, 1.2 (0.5-2.5) pmol/L, 62 (22-125) pmol/L, respectively. Our data demonstrate that PTHrP is a normal constituent of human CSF with median concentrations 51 fold higher than in serum. Elevated serum PTHrP concentrations were positively correlated with age and significantly higher in women. Our data suggest that CSF could be a significant source of circulating PTHrP. Copyright © 2017 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Comparison of Serum Protein Electrophoresis Values in Wild and Captive Whooping Cranes ( Grus americana ).

PubMed

Hausmann, Jennifer C; Cray, Carolyn; Hartup, Barry K

2015-09-01

Protein electrophoresis of serum samples from endangered, wild whooping cranes ( Grus americana ) was performed to help assess the health of the only self-sustaining, migratory population in North America. Serum samples from wild adult cranes (n = 22) were taken at Aransas National Wildlife Refuge, Texas, USA during winter. Wild juvenile cranes (n = 26) were sampled at Wood Buffalo National Park, Northwest Territories, Canada, in midsummer. All captive crane samples were acquired from the International Crane Foundation, Baraboo, WI, USA. Captive adult cranes (n = 30) were sampled during annual examinations, and archived serum samples from captive juvenile cranes (n = 19) were selected to match the estimated age of wild juveniles. Wild juveniles had significantly lower concentrations of all protein fractions than wild adults, except for prealbumin and γ globulins. All protein fraction concentrations for wild juveniles were significantly lower compared with captive juveniles, except for prealbumin and γ globulins, which were higher. Wild adults had significantly greater γ globulin concentrations than captive adults. Captive juveniles had significantly lower prealbumin and albumin concentrations and albumin : globulin ratios than captive adults. The higher γ globulin concentrations in wild versus captive cranes are likely because of increased antigenic exposure and immune stimulation. Protein fraction concentrations vary significantly with age and natural history in this species. Reference intervals for serum protein electrophoresis results from captive adult whooping cranes are provided in this study.

Detection of EBV-DNA in serum samples of an immunosuppressed child during a three years follow-up: association of clinical and PCR data with active infection.

PubMed

Okay, Thelma Suely; Del Negro, Gilda Maria Barbaro; Yamamoto, Lídia; Raiz Júnior, Roberto

2005-01-01

Twenty-four whole blood and serum samples were drawn from an eight year-old heart transplant child during a 36 months follow-up. EBV serology was positive for VCA-IgM and IgG, and negative for EBNA-IgG at the age of five years old when the child presented with signs and symptoms suggestive of acute infectious mononucleosis. After 14 months, serological parameters were: positive VCA-IgG, EBNA-IgG and negative VCA-IgM. This serological pattern has been maintained since then even during episodes suggestive of EBV reactivation. PCR amplified a specific DNA fragment from the EBV gp220 (detection limit of 100 viral copies). All twenty-four whole blood samples yielded positive results by PCR, while 12 out of 24 serum samples were positive. We aimed at analyzing whether detection of EBV-DNA in serum samples by PCR was associated with overt disease as stated by the need of antiviral treatment and hospitalization. Statistical analysis showed agreement between the two parameters evidenced by the Kappa test (value 0.750; p < 0.001). We concluded that detection of EBV-DNA in serum samples of immunosuppressed patients might be used as a laboratory marker of active EBV disease when a Real-Time PCR or another quantitative method is not available.

Direct and quantitative analysis of underivatized acylcarnitines in serum and whole blood using paper spray mass spectrometry

PubMed Central

Yang, Qian; Manicke, Nicholas E.; Wang, He; Petucci, Christopher; Cooks, R. Graham

2013-01-01

A simple protocol for rapid quantitation of acylcarnitines in serum and whole blood has been developed using paper spray mass spectrometry. Dried serum and whole blood containing a mixture of ten acylcarnitines at various concentrations were analyzed as spots from paper directly without any sample pretreatment, separation, or derivatization. The composition of the spray solvent was found to be a critical factor: for serum samples, spray solvent of methanol/water/formic acid (80:20:0.1) gave the best signal intensity while for blood samples which contain more matrix components, acetonitrile/water (90:10) was a much more suitable spray solvent. For the paper type and size used, 0.5 μL of sample provided an optimal signal for both serum and whole blood samples. For quantitative profiling, the limits of quantitation obtained from both serum and blood were much lower than the clinically validated cutoff values for diagnosis of fatty acid oxidation disorders in newborn screening. Linearity (R2>0.95) and reproducibility (RSD ~10 %) were achieved in the concentration ranges from 100 nM to 5 μM for the C2 acylcarnitine, and for other acylcarnitines, these values were from 10 to 500 nM. Acylcarnitine profiles offer an effective demonstration of the fact that paper spray mass spectrometry is an appropriate, simple, rapid method with high sensitivity and high reproducibility applicable to newborn screening tests. PMID:22760507

Partition of environmental chemicals between maternal and fetal blood and tissues.

PubMed

Needham, Larry L; Grandjean, Philippe; Heinzow, Birger; Jørgensen, Poul J; Nielsen, Flemming; Patterson, Donald G; Sjödin, Andreas; Turner, Wayman E; Weihe, Pal

2011-02-01

Passage of environmental chemicals across the placenta has important toxicological consequences, as well as for choosing samples for analysis and for interpreting the results. To obtain systematic data, we collected in 2000 maternal and cord blood, cord tissue, placenta, and milk in connection with births in the Faroe Islands, where exposures to marine contaminants is increased. In 15 sample sets, we measured a total of 87 environmental chemicals, almost all of which were detected both in maternal and fetal tissues. The maternal serum lipid-based concentrations of organohalogen compounds averaged 1.7 times those of cord serum, 2.8 times those of cord tissue and placenta, and 0.7 those of milk. For organohalogen compounds detectable in all matrices, a high degree of correlation between concentrations in maternal serum and the other tissues investigated was generally observed (r(2) > 0.5). Greater degree of chlorination resulted in lower transfer from maternal serum into milk. Concentrations of pentachlorbenzene, γ-hexachlorocyclohexane, and several polychlorinated biphenyl congeners with low chlorination were higher in fetal samples and showed poor correlation with maternal levels. Perfluorinated compounds occurred in lower concentrations in cord serum than in maternal serum. Cadmium, lead, mercury, and selenium were all detected in fetal samples, but only mercury showed close correlations among concentrations in different matrices. Although the environmental chemicals examined pass through the placenta and are excreted into milk, partitions between maternal and fetal samples are not uniform.

A comprehensive biosensor integrated with a ZnO nanorod FET array for selective detection of glucose, cholesterol and urea.

PubMed

Ahmad, Rafiq; Tripathy, Nirmalya; Park, Jin-Ho; Hahn, Yoon-Bong

2015-08-04

We report a novel straightforward approach for simultaneous and highly-selective detection of multi-analytes (i.e. glucose, cholesterol and urea) using an integrated field-effect transistor (i-FET) array biosensor without any interference in each sensor response. Compared to analytically-measured data, performance of the ZnO nanorod based i-FET array biosensor is found to be highly reliable for rapid detection of multi-analytes in mice blood, and serum and blood samples of diabetic dogs.

Characterizing concentrations of diethylene glycol and suspected metabolites in human serum, urine, and cerebrospinal fluid samples from the Panama DEG mass poisoning.

PubMed

Schier, J G; Hunt, D R; Perala, A; McMartin, K E; Bartels, M J; Lewis, L S; McGeehin, M A; Flanders, W D

2013-12-01

Diethylene glycol (DEG) mass poisoning is a persistent public health problem. Unfortunately, there are no human biological data on DEG and its suspected metabolites in poisoning. If present and associated with poisoning, the evidence for use of traditional therapies such as fomepizole and/or hemodialysis would be much stronger. To characterize DEG and its metabolites in stored serum, urine, and cerebrospinal fluid (CSF) specimens obtained from human DEG poisoning victims enrolled in a 2006 case-control study. In the 2006 study, biological samples from persons enrolled in a case-control study (42 cases with new-onset, unexplained AKI and 140 age-, sex-, and admission date-matched controls without AKI) were collected and shipped to the Centers for Disease Control and Prevention (CDC) in Atlanta for various analyses and were then frozen in storage. For this study, when sufficient volume of the original specimen remained, the following analytes were quantitatively measured in serum, urine, and CSF: DEG, 2-hydroxyethoxyacetic acid (HEAA), diglycolic acid, ethylene glycol, glycolic acid, and oxalic acid. Analytes were measured using low resolution GC/MS, descriptive statistics calculated and case results compared with controls when appropriate. Specimens were de-identified so previously collected demographic, exposure, and health data were not available. The Wilcoxon Rank Sum test (with exact p-values) and bivariable exact logistic regression were used in SAS v9.2 for data analysis. The following samples were analyzed: serum, 20 case, and 20 controls; urine, 11 case and 22 controls; and CSF, 11 samples from 10 cases and no controls. Diglycolic acid was detected in all case serum samples (median, 40.7 mcg/mL; range, 22.6-75.2) and no controls, and in all case urine samples (median, 28.7 mcg/mL; range, 14-118.4) and only five (23%) controls (median, < Lower Limit of Quantitation (LLQ); range, < LLQ-43.3 mcg/mL). Significant differences and associations were identified between case status and the following: 1) serum oxalic acid and serum HEAA (both OR = 14.6; 95% C I = 2.8-100.9); 2) serum diglycolic acid and urine diglycolic acid (both OR > 999; exact p < 0.0001); and 3) urinary glycolic acid (OR = 0.057; 95% C I = 0.001-0.55). Two CSF sample results were excluded and two from the same case were averaged, yielding eight samples from eight cases. Diglycolic acid was detected in seven (88%) of case CSF samples (median, 2.03 mcg/mL; range, < LLQ, 7.47). Significantly elevated HEAA (serum) and diglycolic acid (serum and urine) concentrations were identified among cases, which is consistent with animal data. Low urinary glycolic acid concentrations in cases may have been due to concurrent AKI. Although serum glycolic concentrations among cases may have initially increased, further metabolism to oxalic acid may have occurred thereby explaining the similar glycolic acid concentrations in cases and controls. The increased serum oxalic acid concentration results in cases versus controls are consistent with this hypothesis. Diglycolic acid is associated with human DEG poisoning and may be a biomarker for poisoning. These findings add to animal data suggesting a possible role for traditional antidotal therapies. The detection of HEAA and diglycolic acid in the CSF of cases suggests a possible association with signs and symptoms of DEG-associated neurotoxicity. Further work characterizing the pathophysiology of DEG-associated neurotoxicity and the role of traditional toxic alcohol therapies such as fomepizole and hemodialysis is needed.

Sipa1l1 is an early biomarker of liver fibrosis in CCl4-treated rats

PubMed Central

Marfà, Santiago; Morales-Ruiz, Manuel; Oró, Denise; Ribera, Jordi; Fernández-Varo, Guillermo; Jiménez, Wladimiro

2016-01-01

ABSTRACT At present, several procedures are used for staging liver fibrosis. However, these methods may involve clinical complications and/or present diagnostic uncertainty mainly in the early stages of the disease. Thus, this study was designed to unveil new non-invasive biomarkers of liver fibrosis in an in vivo model of fibrosis/cirrhosis induction by CCl4 inhalation by using a label-free quantitative LC-MS/MS approach. We analyzed 94 serum samples from adult Wistar rats with different degrees of liver fibrosis and 36 control rats. Firstly, serum samples from 18 CCl4-treated rats were clustered into three different groups according to the severity of hepatic and the serum proteome was characterized by label-free LC-MS/MS. Furthermore, three different pooled serum samples obtained from 16 control Wistar rats were also analyzed. Based on the proteomic data obtained, we performed a multivariate analysis which displayed three main cell signaling pathways altered in fibrosis. In cirrhosis, more biological imbalances were detected as well as multi-organ alterations. In addition, hemopexin and signal-induced proliferation-associated 1 like 1 (SIPA1L1) were selected as potential serum markers of liver fibrogenesis among all the analyzed proteins. The results were validated by ELISA in an independent group of 76 fibrotic/cirrhotic rats and 20 controls which confirmed SIPA1L1 as a potential non-invasive biomarker of liver fibrosis. In particular, SIPA1L1 showed a clear diminution in serum samples from fibrotic/cirrhotic rats and a great accuracy at identifying early fibrotic stages. In conclusion, the proteomic analysis of serum samples from CCl4-treated rats has enabled the identification of SIPA1L1 as a non-invasive marker of early liver fibrosis. PMID:27230648

Serum and urine chromium as indices of chromium status in tannery workers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Randall, J.A.; Gibson, R.S.

Serum and urinary Cr levels of a selected group of men exposed to CrIII in four Southern Ontario tanneries were compared with those of men not exposed to Cr. Fasted blood samples were obtained from 72 tannery workers (TW; mean age +/- SD = 36 +/- 12 years) and from 52 controls (CS; mean age +/- SD = 41 +/- 13 years). Serum Cr levels as determined by graphite furnace atomic absorption spectrophotometry were significantly higher (P = 0.0001) for TW (median 0.49 ng/ml, range 0.37-0.81) than for CS (median 0.15 ng/ml, range 0.12-0.20). Urine samples were collected from 49more » TW and 43 CS on a Friday pm and from 42 TW on a Monday am. Urinary creatine (Cre) was determined by the Jaffe reaction. For Friday samples, the median urinary Cr/Cre ratio was significantly higher (P = 0.0001) for TW (median 0.83 ng/mg, range 0.48-1.82) than for CS (median 0.18 ng/mg, range 0.13-0.26). For TW, Cr/Cre was correlated with serum Cr (r = 0.72, P = 0.0001). Neither urinary Cr/Cre nor serum Cr was correlated with length of employment in the tanning industry. There were significant differences in serum Cr levels and urinary Cr/Cre ratios among TW employed in different areas of the tanneries. For TW, the median urinary Cr/Cre ratio for Monday morning samples was significantly lower than for Friday afternoon samples (P = 0.03). These data indicate that CrIII is absorbed and that serum and urine Cr in tannery workers may be indices of Cr exposure and status.« less

Serum and urine chromium as indices of chromium status in tannery workers.

PubMed

Randall, J A; Gibson, R S

1987-05-01

Serum and urinary Cr levels of a selected group of men exposed to CrIII in four Southern Ontario tanneries were compared with those of men not exposed to Cr. Fasted blood samples were obtained from 72 tannery workers (TW; mean age +/- SD = 36 +/- 12 years) and from 52 controls (CS; mean age +/- SD = 41 +/- 13 years). Serum Cr levels as determined by graphite furnace atomic absorption spectrophotometry were significantly higher (P = 0.0001) for TW (median 0.49 ng/ml, range 0.37-0.81) than for CS (median 0.15 ng/ml, range 0.12-0.20). Urine samples were collected from 49 TW and 43 CS on a Friday pm and from 42 TW on a Monday am. Urinary creatine (Cre) was determined by the Jaffe reaction. For Friday samples, the median urinary Cr/Cre ratio was significantly higher (P = 0.0001) for TW (median 0.83 ng/mg, range 0.48-1.82) than for CS (median 0.18 ng/mg, range 0.13-0.26). For TW, Cr/Cre was correlated with serum Cr (r = 0.72, P = 0.0001). Neither urinary Cr/Cre nor serum Cr was correlated with length of employment in the tanning industry. There were significant differences in serum Cr levels and urinary Cr/Cre ratios among TW employed in different areas of the tanneries. For TW, the median urinary Cr/Cre ratio for Monday morning samples was significantly lower than for Friday afternoon samples (P = 0.03). These data indicate that CrIII is absorbed and that serum and urine Cr in tannery workers may be indices of Cr exposure and status.

Association between serum soluble CD30 and serum creatinine before and after renal transplantation.

PubMed

López-Hoyos, M; San Segundo, D; Benito, M J; Fernández-Fresnedo, G; Ruiz, J C; Rodrigo, E; Gómez-Alamillo, C; Benito, A; Arias, M

2008-11-01

There is increasing evidence that circulating levels of soluble CD30 (sCD30) may represent a biomarker for outcome in kidney transplantation. The aim of this study was to measure the pre- and posttransplantation serum levels of sCD30 in cadaveric kidney transplant recipients and correlate them with serum creatinine. Serum sCD30 was measured by a commercial enzyme-linked immunosorbent assay (ELISA) from prospective samples of 38 kidney allograft recipients serially transplanted at our center. Samples were collected at day 0 pretransplantation and at months 6, 12, 18, and 24 posttransplantation. We also studied sera from 29 patients with chronic kidney disease (CKD) at different stages of the K/DOQI guidelines, as a control group. Serum levels of sCD30 decreased significantly in samples posttransplantation compared with pretransplantation. The significant decrease after transplantation may be related to the improvement in renal function since we observed a significant correlation between serum levels of sCD30 and creatinine (sCr) at all times of the study. In addition, the patients with chronic renal failure showed a significant association between serum sCD30 and sCr (r = .454; P = .013). Our results did not suggest that the measurement of sCD30 may be used as a valuable biomarker in renal transplantation. Increased levels may be related to a decrease in its renal elimination.

Antibody titres at diagnosis and during follow-up of anti-NMDA receptor encephalitis: a retrospective study.

PubMed

Gresa-Arribas, Nuria; Titulaer, Maarten J; Torrents, Abiguei; Aguilar, Esther; McCracken, Lindsey; Leypoldt, Frank; Gleichman, Amy J; Balice-Gordon, Rita; Rosenfeld, Myrna R; Lynch, David; Graus, Francesc; Dalmau, Josep

2014-02-01

Anti-N-methyl-d-aspartate (NMDA) receptor encephalitis is a severe but treatable autoimmune disorder which diagnosis depends on sensitive and specific antibody testing. We aimed to assess the sensitivity and specificity of serum and CSF antibody testing in patients with anti-NMDA receptor encephalitis, and the relation between titres, relapses, outcome, and epitope repertoire. In this observational study, we used rat brain immunohistochemistry and cell-based assays (CBA) with fixed or live NMDA receptor-expressing cells to determine the sensitivity and specificity of antibody testing in paired serum and CSF samples. Samples were obtained at diagnosis from patients with anti-NMDA receptor encephalitis and from control participants worldwide. We deemed a patient to be antibody positive if their serum, their CSF, or both tested positive with both immunohistochemistry and CBA techniques; we determined titres with serial sample dilution using brain immunohistochemistry. We examined samples from 45 patients (25 with good outcome [modified Rankin Scale, mRS 0-2], ten with poor outcome [mRS 3-6], and ten with relapses) at three or more timepoints. We determined the epitope repertoire in the samples of 23 patients with CBA expressing GluN1-NMDA receptor mutants. We analysed samples from 250 patients with anti-NMDA receptor encephalitis and 100 control participants. All 250 patients had NMDA receptor antibodies in CSF but only 214 had antibodies in serum (sensitivity 100.0% [98.5-1000%] vs 85.6% [80.7-89.4%], p<0.0001). Serum immunohistochemistry testing was more often in agreement with CBA with fixed cells (77 [71%] of 108) than with CBA with live cells (63 [58%] of 108, p=0.0056). In multivariable analysis, CSF and serum titres were higher in patients with poor outcome than in those with good outcome (CSF dilution 340 vs 129, difference 211, [95% CI 1-421], p=0.049; serum dilution 7370 vs 1243, difference 6127 [2369-9885], p=0.0025), and in patients with teratoma than in those without teratoma (CSF 395 vs 110, difference 285 [134-437], p=0.0079; serum 5515 vs 1644, difference 3870 [548-7193], p=0.024). Over time there was a decrease of antibody titres in the 35 patients with good or poor outcome and samples followed at three timepoints regardless of outcome (from diagnosis to last follow-up: CSF 614 to 76, difference 538 [288-788]; serum 5460 to 1564, difference 3896 [2428-5362]; both p<0.0001). Relapses were associated with a change in titre more often in CSF than in serum (14 of 19 vs seven of 16, p=0.037). After recovery, 24 of 28 CSF samples and 17 of 23 serum samples from patients remained antibody positive. Patients' antibodies targeted a main epitope region at GluN1 aminoacid 369; the epitope repertoire did not differ between patients with different outcomes, and did not change during relapses. The sensitivity of NMDA receptor antibody testing is higher in CSF than in serum. Antibody titres in CSF and serum were higher in patients with poor outcome or teratoma than in patients with good outcome or no tumour. The titre change in CSF was more closely related with relapses than was that in serum. These findings emphasise the importance of including CSF in antibody studies, and that antibody titres can complement clinical assessments. Dutch Cancer Society, National Institutes of Health, McKnight Neuroscience of Brain Disorders award, the Fondo de Investigaciones Sanitarias, ErasmusMC fellowship, and Fundació la Marató de TV3. Copyright © 2014 Elsevier Ltd. All rights reserved.

Simultaneous determination of creatinine and creatine in human serum by double-spike isotope dilution liquid chromatography-tandem mass spectrometry (LC-MS/MS) and gas chromatography-mass spectrometry (GC-MS).

PubMed

Fernández-Fernández, Mario; Rodríguez-González, Pablo; Añón Álvarez, M Elena; Rodríguez, Felix; Menéndez, Francisco V Álvarez; García Alonso, J Ignacio

2015-04-07

This work describes the first multiple spiking isotope dilution procedure for organic compounds using (13)C labeling. A double-spiking isotope dilution method capable of correcting and quantifying the creatine-creatinine interconversion occurring during the analytical determination of both compounds in human serum is presented. The determination of serum creatinine may be affected by the interconversion between creatine and creatinine during sample preparation or by inefficient chemical separation of those compounds by solid phase extraction (SPE). The methodology is based on the use differently labeled (13)C analogues ((13)C1-creatinine and (13)C2-creatine), the measurement of the isotopic distribution of creatine and creatinine by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and the application of multiple linear regression. Five different lyophilized serum-based controls and two certified human serum reference materials (ERM-DA252a and ERM-DA253a) were analyzed to evaluate the accuracy and precision of the proposed double-spike LC-MS/MS method. The methodology was applied to study the creatine-creatinine interconversion during LC-MS/MS and gas chromatography-mass spectrometry (GC-MS) analyses and the separation efficiency of the SPE step required in the traditional gas chromatography-isotope dilution mass spectrometry (GC-IDMS) reference methods employed for the determination of serum creatinine. The analysis of real serum samples by GC-MS showed that creatine-creatinine separation by SPE can be a nonquantitative step that may induce creatinine overestimations up to 28% in samples containing high amounts of creatine. Also, a detectable conversion of creatine into creatinine was observed during sample preparation for LC-MS/MS. The developed double-spike LC-MS/MS improves the current state of the art for the determination of creatinine in human serum by isotope dilution mass spectrometry (IDMS), because corrections are made for all the possible errors derived from the sample preparation step.

Serum Albumin Domain Structures in Human Blood Serum by Mass Spectrometry and Computational Biology.

PubMed

Belsom, Adam; Schneider, Michael; Fischer, Lutz; Brock, Oliver; Rappsilber, Juri

2016-03-01

Chemical cross-linking combined with mass spectrometry has proven useful for studying protein-protein interactions and protein structure, however the low density of cross-link data has so far precluded its use in determining structures de novo. Cross-linking density has been typically limited by the chemical selectivity of the standard cross-linking reagents that are commonly used for protein cross-linking. We have implemented the use of a heterobifunctional cross-linking reagent, sulfosuccinimidyl 4,4'-azipentanoate (sulfo-SDA), combining a traditional sulfo-N-hydroxysuccinimide (sulfo-NHS) ester and a UV photoactivatable diazirine group. This diazirine yields a highly reactive and promiscuous carbene species, the net result being a greatly increased number of cross-links compared with homobifunctional, NHS-based cross-linkers. We present a novel methodology that combines the use of this high density photo-cross-linking data with conformational space search to investigate the structure of human serum albumin domains, from purified samples, and in its native environment, human blood serum. Our approach is able to determine human serum albumin domain structures with good accuracy: root-mean-square deviation to crystal structure are 2.8/5.6/2.9 Å (purified samples) and 4.5/5.9/4.8Å (serum samples) for domains A/B/C for the first selected structure; 2.5/4.9/2.9 Å (purified samples) and 3.5/5.2/3.8 Å (serum samples) for the best out of top five selected structures. Our proof-of-concept study on human serum albumin demonstrates initial potential of our approach for determining the structures of more proteins in the complex biological contexts in which they function and which they may require for correct folding. Data are available via ProteomeXchange with identifier PXD001692. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Canis lupus familiaris involved in the transmission of pathogenic Yersinia spp. in China.

PubMed

Wang, Xin; Liang, Junrong; Xi, Jinxiao; Yang, Jinchuan; Wang, Mingliu; Tian, Kecheng; Li, Jicheng; Qiu, Haiyan; Xiao, Yuchun; Duan, Ran; Yang, Haoshu; Li, Kewei; Cui, Zhigang; Qi, Meiying; Jing, Huaiqi

2014-08-06

To investigate canines carrying pathogens associated with human illness, we studied their roles in transmitting and maintaining pathogenic Yersinia spp. We examined different ecological landscapes in China for the distribution of pathogenic Yersinia spp. in Canis lupus familiaris, the domestic dog. The highest number of pathogenic Yersinia enterocolitica was shown from the tonsils (6.30%), followed by rectal swabs (3.63%) and feces (1.23%). Strains isolated from plague free areas for C. lupus familiaris, local pig and diarrhea patients shared the same pulsed-field gel electrophoresis (PFGE) pattern, indicating they may be from the same clone and the close transmission source of pathogenic Y. enterocolitica infections in these areas. Among 226 dogs serum samples collected from natural plague areas of Yersinia pestis in Gansu and Qinghai Provinces, 49 were positive for F1 antibody, while the serum samples collected from plague free areas were all negative, suggested a potential public health risk following exposure to dogs. No Y. enterocolitica or Yersinia pseudotuberculosis was isolated from canine rectal swabs in natural plague areas. Therefore, pathogenic Yersinia spp. may be regionally distributed in China. Copyright © 2014 Elsevier B.V. All rights reserved.

Determination of polychlorinated biphenyl levels in the serum of residents and in the homogenates of seafood from the New Bedford, Massachusetts, area: a comparison of exposure sources through pattern recognition techniques.

PubMed

Burse, V W; Groce, D F; Caudill, S P; Korver, M P; Phillips, D L; McClure, P C; Lapeza, C R; Head, S L; Miller, D T; Buckley, D J

1994-04-29

We measured the residues of polychlorinated biphenyls (PCBs) in the serum of 23 residents of the New Bedford, Massachusetts, area and from two homogenates each of bluefish and lobsters from the same area. We used congener-specific and total Aroclor quantitative approaches, both of which involved gas chromatography with electron capture detection. Using gas chromatography mass spectrometry (electron ionization mode), we confirmed the presence of PCBs in the combined serum samples and in the aliquots of bluefish and lobsters. In measuring the PCB levels in serum, we found good agreement between the two electron capture detector approaches (r > or = 0.97) when the serum of specific congeners was compared to total Aroclor. We used univariate and multivariate quality control approaches to monitor these analyses. Analytical results for bluefish showed a better agreement between the two techniques than did those for lobsters; however, the small number of samples precluded any statistical comparison. We also measured levels of chlorinated pesticides in the serum samples of two groups of New Bedford residents, those with low PCB levels (< 15 ng/ml) and those with high PCB levels (> or = 15 ng/ml). We found that residents with high PCB levels also tended to have higher levels of hexachlorobenzene (HCB) and 1,1-dichloro-2,2-di-(p-chlorophenyl) ethylene (p,p'-DDE). The higher concentration of all three analytes appears to be influenced by employment in the capacitor industry, by seafood consumption, or both. Using Jaccard measures of similarity and principal component analysis we compared the gas chromatographic patterns of PCBs found in the serum of New Bedford area residents with high serum PCBs with the patterns found in homogenates of lobsters (inclusive of all edible portions except the roe), in homogenates of bluefish fillets taken from local waters, and in serum from goats fed selected technical Aroclors (e.g. Aroclors 1016, 1242, 1254, or 1260). The patterns found in human serum samples were similar to the patterns found in lobster homogenates. Both of these patterns closely resembled patterns found in the serum samples of the goat fed aroclor 1254, as demonstrated by both pattern recognition techniques. In addition, the chromatographic patterns of human serum and of lobsters and bluefish homogenates all indicated the presence of PCBs more characteristic of Aroclors 1016 or 1242.

Polychlorinated biphenyls in umbilical cord serum of newborns from Rio Grande do Sul state, Brazil.

PubMed

Mohr, Susana; Sifuentes dos Santos, Joice; Schwanz, Thiago Guilherme; Wagner, Roger; Mozzaquatro, Joseane Oliveira; Lorenzoni, Alessandra Scherer; Costabeber, Ijoni Hilda

2015-12-07

Polychlorinated biphenyls (PCBs) are food-chain contaminants that have been shown to contaminate foods worldwide. The newborn are exposed to these organochlorine compounds across the placenta and through breastfeeding. They are proven to be carcinogenic and may contribute to congenital malformation etiology. This study examined levels of five PCB congeners (28, 52, 138, 153 and 180) in umbilical cord serum samples from 148 newborns from Rio Grande do Sul state, Brazil. Serum concentrations of PCBs were analyzed by gas chromatography with electron capture detection and mass spectrometry. Levels of ∑PCBs ranged from 0.35 to 55.17 ng/ml in umbilical cord serum positive samples, and PCB 138 was the most prevalent congener. Only 7.4% of samples presented no PCB congener. Some PCB congener cord serum levels were related to the locale of the mothers' residence, smoking and drinking habits, fruit consumption, and congenital malformation. Copyright © 2015 Elsevier B.V. All rights reserved.

Polyvalent immunoglobulin binding is an obstacle to accurate measurement of specific antibodies with ELISA despite inclusion of blocking agents.

PubMed

Loeffler, David A; Klaver, Andrea C

2017-11-01

Specific antibody concentrations are frequently measured in serum (and plasma and intravenous immunoglobulin) samples by enzyme-linked immunosorbent assay (ELISA). The standard negative control involves incubation of buffer alone on antigen-coated wells. The immunoreactivity that develops in antigen-coated wells in which diluted serum has been incubated is assumed to represent specific antibody binding. This approach can result in marked overestimation of specific antibody levels, because serum contains specific polyvalent antibodies which bind, primarily with low affinity, to multiple antigens (including those on ELISA plates) despite the use of blocking agents. Non-denaturing purification of serum IgG, followed by assessment of the antigen binding or antigen-binding affinity of this purified IgG, can reduce but not eliminate the problem of polyvalent antibody binding in indirect ELISAs. Alternatively, polyvalent antibody binding can be estimated by incubating a diluted serum sample on wells coated with an irrelevant protein (such as bovine serum albumin or a scrambled peptide sequence) or buffer alone, then subtracting this reactivity from the sample's binding to wells coated with the antigen of interest. Polyvalent binding of immunoglobulins must be accounted for in order to obtain accurate ELISA measurements of serum, plasma, or intravenous immunoglobulin antibodies. Copyright © 2017 Elsevier B.V. All rights reserved.

A 3D-Printed, Portable, Optical-Sensing Platform for Smartphones Capable of Detecting the Herbicide 2,4-Dichlorophenoxyacetic Acid.

PubMed

Wang, Yijia; Zeinhom, Mohamed M A; Yang, Mingming; Sun, Rongrong; Wang, Shengfu; Smith, Jordan N; Timchalk, Charles; Li, Lei; Lin, Yuehe; Du, Dan

2017-09-05

Onsite rapid detection of herbicides and herbicide residuals in environmental and biological specimens are important for agriculture, environmental concerns, food safety, and health care. The traditional method for herbicide detection requires expensive laboratory equipment and a long turnaround time. In this work, we developed a single-stripe microliter plate smartphone-based colorimetric device for rapid and low-cost in-field tests. This portable smartphone platform is capable of screening eight samples in a single-stripe microplate. The device combined the advantages of small size (50 × 100 × 160 mm 3 ) and low cost ($10). The platform was calibrated by using two different dye solutions, i.e. methyl blue (MB) and rhodamine B, for the red and green channels. The results showed good correlation with results attained from a traditional laboratory reader. We demonstrated the application of this platform for detection of the herbicide 2,4-dichlorophenoxyacetic acid in the range of 1 to 80 ppb. Spiked samples of tap water, rat serum, plasma, and human serum were tested by our device. Recoveries obtained varied from 95.6% to 105.2% for all of the spiked samples using the microplate reader and from 93.7% to 106.9% for all of the samples using the smartphone device. This work validated that the smartphone optical-sensing platform is comparable to the commercial microplate reader; it is eligible for onsite, rapid, and low-cost detection of herbicides for environmental evaluation and biological monitoring.

[The Competence Network for HIV/AIDS. Data, Samples, Facts].

PubMed

Michalik, Claudia; Skaletz-Rorowski, Adriane; Brockmeyer, Norbert H

2016-04-01

With funding for the Competence Networks in Medicine from the Federal Ministry of Education and Research, the Competence Network for HIV/AIDS (KompNet HIV/AIDS) was established as an interdisciplinary research association. Essential working groups were incorporated all over Germany, which are active in clinical and basic HIV/AIDS research. After successful establishment, providing research infrastructure for national and international cooperation in the field of HIV/AIDS was the focus of the network. By bringing together research activities, preconditions are created for improving HIV infection treatment and increasing life expectancy of HIV-infected patients. The members of KompNet HIV/AIDS are HIV experts from university clinics, HIV physicians, patient representatives, as well as national reference centers. As a scientific research basis, the network established an HIV patient cohort. Clinical and sociodemographic data of HIV patients were documented biannually and complemented by serum and DNA-samples collected twice per year. Furthermore, a child cohort was set up. Within the KompNet HIV/AIDS, a research infrastructure for HIV was established for internal, external as well international scientists. Within the HIV cohort a total of 16,500 patients are documented. The associated biobank comprises ~ 56,000 serum samples and ~ 16,000 DNA samples. The child cohort consists of 647 HIV-exposed and 230 infected children. The KompNet HIV/AIDS cohorts became an important partner in several international collaborations. Nevertheless, the maintenance of such infrastructures without public funding is a challenge.

Determination of serum calcium levels by 42Ca isotope dilution inductively coupled plasma mass spectrometry.

PubMed

Han, Bingqing; Ge, Menglei; Zhao, Haijian; Yan, Ying; Zeng, Jie; Zhang, Tianjiao; Zhou, Weiyan; Zhang, Jiangtao; Wang, Jing; Zhang, Chuanbao

2017-11-27

Serum calcium level is an important clinical index that reflects pathophysiological states. However, detection accuracy in laboratory tests is not ideal; as such, a high accuracy method is needed. We developed a reference method for measuring serum calcium levels by isotope dilution inductively coupled plasma mass spectrometry (ID ICP-MS), using 42Ca as the enriched isotope. Serum was digested with 69% ultrapure nitric acid and diluted to a suitable concentration. The 44Ca/42Ca ratio was detected in H2 mode; spike concentration was calibrated by reverse IDMS using standard reference material (SRM) 3109a, and sample concentration was measured by a bracketing procedure. We compared the performance of ID ICP-MS with those of three other reference methods in China using the same serum and aqueous samples. The relative expanded uncertainty of the sample concentration was 0.414% (k=2). The range of repeatability (within-run imprecision), intermediate imprecision (between-run imprecision), and intra-laboratory imprecision were 0.12%-0.19%, 0.07%-0.09%, and 0.16%-0.17%, respectively, for two of the serum samples. SRM909bI, SRM909bII, SRM909c, and GBW09152 were found to be within the certified value interval, with mean relative bias values of 0.29%, -0.02%, 0.10%, and -0.19%, respectively. The range of recovery was 99.87%-100.37%. Results obtained by ID ICP-MS showed a better accuracy than and were highly correlated with those of other reference methods. ID ICP-MS is a simple and accurate candidate reference method for serum calcium measurement and can be used to establish and improve serum calcium reference system in China.

Detection of a Serum Siderophore by LC-MS/MS as a Potential Biomarker of Invasive Aspergillosis

PubMed Central

Carroll, Cassandra S.; Amankwa, Lawrence N.; Pinto, Linda J.; Fuller, Jeffrey D.; Moore, Margo M.

2016-01-01

Invasive aspergillosis (IA) is a life-threatening systemic mycosis caused primarily by Aspergillus fumigatus. Early diagnosis of IA is based, in part, on an immunoassay for circulating fungal cell wall carbohydrate, galactomannan (GM). However, a wide range of sensitivity and specificity rates have been reported for the GM test across various patient populations. To obtain iron in vivo, A. fumigatus secretes the siderophore, N,N',N"-triacetylfusarinine C (TAFC) and we hypothesize that TAFC may represent a possible biomarker for early detection of IA. We developed an ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for TAFC analysis from serum, and measured TAFC in serum samples collected from patients at risk for IA. The method showed lower and upper limits of quantitation (LOQ) of 5 ng/ml and 750 ng/ml, respectively, and complete TAFC recovery from spiked serum. As proof of concept, we evaluated 76 serum samples from 58 patients with suspected IA that were investigated for the presence of GM. Fourteen serum samples obtained from 11 patients diagnosed with probable or proven IA were also analyzed for the presence of TAFC. Control sera (n = 16) were analyzed to establish a TAFC cut-off value (≥6 ng/ml). Of the 36 GM-positive samples (≥0.5 GM index) from suspected IA patients, TAFC was considered positive in 25 (69%). TAFC was also found in 28 additional GM-negative samples. TAFC was detected in 4 of the 14 samples (28%) from patients with proven/probable aspergillosis. Log-transformed TAFC and GM values from patients with proven/probable IA, healthy individuals and SLE patients showed a significant correlation with a Pearson r value of 0.77. In summary, we have developed a method for the detection of TAFC in serum that revealed this fungal product in the sera of patients at risk for invasive aspergillosis. A prospective study is warranted to determine whether this method provides improved early detection of IA. PMID:26974544

Variation of calcium, copper and iron levels in serum, bile and stone samples of patients having different types of gallstone: A comparative study.

PubMed

Khan, Mustafa; Kazi, Tasneem Gul; Afridi, Hassan Imran; Sirajuddin; Bilal, Muhammad; Akhtar, Asma; Khan, Sabir; Kadar, Salma

2017-08-01

Epidemiological data among the human population has shown a significantly increased incidence of gallstone (GS) disease worldwide. It was studied that some essential (calcium) and transition elements (iron and copper) in bile play an important role in the development of GS. The estimation of calcium, copper and iron were carried out in the serum, gall bladder bile and different types of GS (cholesterol, mixed and pigmented) of 172 patients, age ranged 20-55years. For comparative purpose age matched referents not suffering from GS diseases were also selected. Biliary concentrations of calcium (Ca), iron (Fe) and copper (Cu) were correlated with their concentrations in serum and different types of GS samples. The ratio of Ca, Fe and Cu in bile with serum was also calculated. Understudy metals were determined by flame atomic absorption spectroscopy after acid decomposition of matrices of selected samples. The Ca concentrations in serum samples were significantly higher in patients with pigmented GS as compared to controls (p<0.005), whereas for patients having cholesterol and mixed GS the concentrations were on the lower side. Biliary Ca concentrations of patients were found to be higher than controls, but difference was significant for pigmented GS patients (p>0.001). The contents of Cu and Fe in serum and bile of all patients (except female cholesterol GS patient have low serum iron concentration) were found to be higher than control, but difference was significant in those patients who have pigmented GS. The concentration of Ca, Fe and Cu in different types GS were found in the order, Pigmented>mixed>cholesterol. The bile/serum ratio for Ca, Cu and Fe was found to be significantly higher in pigmented GS patients. Gall bladder bile was slightly alkaline in patients as compared to referents. The density of bile was found to be higher in patients as compared to the referents. Various functional groups present in different types of GS samples were confirmed by Fourier transform infra-red spectroscopy. The higher density and pH of bile, elevated concentrations of transition elements in all types of biological samples (serum, bile and GS), could be an important factor for the formation of different types of GS. Copyright © 2017 Elsevier B.V. All rights reserved.

Survival of the North American strain of viral hemorrhagic septicemia virus (VHSV) in filtered seawater and seawater containing ovarian fluid, crude oil and serum-enriched culture medium

USGS Publications Warehouse

Kocan, R.M.; Hershberger, P.K.; Elder, N.E.

2001-01-01

 The North American strain of viral hemorrhagic septicemia virus (NA-VHSV) could be recovered for up to 40 h in natural filtered seawater (27 ppt) with a 50% loss of infectivity after approximately 10 h at 15°C. Addition of 10 ppb North Slope crude oil to the seawater had no effect on virus survival. However, when various concentrations of teleost ovarian fluid were added to seawater, virus could be recovered after 72 h at 0.01% ovarian fluid and after 96 h at 1.0%. When cell culture medium supplemented with 10% fetal bovine serum was added to the seawater, 100% of the virus could be recovered for the first 15 d and 60% of the virus remained after 36 d. These findings quantify NA-VHSV infectivity in natural seawater and demonstrate that ovarian fluid, which occurs naturally during spawning events, significantly prolongs the survival and infectivity of the virus. The extended stabilization of virus in culture medium supplemented with serum allows for low titer field samples to be collected and transported in an unfrozen state without significant loss of virus titer.

Large-Scale Evaluation of the Immuno-Mycologics Lateral Flow and Enzyme-Linked Immunoassays for Detection of Cryptococcal Antigen in Serum and Cerebrospinal Fluid

PubMed Central

Hansen, Jessica; Slechta, E. Susan; Gates-Hollingsworth, Marcellene A.; Neary, Brandon; Barker, Adam P.; Bauman, Sean; Kozel, Thomas R.

2013-01-01

Cryptococcosis is a systemic infection caused by the pathogenic yeasts Cryptococcus neoformans and C. gattii. Detection of cryptococcal capsular antigen (CrAg) in serum and cerebrospinal fluid (CSF) plays an important diagnostic role. We prospectively compared the new Immuno-Mycologics Inc. (IMMY) lateral flow assay (LFA) and enzyme immunoassay (EIA) to our current CrAg test (Premier EIA; Meridian Bioscience Inc.). Discordant samples were retested with the latex-Cryptococcus antigen test (IMMY) and using serotype-specific monoclonal antibodies (MAbs). A total of 589 serum and 411 CSF specimens were tested in parallel. Qualitative agreement across assays was 97.7%. In all, 56 (41 serum and 15 CSF) samples were positive and 921 (527 serum and 394 CSF) samples were negative by all three assays. The 23 discrepant specimens were all Meridian EIA negative. Of 23 discordant specimens, 20 (87.0%) were positive by both the IMMY LFA and EIA, 2 were LFA positive only, and 1 was EIA positive only. Eleven discrepant specimens had adequate volume for latex agglutination (LA) testing; 8 were LA positive, and 3 were LA negative. LA-negative samples (2 CSF samples and 1 serum) had low IMMY LFA/EIA titers (≤1:10). Serotype-specific MAb analysis of the LA-positive samples suggested that these specimens contained CrAg epitopes similar to those of serotype C strains. In conclusion, the IMMY assays showed excellent overall concordance with the Meridian EIA. Assay performance differences were related to issues of analytic sensitivity and possible serotype bias. Incomplete access to patient-level data combined with low specimen volumes limited our ability to fully resolve discrepant results. PMID:23114703

Large-scale evaluation of the immuno-mycologics lateral flow and enzyme-linked immunoassays for detection of cryptococcal antigen in serum and cerebrospinal fluid.

PubMed

Hansen, Jessica; Slechta, E Susan; Gates-Hollingsworth, Marcellene A; Neary, Brandon; Barker, Adam P; Bauman, Sean; Kozel, Thomas R; Hanson, Kimberly E

2013-01-01

Cryptococcosis is a systemic infection caused by the pathogenic yeasts Cryptococcus neoformans and C. gattii. Detection of cryptococcal capsular antigen (CrAg) in serum and cerebrospinal fluid (CSF) plays an important diagnostic role. We prospectively compared the new Immuno-Mycologics Inc. (IMMY) lateral flow assay (LFA) and enzyme immunoassay (EIA) to our current CrAg test (Premier EIA; Meridian Bioscience Inc.). Discordant samples were retested with the latex-Cryptococcus antigen test (IMMY) and using serotype-specific monoclonal antibodies (MAbs). A total of 589 serum and 411 CSF specimens were tested in parallel. Qualitative agreement across assays was 97.7%. In all, 56 (41 serum and 15 CSF) samples were positive and 921 (527 serum and 394 CSF) samples were negative by all three assays. The 23 discrepant specimens were all Meridian EIA negative. Of 23 discordant specimens, 20 (87.0%) were positive by both the IMMY LFA and EIA, 2 were LFA positive only, and 1 was EIA positive only. Eleven discrepant specimens had adequate volume for latex agglutination (LA) testing; 8 were LA positive, and 3 were LA negative. LA-negative samples (2 CSF samples and 1 serum) had low IMMY LFA/EIA titers (≤1:10). Serotype-specific MAb analysis of the LA-positive samples suggested that these specimens contained CrAg epitopes similar to those of serotype C strains. In conclusion, the IMMY assays showed excellent overall concordance with the Meridian EIA. Assay performance differences were related to issues of analytic sensitivity and possible serotype bias. Incomplete access to patient-level data combined with low specimen volumes limited our ability to fully resolve discrepant results.

The influence of storage time and temperature on the measurement of serum, plasma and urine osmolality.

PubMed

Bezuidenhout, Karla; Rensburg, Megan A; Hudson, Careen L; Essack, Younus; Davids, M Razeen

2016-07-01

Many clinical laboratories require that specimens for serum and urine osmolality determination be processed within 3 h of sampling or need to arrive at the laboratory on ice. This protocol is based on the World Health Organization report on sample storage and stability, but the recommendation lacks good supporting data. We studied the effect of storage temperature and time on osmolality measurements. Blood and urine samples were obtained from 16 patients and 25 healthy volunteers. Baseline serum, plasma and urine osmolality measurements were performed within 30 min. Measurements were then made at 3, 6, 12, 24 and 36 h on samples stored at 4-8℃ and room temperature. We compared baseline values with subsequent measurements and used difference plots to illustrate changes in osmolality. At 4-8℃, serum and plasma osmolality were stable for up to 36 h. At room temperature, serum and plasma osmolality were very stable for up to 12 h. At 24 and 36 h, changes from baseline osmolality were statistically significant and exceeded the total allowable error of 1.5% but not the reference change value of 4.1%. Urine osmolality was extremely stable at room temperature with a mean change of less than 1 mosmol/kg at 36 h. Serum and plasma samples can be stored at room temperature for up to 36 h before measuring osmolality. Cooling samples to 4-8℃ may be useful when delays in measurement beyond 12 h are anticipated. Urine osmolality is extremely stable for up to 36 h at room temperature. © The Author(s) 2015.

Use of a porous silicon-gold plasmonic nanostructure to enhance serum peptide signals in MALDI-TOF analysis.

PubMed

Li, Xiao; Tan, Jie; Yu, Jiekai; Feng, Jiandong; Pan, Aiwu; Zheng, Shu; Wu, Jianmin

2014-11-07

Small peptides in serum are potential biomarkers for the diagnosis of cancer and other diseases. The identification of peptide biomarkers in human plasma/serum has become an area of high interest in medical research. However, the direct analysis of peptides in serum samples using mass spectrometry is challenging due to the low concentration of peptides and the high abundance of high-molecular-weight proteins in serum, the latter of which causes severe signal suppression. Herein, we reported that porous semiconductor-noble metal hybrid nanostructures can both eliminate the interference from large proteins in serum samples and significantly enhance the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) yields of peptides captured on the nanostructure. Serum peptide fingerprints with high fidelity can be acquired rapidly, and successful discrimination of colorectal cancer patients based on peptide fingerprints is demonstrated. Copyright © 2014 Elsevier B.V. All rights reserved.

Polycyclic Aromatic Hydrocarbons and Polychlorinated Dibenzo-p-Dioxins/Dibenzofurans in Microliter Samples of Human Serum as Exposure Indicators

PubMed Central

Xia, Xiaoyan; Carroll-Haddad, Alesia; Brown, Nicole; Utell, Mark J.; Mallon, Timothy; Hopke, Philip K.

2016-01-01

Objective The objectives were: 1) measure polycyclic aromatic hydrocarbons (PAHs), polychlorinated dibenzo-p-dioxins (PCDDs) and dibenzofurans (PCDFs) in 100 μL of human serum and 2) assess PAH and PCDD/PCDF as markers of burn pit exposures during military deployments. Methods PAHs and PCDDs/PCDFs were analyzed in 100μL serum samples collected pre- and post-deployment from 200 persons deployed to Iraq or Afghanistan (CASE); 200 persons not deployed (CONTROL) with GC/MS. Results Naphthalene was found in ~83% of the samples and was statistically different between post-deployment CASE personnel and pre-deployment. 1,2,3,4,6,7,8-Heptachlorodibenzo-p-dioxin, Octachlorodibenzo-p-dioxin, 1,2,3,7,8,9-Hexachlorodibenzofuran, and 1,2,3,4,6,7,8-Heptachlorodibenzofuran were found in ~38% of samples. Concentrations were significantly different between CASE and CONTROL and between pre- and post-deployment samples. Conclusions PAH and PCDD/PCDF in serum can serve as exposure markers and measurements in small volumes is feasible for quantifying exposure to burn pits. PMID:27501107

Total protein concentration and diagnostic test results for gray wolf (Canis lupus) serum using Nobuto filter paper strips

USGS Publications Warehouse

Jara, Rocio F.; Sepúlveda, Carolina; Ip, Hon S.; Samuel, Michael D.

2015-01-01

Nobuto filter paper strips are widely used for storing blood-serum samples, but the recovery of proteins from these strips following rehydration is unknown. Poor recovery of proteins could reduce the concentration of antibodies and antigens and reduce the sensitivity of diagnostic assays. We compared the protein concentration, and its association with test sensitivity, of eluted Nobuto strip samples with paired sera. We collected and froze serum from five gray wolves (Canis lupus) for 8 mo. When thawed, we used a spectrophotometer (absorbance 280 nm) to determine the serum protein concentration for paired sera and Nobuto eluates for each animal in 2-fold serial dilutions. Total protein concentration was similar for both sample storage methods (Nobuto eluates and control sera), except for the undiluted samples in which Nobuto eluates had higher total protein concentrations. Both sample storage methods appear to produce similar results using the SNAP® 4Dx® Test to detect antibodies against pathogens causing Lyme disease, anaplasmosis, and ehrlichiosis as well as antigen for canine heartworm disease.

Total protein concentration and diagnostic test results for gray wolf (Canis lupus) serum using Nobuto filter paper strips.

PubMed

Jara, Rocío F; Sepúlveda, Carolina; Ip, Hon S; Samuel, Michael D

2015-04-01

Nobuto filter paper strips are widely used for storing blood-serum samples, but the recovery of proteins from these strips following rehydration is unknown. Poor recovery of proteins could reduce the concentration of antibodies and antigens and reduce the sensitivity of diagnostic assays. We compared the protein concentration, and its association with test sensitivity, of eluted Nobuto strip samples with paired sera. We collected and froze serum from five gray wolves (Canis lupus) for 8 mo. When thawed, we used a spectrophotometer (absorbance 280 nm) to determine the serum protein concentration for paired sera and Nobuto eluates for each animal in 2-fold serial dilutions. Total protein concentration was similar for both sample storage methods (Nobuto eluates and control sera), except for the undiluted samples in which Nobuto eluates had higher total protein concentrations. Both sample storage methods appear to produce similar results using the SNAP® 4Dx® Test to detect antibodies against pathogens causing Lyme disease, anaplasmosis, and ehrlichiosis as well as antigen for canine heartworm disease.

Pyridoxal 5'-phosphate, pyridoxal, and 4-pyridoxic acid in the paired serum and cerebrospinal fluid of children.

PubMed

Akiyama, Tomoyuki; Hayashi, Yumiko; Hanaoka, Yoshiyuki; Shibata, Takashi; Akiyama, Mari; Tsuchiya, Hiroki; Yamaguchi, Tokito; Kobayashi, Katsuhiro

2017-09-01

We quantified pyridoxal 5'-phosphate (PLP), pyridoxal (PL), and 4-pyridoxic acid (PA) in paired serum and cerebrospinal fluid (CSF) samples from children and investigated the effect of age on the concentrations and CSF-to-serum ratios of these vitamers. Serum and CSF samples prospectively collected from 49 pediatric patients were analyzed. PLP, PL, and PA were measured using high-performance liquid chromatography with fluorescence detection, using pre-column derivatization by semicarbazide. Effects of age on these vitamers, the PLP-to-PL ratio, CSF-to-serum PLP ratio, and CSF-to-serum PL ratio were evaluated using correlation analysis. The PLP, PL, and PA concentrations in the serum and CSF were higher at younger ages, except for CSF PA concentrations that were mostly below the limit of detection (<1.2nmol/l). The PLP-to-PL ratios in the serum and CSF correlated positively with age. The CSF-to-serum PLP ratio and CSF-to-serum PL ratio were independent of age. Age-related changes in PLP, PL, and PA in serum and in CSF from pediatric patients and CSF-to-serum ratios of PLP and PL demonstrated in this study will provide valuable information for evaluating PLP supply to the central nervous system from the peripheral blood. Copyright © 2017 Elsevier B.V. All rights reserved.

Lack of Sarcocystis neurona antibody response in Virginia opossums (Didelphis virginiana) fed Sarcocystis neurona-infected muscle tissue.

PubMed

Cheadle, M A; Lindsay, D S; Greiner, E C

2006-06-01

Serum was collected from laboratory-reared Virginia opossums (Didelphis virginiana) to determine whether experimentally infected opossums shedding Sarcocystis neurona sporocysts develop serum antibodies to S. neurona merozoite antigens. Three opossums were fed muscles from nine-banded armadillos (Dasypus novemcinctus), and 5 were fed muscles from striped skunks (Mephitis mephitis). Serum was also collected from 26 automobile-killed opossums to determine whether antibodies to S. neurona were present in these opossums. Serum was analyzed using the S. neurona direct agglutination test (SAT). The SAT was modified for use with a filter paper collection system. Antibodies to S. neurona were not detected in any of the serum samples from opossums, indicating that infection in the opossum is localized in the small intestine. Antibodies to S. neurona were detected in filter-paper-processed serum samples from 2 armadillos naturally infected with S. neurona.

Assessment of aflatoxin exposure using serum and urinary biomarkers in São Paulo, Brazil: A pilot study.

PubMed

Jager, Alessandra V; Tonin, Fernando G; Baptista, Gabriela Z; Souto, Pollyana C M C; Oliveira, Carlos A F

2016-05-01

The aim of this study was to evaluate the human exposure of individuals from Pirassununga, Brazil, to dietary aflatoxins B1 (AFB1) and M1 (AFM1) by determination of serum AFB1-lysine and urinary aflatoxin biomarkers (AFM1 and AFB1-N(7)-guanine). The participants were recruited among employees from a Campus of the University of São Paulo, which provided food samples from their homes, as well as serum and urine samples four times every three months, from June 2011 until March 2012. The probable daily intake (PDI) of aflatoxin was estimated by using the results from analysis of food products collected by the time of samples collection, and data from a 24-hour dietary recall questionnaire. Analyses of AFB1 and AFM1 in food samples were conducted by high-performance liquid chromatography with fluorescence detection. Biomarkers in serum and urine were determined by tandem mass spectrometry. AFB1 and AFM1 were detected in 38 samples of cereals (28%, N=136) and 31 milk products (36%, N=86), respectively. AFB1-lysine and AFB1-N(7)-guanine and were not detected in serum or urine samples, respectively. However, AFM1 was found in 74 urine samples (65%), at mean levels in the 4 sampling times ranging from 0.37±0.23 to 1.70±2.88pg/mg creatinine. The mean PDI varied among different sampling times, ranging from 0.09±0.09 to 1.35±5.98ng/kg body weight/day. A modest though significant correlation (r=0.45; p=0.03; N=23) was found for the first time in Brazil between the AFM1 concentration in urine and the PDI for total aflatoxins (AFB1+AFM1) in sampling 1 (June 2011). Urinary AFM1 was confirmed as very sensitive for monitoring the human exposure to dietary aflatoxin. Further studies using serum and urinary biomarkers are needed to estimate the aflatoxin exposure of populations in higher risk areas in Brazil. Copyright © 2015 Elsevier GmbH. All rights reserved.

Distinction of leukemia patients' and healthy persons' serum using FTIR spectroscopy

NASA Astrophysics Data System (ADS)

Sheng, Daping; Liu, Xingcun; Li, Weizu; Wang, Yuchan; Chen, Xianliang; Wang, Xin

2013-01-01

In this paper, FTIR spectroscopy was applied to compare the serum from leukemia patients with the serum from healthy persons. IR spectra of leukemia patients' serum were similar with IR spectra of healthy persons' serum, and they were all made up of proteins, lipids and nucleic acids, etc. In order to identify leukemia patients' serum and healthy persons' serum, the H1075/H1542, H1045/H1467, H2959/H2931 ratios were measured. The H2959/H2931 ratio had the highest significant difference among these ratios and might be a useful factor for identifying leukemia patients' serum and healthy persons' serum. Furthermore, from curve fitting, the RNA/DNA (A1115/A1028) ratios were observed to be lower in leukemia patients' serum than those in healthy persons' serum. The results indicated FTIR spectroscopic study of serum might be a useful tool in the field of leukemia research and diagnosis.

SELDI-TOF-MS proteomic profiling of serum, urine, and amniotic fluid in neural tube defects.

PubMed

Liu, Zhenjiang; Yuan, Zhengwei; Zhao, Qun

2014-01-01

Neural tube defects (NTDs) are common birth defects, whose specific biomarkers are needed. The purpose of this pilot study is to determine whether protein profiling in NTD-mothers differ from normal controls using SELDI-TOF-MS. ProteinChip Biomarker System was used to evaluate 82 maternal serum samples, 78 urine samples and 76 amniotic fluid samples. The validity of classification tree was then challenged with a blind test set including another 20 NTD-mothers and 18 controls in serum samples, and another 19 NTD-mothers and 17 controls in urine samples, and another 20 NTD-mothers and 17 controls in amniotic fluid samples. Eight proteins detected in serum samples were up-regulated and four proteins were down-regulated in the NTD group. Four proteins detected in urine samples were up-regulated and one protein was down-regulated in the NTD group. Six proteins detected in amniotic fluid samples were up-regulated and one protein was down-regulated in the NTD group. The classification tree for serum samples separated NTDs from healthy individuals, achieving a sensitivity of 91% and a specificity of 97% in the training set, and achieving a sensitivity of 90% and a specificity of 97% and a positive predictive value of 95% in the test set. The classification tree for urine samples separated NTDs from controls, achieving a sensitivity of 95% and a specificity of 94% in the training set, and achieving a sensitivity of 89% and a specificity of 82% and a positive predictive value of 85% in the test set. The classification tree for amniotic fluid samples separated NTDs from controls, achieving a sensitivity of 93% and a specificity of 89% in the training set, and achieving a sensitivity of 90% and a specificity of 88% and a positive predictive value of 90% in the test set. These suggest that SELDI-TOF-MS is an additional method for NTDs pregnancies detection.

Urine phenobarbital drug screening: potential use for compliance assessment in neonates.

PubMed

Guillet, Ronnie; Kwon, Jennifer M; Chen, Sixaio; McDermott, Michael P

2012-02-01

This study was done to determine if urine phenobarbital measurements provide a reliable indicator of presence of the drug in neonates. Urine was collected from neonates treated with phenobarbital for clinical indications within 4 to 6 hours of clinically indicated collection of serum phenobarbital levels. Urine samples were also collected from control neonates not treated with phenobarbital. One aliquot was assayed fresh, another frozen at -30°C and assayed 1 to 3 months later. Phenobarbital was assayed using the ONLINE TDM Roche/Hitachi automated clinical chemistry analyzer. Serum and urine concentrations were compared as were fresh and frozen urine measurements. Serum phenobarbital ranged from 5.6 to 52.7 μg/mL. Matched urine samples were 56.6 ± 12.5% of the serum level. Frozen samples were 98.3 ± 8.0% of the fresh samples. Urine phenobarbital concentrations, either fresh or frozen, can be used in neonates as a noninvasive estimate of drug levels.

Occurrence of West Nile virus antibodies in wild birds, horses, and humans in Poland.

PubMed

Niczyporuk, Jowita Samanta; Samorek-Salamonowicz, Elżbieta; Lecollinet, Sylvie; Pancewicz, Sławomir Andrzej; Kozdruń, Wojciech; Czekaj, Hanna

2015-01-01

Serum samples of 474 wild birds, 378 horses, and 42 humans with meningitis and lymphocytic meningitis were collected between 2010 and 2014 from different areas of Poland. West Nile virus (WNV) antibodies were detected using competition enzyme linked immunosorbent assays: ELISA-1 ID Screen West Nile Competition, IDvet, ELISA-2 ID Screen West Nile IgM Capture, and ELISA-3 Ingezim West Nile Compac. The antibodies were found in 63 (13.29%) out of 474 wild bird serum samples and in one (0.26%) out of 378 horse serum samples. Fourteen (33.33%) out of 42 sera from patients were positive against WNV antigen and one serum was doubtful. Positive samples obtained in birds were next retested with virus microneutralisation test to confirm positive results and cross-reactions with other antigens of the Japanese encephalitis complex. We suspect that positive serological results in humans, birds, and horses indicate that WNV can be somehow closely related with the ecosystem in Poland.

Occurrence of West Nile Virus Antibodies in Wild Birds, Horses, and Humans in Poland

PubMed Central

Niczyporuk, Jowita Samanta; Samorek-Salamonowicz, Elżbieta; Lecollinet, Sylvie; Pancewicz, Sławomir Andrzej; Kozdruń, Wojciech; Czekaj, Hanna

2015-01-01

Serum samples of 474 wild birds, 378 horses, and 42 humans with meningitis and lymphocytic meningitis were collected between 2010 and 2014 from different areas of Poland. West Nile virus (WNV) antibodies were detected using competition enzyme linked immunosorbent assays: ELISA-1 ID Screen West Nile Competition, IDvet, ELISA-2 ID Screen West Nile IgM Capture, and ELISA-3 Ingezim West Nile Compac. The antibodies were found in 63 (13.29%) out of 474 wild bird serum samples and in one (0.26%) out of 378 horse serum samples. Fourteen (33.33%) out of 42 sera from patients were positive against WNV antigen and one serum was doubtful. Positive samples obtained in birds were next retested with virus microneutralisation test to confirm positive results and cross-reactions with other antigens of the Japanese encephalitis complex. We suspect that positive serological results in humans, birds, and horses indicate that WNV can be somehow closely related with the ecosystem in Poland. PMID:25866767

[Establishment of A1E3 and B1C4 monoclonal antibody-based ELISA for detecting circulating antigen of Schistosoma japonicum and its preliminary application].

PubMed

Cai, Yu-Chun; Chen, Shao-Hong; Tian, Li-Guang; Chu, Yan-Hong; Lu, Yan; Chen, Mu-Xin; Ai, Lin; Zhou, Yang; Chen, Jia-Xu

2014-02-01

To establish A1E3 and B1C4 monoclonal antibody-based ELISA for detecting circulating antigen of Schistosoma japonicum and explore its application value in the field. The characteristics of A1E3 and B1C4 monoclonal antibodies were analyzed by SDS-PAGE and Western blotting. The SEA-based ELISA was used to evaluate the titers of A1E3 and B1C4. The orthogonal test was used to determine the best concentration of coating antibody B1C4 and optimal working concentration of A1E3-HRP. Under the optimal conditions, the serum samples of 20 acute schistosomiasis cases, 46 chronic schistosomiasis cases, and 20 control sera were tested to evaluate its detection sensitivity and specificity. Seventy-two antibody positive serum samples from Jiangling County of Hubei Province were detected and compared to a commercially available ELISA kit, to evaluate the detection effects of this method. The results of SDS-PAGE demonstrated that the purified A1E3 and B1C4 contained a clear heavy chain with molecular weight of 88,000 and 52,000 respectively and had the same light chain with molecular weight of 20,000; while Western blotting demonstrated that A1E3 and B1C4 could be recognized by SEA and serum samples of acute schistosomiasis cases. The SEA-based ELISA demonstrated the titers of B1C4 and A1E3 were 1:10(5) and 1:30,000, respectively. The serum samples from all the acute cases and 86.9% of the chronic cases showed a positive reaction. All of the control sera from healthy persons gave a negative response. The positive rates of the double monoclonal antibody ELISA and commercial ELISA for detecting the circulating antigen were 45.8% and 43.1% respectively, and there was no significant difference between the results of the two methods. A1E3 and B1C4 monoclonal antibody-based ELISA is established successfully. It exhibits a high sensitivity and specificity in detecting circulating antigen of Schistosoma japonicum.

Analysis of the low molecular weight serum peptidome using ultrafiltration and a hybrid ion trap-Fourier transform mass spectrometer.

PubMed

Zheng, Xiaoyang; Baker, Haven; Hancock, William S

2006-07-07

Advances in proteomics are continuing to expand the ability to analyze the serum proteome. In recent years, it has been realized that in addition to the circulating proteins, human serum also contains a large number of peptides. Many of these peptides are believed to be fragments of larger proteins that have been at least partially degraded by various enzymes such as metalloproteases. Identifying these peptides from a small amount of serum/plasma is difficult due to the complexity of the sample, the low levels of these peptides, and the difficulties in getting a protein identification from a single peptide. In this study, we modified previously published protocols for using centrifugal ultrafiltration, and unlike past studies did not digest the filtrate with trypsin with the intent of identifying endogenous peptides with this method. The filtrate fraction was concentrated and analyzed by a reversed phase-high performance liquid chromatography system connected to a nanospray ionization hybrid ion trap-Fourier transform mass spectrometer (LTQ-FTMS). The mass accuracy of this instrument allows confidence for identifying the protein precursors by a single peptide. The utility of this approach was demonstrated by the identification of over 300 unique peptides with 2 ppm or better mass accuracy per serum sample. With confident identifications, the origin and function of native serum peptides can be more seriously explored. Interestingly, over 34 peptide ladders were observed from over 17 serum proteins. This indicates that a cascade of proteolytic processes affects the serum peptidome. To examine whether this result was an artifact of serum, matched plasma and serum samples were analyzed with similar peptide ladders found in each.

Evaluation of two new commercial tests for the diagnosis of acute dengue virus infection using NS1 antigen detection in human serum.

PubMed

Dussart, Philippe; Petit, Laure; Labeau, Bhety; Bremand, Laetitia; Leduc, Alexandre; Moua, David; Matheus, Séverine; Baril, Laurence

2008-08-20

We compared the performance of two new commercial tests for the detection of dengue NS1 protein during the clinical phase of dengue virus (DENV) infection-an immunochromatographic test allowing rapid detection of the NS1 antigen, Dengue NS1 Ag STRIP (Bio-Rad Laboratories - Marnes La Coquette, France), and a two-step sandwich-format microplate enzyme-linked immunosorbent assay (ELISA), pan-E Dengue Early ELISA (Panbio - Brisbane, Australia)-with a one-step sandwich-format microplate ELISA, the Platelia Dengue NS1 Ag test (Bio-Rad). We tested 272 serum samples from patients with dengue disease. Of these, 222 were from patients with acute infection of one of the four dengue serotypes, detected by RT-PCR and/or virus isolation. Forty-eight acute-phase serum samples from patients not infected with dengue virus were also included. The sensitivity of the Platelia Dengue NS1 Ag test on acute serum samples (n = 222) was 87.4% (95% confidence interval: 82.3% to 91.5%); that of Dengue NS1 Ag STRIP was 81.5% (95% CI: 75.8% to 86.4%) after 15 minutes and 82.4% (95% CI: 76.8% to 87.2%) after 30 minutes. Both tests had a specificity of 100% (97.5% CI, one-sided test: 92.6% to 100.0%). The pan-E Dengue Early ELISA had a sensitivity of 60.4% (95% CI: 53.4% to 66.8%) and a specificity of 97.9% (95% CI: 88.9% to 99.9%). Our findings support the use of diagnostic tools based on the NS1 antigen detection for the diagnosis of acute DENV infection. The immunochromatographic test, Dengue NS1 Ag STRIP-the first rapid diagnostic test for DENV infection-was highly sensitive and specific, and would therefore be a suitable first-line test in the field. The pan-E Dengue Early ELISA was less sensitive than the Platelia test; this two-step ELISA should be combined with DENV IgM antibody detection for the diagnosis of DENV infection.

Large-scale identification of core-fucosylated glycopeptide sites in pancreatic cancer serum using mass spectrometry.

PubMed

Tan, Zhijing; Yin, Haidi; Nie, Song; Lin, Zhenxin; Zhu, Jianhui; Ruffin, Mack T; Anderson, Michelle A; Simeone, Diane M; Lubman, David M

2015-04-03

Glycosylation has significant effects on protein function and cell metastasis, which are important in cancer progression. It is of great interest to identify site-specific glycosylation in search of potential cancer biomarkers. However, the abundance of glycopeptides is low compared to that of nonglycopeptides after trypsin digestion of serum samples, and the mass spectrometric signals of glycopeptides are often masked by coeluting nonglycopeptides due to low ionization efficiency. Selective enrichment of glycopeptides from complex serum samples is essential for mass spectrometry (MS)-based analysis. Herein, a strategy has been optimized using LCA enrichment to improve the identification of core-fucosylation (CF) sites in serum of pancreatic cancer patients. The optimized strategy was then applied to analyze CF glycopeptide sites in 13 sets of serum samples from pancreatic cancer, chronic pancreatitis, healthy controls, and a standard reference. In total, 630 core-fucosylation sites were identified from 322 CF proteins in pancreatic cancer patient serum using an Orbitrap Elite mass spectrometer. Further data analysis revealed that 8 CF peptides exhibited a significant difference between pancreatic cancer and other controls, which may be potential diagnostic biomarkers for pancreatic cancer.

Microchip systems for immunoassay: an integrated immunoreactor with electrophoretic separation for serum theophylline determination.

PubMed

Chiem, N H; Harrison, D J

1998-03-01

A glass microchip is described in which reagents and serum samples for competitive immunoassay of serum theophylline can be mixed, reacted, separated, and analyzed. The device functions as an automated microfluidic immunoassay system, creating a lab-on-a-chip. Electroosmotic pumping was used to control first the mixing of 50-fold-diluted serum sample with labeled theophylline tracer in a 1:1 ratio, followed by 1:1 mixing and reaction with anti-theophylline antibody. The 51-nL on-chip mixer gave the same concentration as dilution performed off-chip, within 3%. A 100-pL plug of the reacted solution was then injected into an electrophoresis separation channel integrated within the same chip. Measurements of free and bound tracer by fluorescence detection gave linear calibration curves of signal vs log[theophylline] between 0 and 40 mg/L, with a slope of 0.52 +/- 0.03 and an intercept of -0.04 +/- 0.04 after a 90-s reaction time. A detection limit of 0.26 mg/L in serum (expressed before the dilution step, actual concentration of 1.3 micrograms/L at the detector) was obtained. Recovery values were 107% +/- 8% for 15 mg/L serum samples.

High-performance liquid chromatographic determination of methotrexate, 7-hydroxymethotrexate, 5-methyltetrahydrofolic acid and folinic acid in serum and cerebrospinal fluid.

PubMed

Belz, S; Frickel, C; Wolfrom, C; Nau, H; Henze, G

1994-11-04

A method for the simultaneous determination of the antifolates methotrexate and 7-hydroxymethotrexate as well as the folates 5-methyltetrahydrofolic acid and folinic acid (5-formyltetrahydrofolic acid) in serum and cerebrospinal fluid (CSF) is described. High-performance liquid chromatography with gradient elution and dual detection (ultraviolet absorption and fluorescence) was used to separate and quantitate the analytes. Serum samples containing high levels of the substances of interest and CSF samples were injected directly onto the HPLC column. For determination of low concentrations, serum samples were subjected to a solid-phase extraction method for clean-up and concentration purposes. The determination limits were 10 ng/ml for both antifolates, 100 ng/ml for folinic acid, and 0.1 ng/ml for the physiologically occurring methylated folate which is about 1/100 the serum concentration in healthy children. The suitability of the method for pharmacokinetic monitoring of high-dose methotrexate therapy combined with leucovorin rescue administered to children with acute lymphoblastic leukemia was demonstrated. Minimum values of the serum folate during treatment ranged from 0.2 to 3.1 ng/ml. Even those very low concentrations could be reliably measured.

A sup 125 I-radioimmunoassay for measuring androstenedione in serum and in blood-spot samples from neonates

DOE Office of Scientific and Technical Information (OSTI.GOV)

Thomson, S.; Wallace, A.M.; Cook, B.

1989-08-01

We developed a radioimmunoassay with a gamma-emitting radioligand to measure androstenedione in human serum and in dried blood-spot samples from newborns. Antisera were raised in rabbits against androstenedione linked to bovine serum albumin at positions 3, 6, or 11 on the steroid nucleus. Radioligands were prepared by linking ({sup 125}I)iodohistamine at positions 3, 6, or 11. Linkages were through either carboxymethyloxime or hemisuccinate bridges. All label and antibody combinations were examined, and the most sensitive and specific combination (antiserum raised against androstenedione-3-carboxymethyloxime-bovine serum albumin with an androstenedione-carboxymethyloxime-({sup 125}I)iodohistamine label) was selected for full evaluation. We report the performance of thesemore » selected reagents in an immunoassay for androstenedione in both serum and dried blood-spot samples from neonates. We measured concentrations of androstenedione in serum under normal and pathological conditions such as congenital adrenal hyperplasia and polycystic ovarian disease. Diurnal variation in normal men was observed. Androstenedione was measured in blood spots from neonates born at term or prematurely, with respiratory distress syndrome, or with congenital adrenal hyperplasia.« less

Effect of the conditions of isolation on the physicochemical properties of human serum albumin in the norm and with pathology

NASA Astrophysics Data System (ADS)

Ivanov, A. I.; Zhbankov, R. G.; Korolenko, E. A.; Korolik, E. V.; Meleshchenko, L. A.; Sarnatskaya, V. V.; Nikolaev, V. G.; Nikolaichik, V. V.; Yushko, L. A.

1997-01-01

Differential scanning calorimetry and IR spectrosocopy were used to investigate the effect of the procedure of isolation of human serum albumin on its physicochemical characteristics. It is shown that fractionation of blood plasma with ethylene glycol followed by ion exchange chromatography can be used to obtain albumin of normal donors that is similar to the albumin in the nonfractionated plasma according to melting thermograms. Endotherms of human serum albumin samples that were obtained by affinity chromatography and preparative electrophoresis are bimodal, unlike the monophasic for albumin obtained by polyethylene glycol precipitation. These changes result from a higher content of nonetherified fatty acids in the albumin samples obtained by affinity chromatography and from modification of the secondary protein structure in the samples obtained by electrophoresis. Analysis of melting thermograms of serum albumin from patients with uremia, chronic hepatitis, and peritonitis shows that fractionation of blood with polyethylene glycol preserves the thermodynamic characteristics of the various pathological serum albumins to the greatest extent. The present results demonstrate the advantage of polyethylene glycol fractionation for isolation of native preparations of normal and “pathological” human serum albumin.

Validation of two commercial real-time RT-PCR kits for rapid and specific diagnosis of classical swine fever virus.

PubMed

Le Dimna, M; Vrancken, R; Koenen, F; Bougeard, S; Mesplède, A; Hutet, E; Kuntz-Simon, G; Le Potier, M F

2008-01-01

Two real-time RT-PCR kits, developed by LSI (TaqVet CSF) and ADIAGENE (Adiavet CSF), obtained an agreement to be commercialised in France, subject to conditions, defined by the French Classical Swine Fever (CSF) National Reference Laboratory. The producers were asked to introduce an internal control to check the RNA extraction efficacy. The different criteria assessed were sensitivity, "pestivirus specificity", reproducibility and ease of handling, using 189 different samples. These samples were either CSFV inactivated strains or blood/serum/organs collected from CSFV experimentally infected pigs or naturally infected wild boars. The reproducibility of the assays was confirmed by the analysis of a batch-to-batch panel control that was used for inter-laboratory tests involving nine laboratories. The two kits were also tested for the use in mass diagnostics and the results proved the kits to be suited using pools of blood, serum and tonsils. Moreover, a field evaluation, carried out on spleen samples collected from the CSF surveillance of wild boars in an area known to be infected and from domestic pigs at a slaughterhouse, confirmed the high sensitivity and specificity of the two kits. This step-by-step evaluation procedure confirmed that the two commercial CSF real-time RT-PCR kits have a higher predictive value than the current diagnostic standard, Virus Isolation.

Aluminium (Al) speciation in serum and urine after subcutaneous venom immunotherapy with Al as adjuvant.

PubMed

Michalke, Bernhard; Kramer, Matthias F; Brehler, Randolf

2018-02-21

Aluminium is associated with disorders and is the commonly used vaccine adjuvant. Understanding the mechanisms of how Al is transported, metabolized or of its toxicity depends on the knowledge of Al-interactions with bioligands, i.e. Al-species. Al-speciation in serum is difficult because of low concentration and the risk of exogenous Al contamination. Furthermore, Al-measurements may be hampered according to various interferences. This study aims for developing quality controlled protocols for reliable Al- and Al-species determination and for investigating probable differences in Al (-speciation) after Al-containing subcutaneous immunotherapy (SIT). Sample donors were recruited either for the control group ("class-0", they never had been treated with SIT containing an Al-depot extract) or for the SIT-group ("class-1", they previously had been treated with SIT for insect venom allergy with an Al-depot extract). Blood was drawn for medical reasons and serum prepared. Additionally, some sample donors collected 24-h-urine. They had been informed (and they consented) about the scientific use of their samples. The study was approved by the ethic committee of the "Medical Association Westphalia-Lippe" and of the University of Münster, evaluating the study positively (No. 2013-667-f-S). We applied quality controlled sample preparation and interference-free Al detection by ICP sectorfield-mass spectrometry. Al-species were analysed using size-exclusion-chromatography-ICP-qMS. Al-concentrations or speciation in urine samples showed no differences between class-0 and class-1. Al-citrate was the main uric Al-species. In serum elevated Al-concentrations were found for both classes, with class-1 samples being significantly higher than class-0 (p = 0.041), but class-0 samples being approximately 10-fold too high compared to reference values from non-exposed persons. We identified gel-monovettes as contamination source. In contamination-free samples from HNO 3 -prewashed gel-free monovettes (n = 27) there was no difference in the serum Al concentration between the two patient groups (p = 0.669) INTERPRETATION: Thorough cleaning of sample preparation ware and use of gel-free monovettes is decisive for an accurate Al analysis in serum. Without these steps, wrong analysis and wrong conclusions are likely. We conclude that gel-monovettes are unsuitable for blood sampling with subsequent Al-analysis. Whether Al in serum is elevated after SIT treatment containing an Al-depot extract, or not, remains inconclusive as the non-contaminated sample size was small. Copyright © 2018 Elsevier GmbH. All rights reserved.

Tamoxifen dose and serum concentrations of tamoxifen and six of its metabolites in routine clinical outpatient care.

PubMed

Jager, N G L; Rosing, H; Schellens, J H M; Linn, S C; Beijnen, J H

2014-02-01

A sensitive and selective HPLC-MS/MS assay was used to analyze steady-state serum concentrations of tamoxifen, N-desmethyltamoxifen (E)-endoxifen, (Z)-endoxifen, N-desmethyl-4'-hydroxytamoxifen, 4-hydroxytamoxifen, and 4'-hydroxytamoxifen to support therapeutic drug monitoring (TDM) in patients treated with tamoxifen according to standard of care. When the (Z)-endoxifen serum concentration was below the predefined therapeutic threshold concentration of 5.9 ng/mL, the clinician was advised to increase the tamoxifen dose and to collect another serum sample. Paired serum samples from patients at one dose level at different time points during the tamoxifen treatment were used to assess the intra-patient variability. A total of 251 serum samples were analyzed, obtained from 205 patients. Of these patients, 197 used 20 mg tamoxifen per day and 8 patients used 10 mg/day. There was wide variability in tamoxifen and metabolite concentrations within the dosing groups. The threshold concentration for (Z)-endoxifen was reached in one patient (12 %) in the 10 mg group, in 153 patients (78 %) in the 20 mg group, and in 26 (96 %) of the patients who received a dose increase to 30 or 40 mg/day. Dose increase from 20 to 30 or 40 mg per day resulted in a significant increase in the mean serum concentrations of all analytes (p < 0.001). The mean intra-patient variability was between 10 and 20 % for all analytes. These results support the suitability of TDM for optimizing the tamoxifen treatment. It is shown that tamoxifen dose is related to (Z)-endoxifen exposure and increasing this dose leads to a higher serum concentration of tamoxifen and its metabolites. The low intra-patient variability suggests that only one serum sample is needed for TDM, making this a relatively noninvasive way to optimize the patient's treatment.

Comparison of lacosamide concentrations in cerebrospinal fluid and serum in patients with epilepsy.

PubMed

May, Theodor W; Brandt, Christian; Helmer, Renate; Bien, Christian G; Cawello, Willi

2015-07-01

This study was carried out to estimate the exposure of the central nervous system (CNS) to the antiepileptic drug (AED) lacosamide, under steady state conditions, in patients with epilepsy who take oral lacosamide alongside up to three other AEDs. Twenty-seven serum and cerebral spinal fluid (CSF) samples were collected from 21 patients receiving lacosamide for the treatment of epilepsy (50-600 mg/day over two or three doses). This included 23 time-matched pairs of serum and CSF samples from 19 patients. The concentration of lacosamide in each sample was determined using high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). Linear regression was used to characterize the relationship between the CSF-to-serum ratio of lacosamide concentration and the time since dosing, the daily lacosamide dose, or the daily dose normalized by volume of distribution (Vd , approximated to total body water), and between the drug concentrations in each compartment (CSF vs. serum). Concentrations of lacosamide in CSF (mean ± standard deviation [SD] 7.37 ± 3.73 μg/ml, range 1.24-14.95, n = 27) and serum (mean ± SD 8.16 ± 3.82 μg/ml, range 2.29-15.45, n = 27) samples showed a good correlation over the dose range investigated. The mean CSF-to-serum ratio of lacosamide concentrations was 0.897 ± 0.193 (range 0.492-1.254, n = 23 time-matched pairs) and was independent of lacosamide dose. Drug concentrations in the CSF are often used to indicate those in the brain interstitial fluid. In patients with epilepsy who follow a stable oral AED dosing regimen, lacosamide concentration in CSF is approximately 85% of that found in serum, suggesting that serum may be a valuable indicator of lacosamide concentration in the CNS. Wiley Periodicals, Inc. © 2015 International League Against Epilepsy.

Evaluation of new monoclonal antibody-based latex agglutination test for detection of cryptococcal polysaccharide antigen in serum and cerebrospinal fluid.

PubMed Central

Kiska, D L; Orkiszewski, D R; Howell, D; Gilligan, P H

1994-01-01

We evaluated the performance of CRYPTO-LEX (Trinity Laboratories, Inc., Raleigh, N. C.), a new mouse immunoglobulin M monoclonal antibody latex agglutination reagent which reacts with the capsular polysaccharide of the four serogroups of Cryptococcus neoformans. This test was compared with CALAS (Meridian Diagnostics, Cincinnati, Ohio) for the ability to detect cryptococcal antigen in serum and cerebrospinal fluid (CSF). A total of 580 clinical specimens (327 serum and 253 CSF samples), primarily from human immunodeficiency virus-infected patients, were tested in this study. Sixty-seven specimens (44 serum and 23 CSF samples) were positive for cryptococcal antigen with both tests, and 511 (282 serum and 229 CSF samples) were negative. The two latex reagents agreed for 326 of 327 serum specimens (44 positives and 282 negatives). One serum specimen with a titer of 1:2 was CALAS positive but CRYPTO-LEX negative. The titer correlation coefficient for the two tests was 0.884 when two highly discordant serum specimens were eliminated from analysis of the data. The two latex tests agreed for 252 of 253 CSF specimens (23 positives and 229 negatives). One specimen with a titer of 1:2 was positive with CALAS and negative by CRYPTO-LEX. The correlation coefficient of the two tests for CSF titers was 0.886. The sensitivity and specificity of CRYPTO-LEX were 97 and 100%, respectively, with a 99.6% correlation with CALAS. These data show that the performance of CRYPTO-LEX is comparable to that of CALAS for detection of cryptococcal antigen in serum and CSF. PMID:7814566

[Clinical evaluation of a novel HBsAg quantitative assay].

PubMed

Takagi, Kazumi; Tanaka, Yasuhito; Naganuma, Hatsue; Hiramatsu, Kumiko; Iida, Takayasu; Takasaka, Yoshimitsu; Mizokami, Masashi

2007-07-01

The clinical implication of the hepatitis B surface antigen (HBsAg) concentrations in HBV-infected individuals remains unclear. The aim of this study was to evaluate a novel fully automated Chemiluminescence Enzyme Immunoassay (Sysmex HBsAg quantitative assay) by comparative measurements of the reference serum samples versus two independent commercial assays (Lumipulse f or Architect HBsAg QT). Furthermore, clinical usefulness was assessed for monitoring of the serum HBsAg levels during antiviral therapy. A dilution test using 5 reference-serum samples showed linear correlation curve in range from 0.03 to 2,360 IU/ml. The HBsAg was measured in total of 400 serum samples and 99.8% had consistent results between Sysmex and Lumipulse f. Additionally, a positive linear correlation was observed between Sysmex and Architect. To compare the Architect and Sysmex, both methods were applied to quantify the HBsAg in serum samples with different HBV genotypes/subgenotypes, as well as in serum contained HBV vaccine escape mutants (126S, 145R). Correlation between the methods was observed in results for escape mutants and common genotypes (A, B, C) in Japan. Observed during lamivudine therapy, an increase in HBsAg and HBV DNA concentrations preceded the aminotransferase (ALT) elevation associated with drug-resistant HBV variant emergence (breakthrough hepatitis). In conclusion, reliability of the Sysmex HBsAg quantitative assay was confirmed for all HBV genetic variants common in Japan. Monitoring of serum HBsAg concentrations in addition to HBV DNA quantification, is helpful in evaluation of the response to lamivudine treatment and diagnosis of the breakthrough hepatitis.

Can nail, hair and urine be used for biomonitoring of human exposure to perfluorooctane sulfonate and perfluorooctanoic acid?

PubMed

Li, Jingguang; Guo, Feifei; Wang, Yuxin; Zhang, Jialing; Zhong, Yuxin; Zhao, Yunfeng; Wu, Yongning

2013-03-01

Because of the disadvantages of invasive sampling, it is desirable to explore non-invasive matrices for human biomonitoring of perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA). The aim of this study was to evaluate the application of nail, hair and urine for human biomonitoring of PFOS and PFOA. The concentrations of PFOS and PFOA in matched nail, hair, urine and serum samples collected from 64 donors were measured. The chemicals of interest were detected with high detection frequency in these matrices (90%-100%) except for PFOA in urine samples (56%). Generally, the gender influences on the levels of PFOS and PFOA in these non-invasive matrices were in agreement with that in serum. For PFOS, the coefficients of Spearman correlation between serum samples and nail, hair and urine samples were 0.786 (p<0.001), 0.545 (p<0.001) and 0.302 (p<0.05), respectively. For PFOA, the correlation was only observed between nail samples and serum samples with a correlation coefficient of 0.299 (p<0.05). The results suggested that nail has more potential than hair and urine to be applied in human biomonitoring for PFOS and PFOA in general populations. Copyright © 2013 Elsevier Ltd. All rights reserved.

Effects of electromagnetic pulse on serum element levels in rat.

PubMed

Li, Kangchu; Ma, Shirong; Ren, Dongqing; Li, Yurong; Ding, Guirong; Liu, Junye; Guo, Yao; Guo, Guozhen

2014-04-01

Electromagnetic pulse (EMP) was a potentially harmful factor to the human body, and a biological dosimetry to evaluate effects of EMP is necessary. Little is known about effects of EMP on concentration of macro and trace elements in serum so far. In this study, Sprague-Dawley rats were randomly divided into 50-kV/m EMP-exposed group (n = 10), 100-kV/m EMP-exposed group (n = 10), 200-kV/m EMP-exposed group (n = 40), and the sham-exposed group (n = 20). The macro and trace element concentrations in serum were examined at 6, 12, 24, and 48 h after EMP exposure at different electric field intensities. Compared with the sham-exposed groups, the concentration of sodium (Na), potassium (K), magnesium (Mg), calcium (Ca), zinc (Zn), copper (Cu), iron (Fe), selenium (Se), and manganese (Mn) in rat serum was not changed significantly within 48 h after 200 pulses of EMP exposure at electric field intensity of 50, 100, and 200 kV/m although the K level was decreased and the Ca level was increased with the electric field intensity of EMP increasing. In addition, there was a tendency that the Zn level was decreased with the time going on within 48 h after EMP exposure. Under our experimental conditions, EMP exposure cannot affect the concentration of macro and trace elements in rat serum. There was no time-effect or dose-effect relationship between EMP exposure and serum element levels. The macro and trace elements in serum are not suitable endpoints of biological dosimetry of EMP.

Molecular epidemiology of malaria in Cameroon. XV. Experimental studies on serum substitutes and supplements and alternative culture media for in vitro drug sensitivity assays using fresh isolates of Plasmodium falciparum.

PubMed

Basco, Leonardo K

2003-08-01

In vitro drug sensitivity assay is an important tool for various on-going studies aiming to establish the correlation between candidate molecular markers for drug resistance and drug response in laboratory-adapted strains and field isolates of Plasmodium falciparum. A widespread use of this technique in the field would require a suitable substitute that can replace human serum. In this study, several alternative sources of serum substitutes and supplements were evaluated for their capacity to sustain parasite growth for a single life cycle and their compatibility with in vitro assays for clinical isolates that have not been adapted to in vitro culture. Albumax, a commercial preparation of lipid-enriched bovine albumin, did not support parasite growth as much as human serum and fetal calf serum in several isolates. Other serum supplements (AmnioMax and Ultroser) supported parasite growth relatively well. The 50% inhibitory concentrations (IC50s) of chloroquine and antifolates determined with 0.05% Albumax were generally two or three times higher than with human serum. With 10% fetal calf serum, IC50s for chloroquine and antifolates were approximately two times higher and three times lower than with human serum, respectively. The use of AmnioMax and OptiMAb resulted in a greater than two-fold increase in IC50s and several uninterpretable assays. Despite possible batch-to-batch differences, fetal calf serum may be a suitable substitute for in vitro drug assays while awaiting the results of further studies on other serum substitutes and supplements.

Analysis of human serum and whole blood for mineral content by ICP-MS and ICP-OES: development of a mineralomics method.

PubMed

Harrington, James M; Young, Daniel J; Essader, Amal S; Sumner, Susan J; Levine, Keith E

2014-07-01

Minerals are inorganic compounds that are essential to the support of a variety of biological functions. Understanding the range and variability of the content of these minerals in biological samples can provide insight into the relationships between mineral content and the health of individuals. In particular, abnormal mineral content may serve as an indicator of illness. The development of robust, reliable analytical methods for the determination of the mineral content of biological samples is essential to developing biological models for understanding the relationship between minerals and illnesses. This paper describes a method for the analysis of the mineral content of small volumes of serum and whole blood samples from healthy individuals. Interday and intraday precision for the mineral content of the blood (250 μL) and serum (250 μL) samples was measured for eight essential minerals--sodium (Na), calcium (Ca), magnesium (Mg), potassium (K), iron (Fe), zinc (Zn), copper (Cu), and selenium (Se)--by plasma spectrometric methods and ranged from 0.635 to 10.1% relative standard deviation (RSD) for serum and 0.348-5.98% for whole blood. A comparison of the determined ranges for ten serum samples and six whole blood samples provided good agreement with literature reference ranges. The results demonstrate that the digestion and analysis methods can be used to reliably measure the content of these minerals and potentially of other minerals.

Serum and urine metabolomic fingerprinting in diagnostics of inflammatory bowel diseases.

PubMed

Dawiskiba, Tomasz; Deja, Stanisław; Mulak, Agata; Ząbek, Adam; Jawień, Ewa; Pawełka, Dorota; Banasik, Mirosław; Mastalerz-Migas, Agnieszka; Balcerzak, Waldemar; Kaliszewski, Krzysztof; Skóra, Jan; Barć, Piotr; Korta, Krzysztof; Pormańczuk, Kornel; Szyber, Przemyslaw; Litarski, Adam; Młynarz, Piotr

2014-01-07

To evaluate the utility of serum and urine metabolomic analysis in diagnosing and monitoring of inflammatory bowel diseases (IBD). Serum and urine samples were collected from 24 patients with ulcerative colitis (UC), 19 patients with the Crohn's disease (CD) and 17 healthy controls. The activity of UC was assessed with the Simple Clinical Colitis Activity Index, while the activity of CD was determined using the Harvey-Bradshaw Index. The analysis of serum and urine samples was performed using proton nuclear magnetic resonance (NMR) spectroscopy. All spectra were exported to Matlab for preprocessing which resulted in two data matrixes for serum and urine. Prior to the chemometric analysis, both data sets were unit variance scaled. The differences in metabolite fingerprints were assessed using partial least-squares-discriminant analysis (PLS-DA). Receiver operating characteristic curves and area under curves were used to evaluate the quality and prediction performance of the obtained PLS-DA models. Metabolites responsible for separation in models were tested using STATISTICA 10 with the Mann-Whitney-Wilcoxon test and the Student's t test (α = 0.05). The comparison between the group of patients with active IBD and the group with IBD in remission provided good PLS-DA models (P value 0.002 for serum and 0.003 for urine). The metabolites that allowed to distinguish these groups were: N-acetylated compounds and phenylalanine (up-regulated in serum), low-density lipoproteins and very low-density lipoproteins (decreased in serum) as well as glycine (increased in urine) and acetoacetate (decreased in urine). The significant differences in metabolomic profiles were also found between the group of patients with active IBD and healthy control subjects providing the PLS-DA models with a very good separation (P value < 0.001 for serum and 0.003 for urine). The metabolites that were found to be the strongest biomarkers included in this case: leucine, isoleucine, 3-hydroxybutyric acid, N-acetylated compounds, acetoacetate, glycine, phenylalanine and lactate (increased in serum), creatine, dimethyl sulfone, histidine, choline and its derivatives (decreased in serum), as well as citrate, hippurate, trigonelline, taurine, succinate and 2-hydroxyisobutyrate (decreased in urine). No clear separation in PLS-DA models was found between CD and UC patients based on the analysis of serum and urine samples, although one metabolite (formate) in univariate statistical analysis was significantly lower in serum of patients with active CD, and two metabolites (alanine and N-acetylated compounds) were significantly higher in serum of patients with CD when comparing jointly patients in the remission and active phase of the diseases. Contrary to the results obtained from the serum samples, the analysis of urine samples allowed to distinguish patients with IBD in remission from healthy control subjects. The metabolites of importance included in this case up-regulated acetoacetate and down-regulated citrate, hippurate, taurine, succinate, glycine, alanine and formate. NMR-based metabolomic fingerprinting of serum and urine has the potential to be a useful tool in distinguishing patients with active IBD from those in remission.

Serum and urine metabolomic fingerprinting in diagnostics of inflammatory bowel diseases

PubMed Central

Dawiskiba, Tomasz; Deja, Stanisław; Mulak, Agata; Ząbek, Adam; Jawień, Ewa; Pawełka, Dorota; Banasik, Mirosław; Mastalerz-Migas, Agnieszka; Balcerzak, Waldemar; Kaliszewski, Krzysztof; Skóra, Jan; Barć, Piotr; Korta, Krzysztof; Pormańczuk, Kornel; Szyber, Przemyslaw; Litarski, Adam; Młynarz, Piotr

2014-01-01

AIM: To evaluate the utility of serum and urine metabolomic analysis in diagnosing and monitoring of inflammatory bowel diseases (IBD). METHODS: Serum and urine samples were collected from 24 patients with ulcerative colitis (UC), 19 patients with the Crohn’s disease (CD) and 17 healthy controls. The activity of UC was assessed with the Simple Clinical Colitis Activity Index, while the activity of CD was determined using the Harvey-Bradshaw Index. The analysis of serum and urine samples was performed using proton nuclear magnetic resonance (NMR) spectroscopy. All spectra were exported to Matlab for preprocessing which resulted in two data matrixes for serum and urine. Prior to the chemometric analysis, both data sets were unit variance scaled. The differences in metabolite fingerprints were assessed using partial least-squares-discriminant analysis (PLS-DA). Receiver operating characteristic curves and area under curves were used to evaluate the quality and prediction performance of the obtained PLS-DA models. Metabolites responsible for separation in models were tested using STATISTICA 10 with the Mann-Whitney-Wilcoxon test and the Student’s t test (α = 0.05). RESULTS: The comparison between the group of patients with active IBD and the group with IBD in remission provided good PLS-DA models (P value 0.002 for serum and 0.003 for urine). The metabolites that allowed to distinguish these groups were: N-acetylated compounds and phenylalanine (up-regulated in serum), low-density lipoproteins and very low-density lipoproteins (decreased in serum) as well as glycine (increased in urine) and acetoacetate (decreased in urine). The significant differences in metabolomic profiles were also found between the group of patients with active IBD and healthy control subjects providing the PLS-DA models with a very good separation (P value < 0.001 for serum and 0.003 for urine). The metabolites that were found to be the strongest biomarkers included in this case: leucine, isoleucine, 3-hydroxybutyric acid, N-acetylated compounds, acetoacetate, glycine, phenylalanine and lactate (increased in serum), creatine, dimethyl sulfone, histidine, choline and its derivatives (decreased in serum), as well as citrate, hippurate, trigonelline, taurine, succinate and 2-hydroxyisobutyrate (decreased in urine). No clear separation in PLS-DA models was found between CD and UC patients based on the analysis of serum and urine samples, although one metabolite (formate) in univariate statistical analysis was significantly lower in serum of patients with active CD, and two metabolites (alanine and N-acetylated compounds) were significantly higher in serum of patients with CD when comparing jointly patients in the remission and active phase of the diseases. Contrary to the results obtained from the serum samples, the analysis of urine samples allowed to distinguish patients with IBD in remission from healthy control subjects. The metabolites of importance included in this case up-regulated acetoacetate and down-regulated citrate, hippurate, taurine, succinate, glycine, alanine and formate. CONCLUSION: NMR-based metabolomic fingerprinting of serum and urine has the potential to be a useful tool in distinguishing patients with active IBD from those in remission. PMID:24415869

[Presence of west Nile virus in northeast Mexico].

PubMed

Fernández-Salas, Ildefonso; de Lourdes Garza-Rodríguez, María; Beaty, Barry J; Jiménez, Javier Ramos; Rivas-Estilla, Ana María

2007-01-01

To investigate the presence of WNV in birds, horses and humans in northeast Mexico. Serum samples from 33 birds, 24 horses and 237 humans were screened by ELISA for Anti-WNV antibodies. Human serum samples were also screened for WNV RNA using an RT-PCR assay. Positive sera were found in three birds and 15 horses. Forty percent of the human serum samples were positive for IgG antibodies and 0% for IgM antibodies and viral RNA. The results of this study show that WNV is present in northeast Mexico and it is a new emergent infectious agent that represents a challenge for research and prevention programs in Mexico.

Saliva, Serum Levels of Interleukin-21, -33 and Prostaglandin E2 in Patients with Generalised Aggressive or Chronic Periodontitis.

PubMed

Gümüş, Pınar; Nizam, Nejat; Nalbantsoy, Ayşe; Özçaka, Özgün; Buduneli, Nurcan

This cross-sectional study aims to evaluate saliva, serum levels of interleukin-21 (IL-21), IL-33, and prostaglandin E2 (PGE2) in patients with generalised chronic periodontitis or aggressive periodontitis. Before initiation of any periodontal treatment, saliva and serum samples were collected and clinical periodontal measurements were recorded from 94 participants (25 aggressive periodontitis patients, 25 chronic periodontitis patients, 44 periodontally healthy individuals). IL-21, IL-33 and PGE2 levels in serum and saliva samples were determined by ELISA. Data were tested statistically using Kruskal-Wallis, Mann-Whitney U-, and Spearman-rho rank tests. Saliva IL-33 levels were statistically significantly higher in the chronic than the aggressive group (p < 0.05). Serum IL-33, saliva and serum IL-21 and PGE2 levels were similar in the two periodontitis groups. Saliva IL-33 levels correlated with age in the chronic periodontitis group (p < 0.05). Statistically significant positive correlations were found between serum, saliva PGE2 levels and plaque index (p < 0.05). IL-33 and IL-21 levels in serum samples positively correlated in the periodontitis groups (p < 0.05). IL-21 and PGE2 analysis did not exhibit discriminating data between generalised chronic and aggressive periodontitis, but the present findings support the role of these cytokines in periodontitis. Statistically significantly higher saliva IL-33 levels in the chronic periodontitis group warrant further research.

Glyphosate and Paraquat in Maternal and Fetal Serums in Thai Women.

PubMed

Kongtip, Pornpimol; Nankongnab, Noppanun; Phupancharoensuk, Ratanavadee; Palarach, Chonlada; Sujirarat, Dusit; Sangprasert, Supha; Sermsuk, Malasod; Sawattrakool, Namthip; Woskie, Susan Renee

2017-01-01

This longitudinal study measured the glyphosate and paraquat concentrations found in maternal and umbilical cord serum in 82 pregnant women who gave birth in three provinces of Thailand. Through questionnaires and biological samples collected at childbirth, factors such as personal characteristics, family members occupation, agricultural activities, and herbicide use in agricultural work were evaluated as predictors of glyphosate and paraquat levels in the pregnant women. Statistical analysis used univariate and binary multiple logistic regression, where the outcome was the probability of exposure to paraquat or glyphosate above the limit of detection associated with occupation and household factors. The glyphosate concentrations in the pregnant women's serum at childbirth (median: 17.5, range: 0.2-189.1 ng/mL) were significantly higher (P < .007) than those in the umbilical cord serum (median: 0.2, range: 0.2-94.9 ng/mL). However, the paraquat concentrations in the serum of the pregnant women at childbirth (83% ≤limit of detection [LOD], with maximum of 58.3 ng/mL) were similar to those in the umbilical cord serum (80% LOD in serum at childbirth were 11.9 times more likely to report work as an agriculturist (P < .001), 3.7 times more likely to live near agricultural areas (P = .006), and 5.9 times more likely to have a family member who worked in agriculture (P < .001). The only factors affecting paraquat exposures in pregnant women at childbirth were reporting the agricultural activity of digging in farm soil and working in the agricultural fields in the third trimester of pregnancy. These results show that pregnant women who work in agriculture or live in families that work in agriculture have higher exposures to the herbicides glyphosate and paraquat. The potential for long-term health impacts of these prenatal exposures to children should be evaluated, and greater regulation of the sale and use of herbicides should be considered in Thailand.

Use of Paired Serum Samples for Serodiagnosis of Typhoid Fever

PubMed Central

House, Deborah; Chinh, Nguyen T.; Diep, To S.; Parry, Christopher M.; Wain, John; Dougan, Gordon; White, Nicholas J.; Hien, Tran Tinh; Farrar, Jeremy J.

2005-01-01

Using an enzyme-linked immunosorbent assay we demonstrate that, in adult patients with typhoid fever, the sensitivity of a serological test based on the detection of anti-lipopolysaccharide immunoglobulin G is increased when used with paired serum samples taken 1 week apart. PMID:16145168

Alemtuzumab: validation of a sensitive and simple enzyme-linked immunosorbent assay.

PubMed

Jilani, Iman; Keating, Michael; Giles, Francis J; O'Brien, Susan; Kantarjian, Hagop M; Albitar, Maher

2004-12-01

Alemtuzumab (MabCampath) is a humanized rat monoclonal antibody that targets the CD52 antigen. It has been approved for the treatment of patients with resistant chronic lymphocytic leukaemia (CLL). Measuring plasma/serum levels of alemtuzumab is important for optimizing the dosing and scheduling of therapy; however, current assays in serum or plasma, based on the capture of alemtuzumab using CD52, are complicated and difficult to adapt for high throughput testing. We developed a simple sandwich enzyme-linked immunosorbent assay (ELISA) to measure alemtuzumab that takes advantage of the remaining rat sequence in alemtuzumab. Using specific anti-rat immunoglobulin (Ig) antibodies (absorbed against human Ig), alemtuzumab levels were measured in the serum and plasma of patients treated with alemtuzumab. Levels were similar between plasma and serum samples, in fresh samples and samples stored at 4 degrees C for 24 h, but were significantly lower in samples stored at room temperature for 24h. The assay was successfully used to determine serum alemtuzumab pre- and post-treatment. This assay is simple and adaptable for high throughput testing, with a limit of detection of 0.05 microg/ml and a coefficient of variation of +/-12.5%. No false positivity was observed in >200 samples tested. This validated assay should help optimize the dosing and scheduling of alemtuzumab therapy. The underlying principles are also applicable to the measurement of other humanized antibodies using an appropriate anti-Ig.

Iron and ADHD: Time to Move beyond Serum Ferritin Levels

ERIC Educational Resources Information Center

Donfrancesco, Renato; Parisi, Pasquale; Vanacore, Nicola; Martines, Francesca; Sargentini, Vittorio; Cortese, Samuele

2013-01-01

Objective: (a) To compare serum ferritin levels in a sample of stimulant-naive children with ADHD and matched controls and (b) to assess the association of serum ferritin to ADHD symptoms severity, ADHD subtypes, and IQ. Method: The ADHD and the control groups included 101 and 93 children, respectively. Serum ferritin levels were determined with…

Blood collected on filter paper for wildlife serology: detecting antibodies to Neospora caninum, West Nile virus, and five bovine viruses in reindeer.

PubMed

Curry, Patricia S; Ribble, Carl; Sears, William C; Hutchins, Wendy; Orsel, Karin; Godson, Dale; Lindsay, Robbin; Dibernardo, Antonia; Kutz, Susan J

2014-04-01

We compared Nobuto filter paper (FP) whole-blood samples to serum for detecting antibodies to seven pathogens in reindeer (Rangifer tarandus tarandus). Serum and FP samples were collected from captive reindeer in 2008-2009. Sample pairs (serum and FP eluates) were assayed in duplicate at diagnostic laboratories with the use of competitive enzyme-linked immunosorbent assays (cELISAs) for Neospora caninum and West Nile virus (WNV); indirect ELISA (iELISAs) for bovine herpesvirus type 1 (BHV-1), parainfluenza virus type 3 (PI-3), and bovine respiratory syncytial virus (BRSV); and virus neutralization (VN) for bovine viral diarrhea virus (BVDV) types I and II. Assay thresholds were evidence-based values employed by each laboratory. Comparable performance to serum was defined as FP sensitivity and specificity ≥ 80%. Filter-paper specificity estimates ranged from 92% in the cELISAs for N. caninum and WNV to 98% in the iELISAs for PI-3 and BRSV. Sensitivity was >85% for five tests (most ≥ 95%) but was insufficient (71-82%) for the PI-3 and BRSV iELISAs. Lowering the threshold for FP samples in these two ELISAs raised sensitivity to ≥ 87% and reduced specificity slightly (≥ 90% in three of the four test runs). Sample size limited the precision of some performance estimates. Based on the criteria of sensitivity and specificity ≥ 80%, and using adjusted FP thresholds for PI-3 and BRSV, FP sensitivity and specificity were comparable to serum in all seven assays. A potential limitation of FP is reduced sensitivity in tests that require undiluted serum (i.e., N. caninum cELISA and BVDV VNs). Possible toxicity to the assay cell layer in VN requires investigation. Results suggested that cELISA is superior to iELISA for detecting antibodies in FP samples from reindeer and other Rangifer tarandus subspecies. Our findings expand the potential utility of FP sampling from wildlife.

Correlation Between Iron and alpha and pi Glutathione-S-Transferase Levels in Humans

DTIC Science & Technology

2012-09-01

assays were performed as described in the Biotrin High Sensitivity Alpha GST EIA kit protocol. First, serum samples were diluted 1:10 with wash solution...immunosorbent assays were performed as described in the Biotrin Pi GST EIA kit protocol. First, plasma samples were diluted 1:5 with sample diluent...immunosorbent assays were performed as described in the AssayMax Human Transferrin ELISA kit protocol. First, serum samples were diluted 1:2000 with MIX

Amoeba-Resisting Bacteria and Ventilator-Associated Pneumonia

PubMed Central

La Scola, Bernard; Boyadjiev, Ioanna; Greub, Gilbert; Khamis, Atieh; Martin, Claude

2003-01-01

To evaluate the role of amoeba-associated bacteria as agents of ventilator-associated pneumonia (VAP), we tested the water from an intensive care unit (ICU) every week for 6 months for such bacteria isolates; serum samples and bronchoalveolar lavage samples (BAL) were also obtained from 30 ICU patients. BAL samples were examined for amoeba-associated bacteria DNA by suicide-polymerase chain reaction, and serum samples were tested against ICU amoeba-associated bacteria. A total of 310 amoeba-associated bacteria from10 species were isolated. Twelve of 30 serum samples seroconverted to one amoeba-associated bacterium isolated in the ICU, mainly Legionella anisa and Bosea massiliensis, the most common isolates from water (p=0.021). Amoeba-associated bacteria DNA was detected in BAL samples from two patients whose samples later seroconverted. Seroconversion was significantly associated with VAP and systemic inflammatory response syndrome, especially in patients for whom no etiologic agent was found by usual microbiologic investigations. Amoeba-associated bacteria might be a cause of VAP in ICUs, especially when microbiologic investigations are negative. PMID:12890321

Lack of Serologic Evidence of Neospora caninum in Humans, England

PubMed Central

McCann, Catherine M.; Vyse, Andrew J.; Salmon, Roland L; Thomas, Daniel; Williams, Diana J.L.; McGarry, John W.; Pebody, Richard

2008-01-01

Retrospective testing of 3,232 serum samples from the general population and 518 serum samples from a high-risk group showed no evidence of human exposure to Neospora caninum in England. Results were obtained by using immunofluorescence antibody testing and ELISA to analyze frequency distribution. PMID:18507920

On-line concentration and determination of all-trans- and 13-cis- retinoic acids in rabbit serum by application of sweeping technique in micellar electrokinetic chromatography.

PubMed

Zhao, Yongxi; Kong, Yu; Wang, Bo; Wu, Yayan; Wu, Hong

2007-03-30

A simple and rapid micellar electrokinetic chromatography (MEKC) method with UV detection was developed for the simultaneous separation and determination of all-trans- and 13-cis-retinoic acids in rabbit serum by on-line sweeping concentration technique. The serum sample was simply deproteinized and centrifuged. Various parameters affecting sample enrichment and separation were systematically investigated. Under optimal conditions, the analytes could be well separated within 17min, and the relative standard deviations (RSD) of migration times and peak areas were less than 3.4%. Compared with the conventional MEKC injection method, the 18- and 19-fold improvements in sensitivity were achieved, respectively. The proposed method has been successfully applied to the determination of all-trans- and 13-cis-retinoic acids in serum samples from rabbits and could be feasible for the further pharmacokinetics study of all-trans-retinoic acid.

Classification of oral cancers using Raman spectroscopy of serum

NASA Astrophysics Data System (ADS)

Sahu, Aditi; Talathi, Sneha; Sawant, Sharada; Krishna, C. Murali

2014-03-01

Oral cancers are the sixth most common malignancy worldwide, with low 5-year disease free survival rates, attributable to late detection due to lack of reliable screening modalities. Our in vivo Raman spectroscopy studies have demonstrated classification of normal and tumor as well as cancer field effects (CFE), the earliest events in oral cancers. In view of limitations such as requirement of on-site instrumentation and stringent experimental conditions of this approach, feasibility of classification of normal and cancer using serum was explored using 532 nm excitation. In this study, strong resonance features of β-carotenes, present differentially in normal and pathological conditions, were observed. In the present study, Raman spectra of sera of 36 buccal mucosa, 33 tongue cancers and 17 healthy subjects were recorded using Raman microprobe coupled with 40X objective using 785 nm excitation, a known source of excitation for biomedical applications. To eliminate heterogeneity, average of 3 spectra recorded from each sample was subjected to PC-LDA followed by leave-one-out-cross-validation. Findings indicate average classification efficiency of ~70% for normal and cancer. Buccal mucosa and tongue cancer serum could also be classified with an efficiency of ~68%. Of the two cancers, buccal mucosa cancer and normal could be classified with a higher efficiency. Findings of the study are quite comparable to that of our earlier study, which suggest that there exist significant differences, other than β- carotenes, between normal and cancerous samples which can be exploited for the classification. Prospectively, extensive validation studies will be undertaken to confirm the findings.

Accuracy of cotinine serum test to detect the smoking habit and its association with periodontal disease in a multicenter study

PubMed Central

Duque, Andrés; Martínez, Paula-Juliana; Giraldo, Astrid; Gualtero, Diego F.; Ardila, Carlos-Martín; Contreras, Adolfo; Duarte, Silvia

2017-01-01

Background The validity of the surveys on self-reported smoking status is often questioned because smokers underestimate cigarette use and deny the habit. It has been suggested that self-report should be accompanied by cotinine test. This report evaluates the usefulness of serum cotinine test to assess the association between smoking and periodontal status in a study with a large sample population to be used in studies with other serum markers in epidemiologic and periodontal medicine researches. Material and Methods 578 patients who were part of a multicenter study on blood biomarkers were evaluated about smoking and its relation to periodontal disease. Severity of periodontal disease was determinate using clinical attachment loss (CAL). Smoking was assessed by a questionnaire and a blood sample drawn for serum cotinine determination. Results The optimal cut-off point for serum cotinine was 10 ng/ml. Serum cotinine showed greater association with severity of CAL than self-report for mild-moderate CAL [OR 2.03 (CI95% 1.16-3.53) vs. OR 1.08 (CI95% 0.62-1.87) ] advanced periodontitis [OR 2.36 (CI95% 1.30- 4.31) vs. OR 2.06 (CI95% 0.97-4.38) ] and extension of CAL > 3 mm [ OR 1.78 (CI95% 1.16-1.71) vs. 1.37 (CI95% 0.89-2.11)]. When the two tests were evaluated together were not shown to be better than serum cotinine test. Conclusions Self-reported smoking and serum cotinine test ≥ 10ng/ml are accurate, complementary and more reliable methods to assess the patient’s smoking status and could be used in studies evaluating serum samples in large population and multicenter studies. Clinical Relevance: The serum cotinine level is more reliable to make associations with the patient’s periodontal status than self-report questionnaire and could be used in multicenter and periodontal medicine studies. Key words:Biological markers, serum, cotinine, periodontitis, smoking. PMID:28578367

Serum testosterone levels in males are not associated with entrepreneurial behavior in two independent observational studies.

PubMed

van der Loos, Matthijs J H M; Haring, Robin; Rietveld, Cornelius A; Baumeister, Sebastian E; Groenen, Patrick J F; Hofman, Albert; de Jong, Frank H; Koellinger, Philipp D; Kohlmann, Thomas; Nauck, Matthias A; Rivadeneira, Fernando; Uitterlinden, André G; van Rooij, Frank J A; Wallaschofski, Henri; Thurik, A Roy

2013-07-02

Previous research has suggested a positive association between testosterone (T) and entrepreneurial behavior in males. However, this evidence was found in a study with a small sample size and has not been replicated. In the present study, we aimed to verify this association using two large, independent, population-based samples of males. We tested the association of T with entrepreneurial behavior, operationalized as self-employment, using data from the Rotterdam Study (N=587) and the Study of Health in Pomerania (N=1697). Total testosterone (TT) and sex hormone-binding globulin (SHBG) were measured in the serum. Free testosterone (FT), non-SHBG-bound T (non-SHBG-T), and the TT/SHBG ratio were calculated and used as measures of bioactive serum T, in addition to TT adjusted for SHBG. Using logistic regression models, we found no significant associations between any of the serum T measures and self-employment in either of the samples. To our knowledge, this is the first large-scale study on the relationship between serum T and entrepreneurial behavior. Copyright © 2013 Elsevier Inc. All rights reserved.

Restricted access carbon nanotubes for direct extraction of cadmium from human serum samples followed by atomic absorption spectrometry analysis.

PubMed

Barbosa, Adriano F; Barbosa, Valéria M P; Bettini, Jefferson; Luccas, Pedro O; Figueiredo, Eduardo C

2015-01-01

In this paper, we propose a new sorbent that is able to extract metal ions directly from untreated biological fluids, simultaneously excluding all proteins from these samples. The sorbent was obtained through the modification of carbon nanotubes (CNTs) with an external bovine serum albumin (BSA) layer, resulting in restricted access carbon nanotubes (RACNTs). The BSA layer was fixed through the interconnection between the amine groups of the BSA using glutaraldehyde as cross-linker. When a protein sample is percolated through a cartridge containing RACNTs and the sample pH is higher than the isoelectric point of the proteins, both proteins from the sample and the BSA layer are negatively ionized. Thus, an electrostatic repulsion prevents the interaction between the proteins from the sample on the RACNTs surface. At the same time, metal ions are adsorbed in the CNTs (core) after their passage through the chains of proteins. The Cd(2+) ion was selected for a proof-of-principle case to test the suitability of the RACNTs due to its toxicological relevance. RACNTs were able to extract Cd(2+) and exclude almost 100% of the proteins from the human serum samples in an online solid-phase extraction system coupled with thermospray flame furnace atomic absorption spectrometry. The limits of detection and quantification were 0.24 and 0.80 μg L(-1), respectively. The sampling frequency was 8.6h(-1), and the intra- and inter-day precisions at the 0.80, 15.0, and 30.0 μg L(-1) Cd(2+) levels were all lower than 10.1% (RSD). The recoveries obtained for human blood serum samples fortified with Cd(2+) ranged from 85.0% to 112.0%. The method was successfully applied to analyze Cd(2+) directly from six human blood serum samples without any pretreatment, and the observed concentrations ranged from

Multiplexed Analysis of Serum Breast and Ovarian Cancer Markers by Means of Suspension Bead-quantum Dot Microarrays

NASA Astrophysics Data System (ADS)

Brazhnik, Kristina; Sokolova, Zinaida; Baryshnikova, Maria; Bilan, Regina; Nabiev, Igor; Sukhanova, Alyona

Multiplexed analysis of cancer markers is crucial for early tumor diagnosis and screening. We have designed lab-on-a-bead microarray for quantitative detection of three breast cancer markers in human serum. Quantum dots were used as bead-bound fluorescent tags for identifying each marker by means of flow cytometry. Antigen-specific beads reliably detected CA 15-3, CEA, and CA 125 in serum samples, providing clear discrimination between the samples with respect to the antigen levels. The novel microarray is advantageous over the routine single-analyte ones due to the simultaneous detection of various markers. Therefore the developed microarray is a promising tool for serum tumor marker profiling.

Headspace Solid Phase Micro Extraction Gas Chromatographic Determination of Fenthion in Human Serum

PubMed Central

Kasiotis, Konstantinos M.; Souki, Helen; Tsakirakis, Angelos N.; Carageorgiou, Haris; Theotokatos, Spiridon A.; Haroutounian, Serkos A.; Machera, Kyriaki

2008-01-01

A simple and effective analytical procedure was developed for the determination of fenthion residues in human serum samples. The sample treatment was performed using the headspace solid-phase micro extraction with polyacrylate fiber, which has the advantage to require low amount of serum (1 mL) without tedious pre-treatment. The quantification of fenthion was carried out by gas chromatography-mass spectrometry and the recoveries ranged from 79 to 104% at two spiking levels for 6 replicates. Detection and quantification limits were calculated as 1.51 and 4.54 ng/mL of serum respectively. Two fenthion metabolites fenoxon and fenthion–sulfoxide were also identified. PMID:19325792

Analysis of serum and cerebrospinal fluid in clinically normal adult miniature donkeys.

PubMed

Mozaffari, A A; Samadieh, H

2013-09-01

To establish reference intervals for serum and cerebrospinal fluid (CSF) parameters in clinically healthy adult miniature donkeys. Experiments were conducted on 10 female and 10 male clinically normal adult miniature donkeys, randomly selected from five herds. Lumbosacral CSF collection was performed with the sedated donkey in the standing position. Cell analysis was performed immediately after the samples were collected. Blood samples were obtained from the jugular vein immediately after CSF sample collection. Sodium, potassium, glucose, urea nitrogen, total protein, calcium, chloride, phosphorous and magnesium concentrations were measured in CSF and serum samples. A paired t-test was used to compare mean values between female and male donkeys. The CSF was uniformly clear, colourless and free from flocculent material, with a specific gravity of 1.002. The range of total nucleated cell counts was 2-4 cells/μL. The differential white cell count comprised only small lymphocytes. No erythrocytes or polymorphonuclear cells were observed on cytological examination. Reference values were obtained for biochemical analysis of serum and CSF. Gender had no effect on any variables measured in serum or CSF (p>0.05). CSF analysis can provide important information in addition to that gained by clinical examination. CSF analysis has not previously been performed in miniature donkeys; this is the first report on the subject. In the present study, reference intervals for total nucleated cell count, total protein, glucose, urea nitrogen, sodium, potassium, chloride, calcium, phosphorous and magnesium concentrations of serum and CSF were determined for male and female miniature donkeys.

Analysis of human serum proteins by liquid phase isoelectric focusing and matrix-assisted laser desorption/ionization-mass spectrometry.

PubMed

Wang, Michael Z; Howard, Brandon; Campa, Michael J; Patz, Edward F; Fitzgerald, Michael C

2003-09-01

Direct matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis of human serum yielded ion signals from only a fraction of the total number of peptides and proteins expected to be in the sample. We increased the number of peptide and protein ion signals observed in the MALDI-TOF mass spectra analysis of human serum by using a prefractionation protocol based on liquid phase isoelectric focusing electrophoresis. This pre-fractionation technique facilitated the MALDI-TOF MS detection of as many as 262 different peptide and protein ion signals from human serum. The results obtained from three replicate fractionation experiments on the same serum sample indicated that 148 different peptide and protein ion signals were reproducibly detected using our isoelectric focusing and MALDI-TOF MS protocol.

Comparison of different mass spectrometric approaches coupled to gas chromatography for the analysis of organochlorine pesticides in serum samples.

PubMed

Fang, Jing; Wu, Qian; Zhao, Yun; Zhao, Hongzhi; Xu, Shunqing; Cai, Zongwei

2017-01-01

Gas chromatography-triple quadrupole mass spectrometry (GC-QqQMS) was applied for the determination of eight organochlorine pesticides (OCPs) in human serum. OCPs were extracted from the serum sample by solid phase extraction (SPE) and analyzed by gas chromatography mass spectrometry (GC-MS) or gas chromatography tandem mass spectrometry (GC-MS/MS). Electron ionization (EI) and negative chemical ionization (NCI) under two data acquisition modes, namely selected ion monitoring (SIM) and multiple reaction monitoring (MRM), were compared. The use of MRM generally provided higher selectivity and sensitivity because less interference from the sample matrix existed. The EI mode is more suitable for less electronegative compounds such as dichlorodiphenyldichloroethanes (DDDs) with detection limits ranging from 0.0060 to 0.060ng/mL. In the NCI mode, MRM analysis provided good and lower detection limits (0.0011-0.0030ng/mL) for pesticides containing more chlorines. The methods were validated by analyzing the pesticides in spiked serum at different levels with recoveries ranged from 83% to 116% and relative standard deviations of less than 10%. The developed method was applied for the determination of the OCPs in real human serum samples. Copyright © 2016 Elsevier B.V. All rights reserved.

Analysis of Serum Total and Free PSA Using Immunoaffinity Depletion Coupled to SRM: Correlation with Clinical Immunoassay Tests

PubMed Central

Liu, Tao; Hossain, Mahmud; Schepmoes, Athena A.; Fillmore, Thomas L.; Sokoll, Lori J.; Kronewitter, Scott R.; Izmirlian, Grant; Shi, Tujin; Qian, Wei-Jun; Leach, Robin J.; Thompson, Ian M.; Chan, Daniel W.; Smith, Richard D.; Kagan, Jacob; Srivastava, Sudhir; Rodland, Karin D.; Camp, David G.

2012-01-01

Recently, selected reaction monitoring mass spectrometry (SRM-MS) has been more frequently applied to measure low abundance biomarker candidates in tissues and biofluids, owing to its high sensitivity and specificity, simplicity of assay configuration, and exceptional multiplexing capability. In this study, we report for the first time the development of immunoaffinity depletion-based workflows and SRM-MS assays that enable sensitive and accurate quantification of total and free prostate-specific antigen (PSA) in serum without the requirement for specific PSA antibodies. Low ng/mL level detection of both total and free PSA was consistently achieved in both PSA-spiked female serum samples and actual patient serum samples. Moreover, comparison of the results obtained when SRM PSA assays and conventional immunoassays were applied to the same samples showed good correlation in several independent clinical serum sample sets. These results demonstrate that the workflows and SRM assays developed here provide an attractive alternative for reliably measuring candidate biomarkers in human blood, without the need to develop affinity reagents. Furthermore, the simultaneous measurement of multiple biomarkers, including the free and bound forms of PSA, can be performed in a single multiplexed analysis using high-resolution liquid chromatographic separation coupled with SRM-MS. PMID:22846433

Blood Phosphorus and Magnesium Levels in 130 Elite Track and Field Athletes

PubMed Central

Malliaropoulos, Nikolaos; Tsitas, Kostas; Porfiriadou, Anthoula; Papalada, Agapi; R.Ames, Paul; Del Buono, Angelo; Lippi, Giuseppe; Maffulli, Nicola

2012-01-01

Purpose This study tested the clinical utility and relevance of serum phosphorus and magnesium as markers possibly useful to monitor training in athletes. Methods Phosphorus and magnesium serum concentrations of 130 elite track and field athletes (65 males and 65 females, age range 20-30 years) from the National Athletics Sports Medicine Center database in Thessaloniki, Greece were measured. Results Abnormal results were found in 61 (47%) athletes (32 men and 29 women). In male athletes, serum phosphate was higher than normal in 18% and decreased in 1.5%, whereas serum magnesium concentration was higher in 26%, and lower in 3%. Regarding female athletes, higher serum phosphate and magnesium levels were detected in 26% and 17% respectively, whereas decreased serum magnesium was found in 3%. The most common alterations were higher serum phosphate (29/61, 47%) and magnesium concentrations (28/61, 46%). Abnormalities of serum phosphorus and magnesium concentrations were detected in almost half of the athletes. Hyperphosphataemia and hypermagnesaemia were the most common abnormalities. Conclusion The reference intervals used for general population cannot be used for athletes. Given the lack of pathological manifestations, the physiopathological significance of these findings is uncertain. Further studies on the interpretation of reported ion concentrations in athletes should take in account the type of sport practiced and also the possible variations during the training and competition season. PMID:23785576

Determinants of serum zinc in a random population sample of four Belgian towns with different degrees of environmental exposure to cadmium

PubMed Central

Thijs, Lutgarde; Staessen, Jan; Amery, Antoon; Bruaux, Pierre; Buchet, Jean-Pierre; Claeys, FranÇoise; De Plaen, Pierre; Ducoffre, Geneviève; Lauwerys, Robert; Lijnen, Paul; Nick, Laurence; Remy, Annie Saint; Roels, Harry; Rondia, Désiré; Sartor, Francis

1992-01-01

This report investigated the distribution of serum zinc and the factors determining serum zinc concentration in a large random population sample. The 1977 participants (959 men and 1018 women), 20–80 years old, constituted a stratified random sample of the population of four Belgian districts, representing two areas with low and two with high environmental exposure to cadmium. For each exposure level, a rural and an urban area were selected. The serum concentration of zinc, frequently used as an index for zinc status in human subjects, was higher in men (13.1 μmole/L, range 6.5–23.0 μmole/L) than in women (12.6 μmole/L, range 6.3–23.2 μmole/L). In men, 20% of the variance of serum zinc was explained by age (linear and squared term, R = 0.29), diurnal variation (r = 0.29), and total cholesterol (r = 0.16). After adjustment for these covariates, a negative relationship was observed between serum zinc and both blood (r = −0.10) and urinary cadmium (r = −0.14). In women, 11% of the variance could be explained by age (linear and squared term, R = 0.15), diurnal variation in serum zinc (r = 0.27), creatinine clearance (r = −0.11), log γ-glutamyltranspeptidase (r = 0.08), cholesterol (r = 0.07), contraceptive pill intake (r = −0.07), and log serum ferritin (r = 0.06). Before and after adjustment for significant covariates, serum zinc was, on average, lowest in the two districts where the body burden of cadmium, as assessed by urinary cadmium excretion, was highest. These results were not altered when subjects exposed to heavy metals at work were excluded from analysis. PMID:1486857

[Study on the neutralization capacity of different types of human measles virus vaccine and the epidemic strains].

PubMed

Feng, Yan; Lu, Yi-yu; Yan, Ju-ying; Jiang, Xiao-hui; Shi, Wen; Xu, Chang-ping; Li, Zhen

2007-11-01

To explore the neutralization capacities of different types of human serum to measles virus epidemic strains and vaccine strain. Neutralization antibody (NT) to Shanghai 191 and measles virus isolates in 2005 were tested using acute and convalescent serum samples from diagnosed measles patients, children serum samples collected before and after vaccination and serum samples of migrant residents, from 3 different regions. Additionally, animal immune serum referring to vaccine strain and 3 epidemic strains were prepared and used to undergo crossing neutralization test with corresponding strains mentioned-above. Antigenic ratios were calculated. GMT value of NT of after-immune serum to vaccine strains was 50.82,1.86 times higher than that to MVi/ZJ/05/7 (GMT was 27.35), whereas GMT value of convalescent serum to MVi/ZJ/05/7 (GMT was 386.95) was obviously higher than that to vaccine strain (GMT was 1:151.83),and GMT value of migrant residents' serum in 3 regions to MVi/ZJ/05/7 were 2.22-4.17 times lower than that to vaccine strain. Meanwhile,the antigenic ratios between MVi/ZJ/ 99/1, MVi/ZJ/04/1, MVi/ZJ/05/7 and vaccine strain were found to be 4.28,5.24 and 5.66 respectively. Additionally,low NT titers to vaccine strain were found in patients' acute sera and GMT value was over 1:4. There were obvious differences on neutralization antibody of different types of serum to measles vaccine strain and epidemic strains which indicating the antigenic diversity of epidemic strains had influenced the protective effectiveness of vaccine antibody to epidemic strains. It was of significance to carry on research projects on the antigenic diversity and effectiveness of measles vaccine.

Multiple diagnostic tests to identify cattle with Bovine viral diarrhea virus and duration of positive test results in persistently infected cattle

PubMed Central

Fulton, Robert W.; Hessman, Bill E.; Ridpath, Julia F.; Johnson, Bill J.; Burge, Lurinda J.; Kapil, Sanjay; Braziel, Barbara; Kautz, Kira; Reck, Amy

2009-01-01

Several tests for Bovine viral diarrhea virus (BVDV) were applied to samples collected monthly from December 20, 2005, through November 27, 2006 (day 0 to day 342) from 12 persistently infected (PI) cattle with BVDV subtypes found in US cattle: BVDV-1a, BVDV-1b, and BVDV-2a. The samples included clotted blood for serum, nasal swabs, and fresh and formalin-fixed ear notches. The tests were as follows: titration of infectious virus in serum and nasal swabs; antigen-capture (AC) enzyme-linked immunosorbent assay (ELISA), or ACE, on serum, nasal swabs, and fresh ear notches; gel-based polymerase chain reaction (PCR) testing of serum, nasal swabs, and fresh ear notches; immunohistochemical (IHC) testing of formalin-fixed ear notches; and serologic testing for BVDV antibodies in serum. Of the 12 animals starting the study, 3 died with mucosal disease. The ACE and IHC tests on ear notches had positive results throughout the study, as did the ACE and PCR tests on serum. There was detectable virus in nasal swabs from all the cattle throughout the study except for a few samples that were toxic to cell cultures. The serum had a virus titer ≥ log10 1.60 in all samples from all the cattle except for 3 collections from 1 animal. Although there were several equivocal results, the PCR test most often had positive results. The BVDV antibodies were due to vaccination or exposure to heterologous strains and did not appear to interfere with any BVDV test. These findings illustrate that PI cattle may be identified by several tests, but differentiation of PI cattle from cattle with acute BVDV infection requires additional testing, especially of blood samples and nasal swabs positive on initial testing. Also, calves PI with BVDV are continual shedders of infectious virus, as shown by the infectivity of nasal swabs over the 11-mo study. PMID:19436580

Partitioning behavior of heavy metals and persistent organic pollutants among feto-maternal bloods and tissues.

PubMed

Kim, Jun-Tae; Son, Min-Hui; Lee, Duk-Hee; Seong, Won Joon; Han, Seunghee; Chang, Yoon-Seok

2015-06-16

Heavy metals and persistent organic pollutants (POPs), including Pb, Cd, T-Hg, MeHg, PCDD/Fs, PCBs, PBDEs, PCNs, and PBDD/Fs, were analyzed in 20 paired samples of cord blood, maternal blood, maternal urine, and placenta. The samples were collected from pregnant mothers and neonates from South Korea in 2010. The distribution of heavy metals among the samples varied with their physicochemical characteristics. The concentrations of Pb and Hg in the maternal and the cord blood samples were significantly correlated each other, implying efficient transplacental transport (TPT). Cd and Hg were accumulated in the placenta, forming protein conjugates, and T-Hg was higher in the cord blood samples than the maternal blood samples due to the binding affinity of Hg with fetal proteins. POPs generally showed the highest concentrations in the maternal serum samples, and the POPs levels in the cord serum and the placenta samples were dependent on the degree of halogenation. The TPT of POPs was seemingly related to lipoprotein transportation. Some PBDE congeners, however, showed their highest concentrations in the cord serum samples, suggesting an additional TPT mechanism. This is the first study to detect PCNs and PBDD/Fs in the cord serum samples, showing that the PCN levels were comparable to other POPs. According to the principal component analysis (PCA) results of the contaminant levels, POPs and heavy metals showed significantly different characteristics, whereas PBDEs had an intermediate attribute. Despite the limited number of participants, the comprehensive analysis of trace contaminants in the paired sample sets enabled us to infer the distribution and TPT mechanism of various contaminants.

Assessment of serum selenium levels in 2-month-old sucking calves using total reflection X-ray fluorescence technique

NASA Astrophysics Data System (ADS)

Buoso, M. C.; Ceccato, D.; Moschini, G.; Bernardini, D.; Testoni, S.; Torboli, A.; Valdes, M.

2001-11-01

The assessment of selenium status of livestock plays an important role in the production of medicine since low serum Se levels influence disease resistance in ruminants. It has been proved that Se deficiency may cause muscular dystrophy, cardiomyopathy and even death. Serum level has been widely used to evaluate the Se short-term status in animals since there is a good association between serum Se level and the dietary intake of the element over a wide range. The purpose of this work was to determine the Se serum concentration in a population of 78 sucking 2-month-old calves, in order to corroborate a clinical diagnosis of severe deficiency status. The samples were analyzed by total reflection X-ray fluorescence (TXRF) at the ITAL STRUCTURES Research Laboratory. The results obtained from the serum samples presented Se concentrations varying from 10 to 66 ng/ml. The comparison between the obtained values and the expected serum selenium values (60-80 ng/ml), confirmed a mild to severe deficiency status in the investigated population.

Quantitative Evaluation of Serum Proteins Uncovers a Protein Signature Related to Maturity-Onset Diabetes of the Young (MODY).

PubMed

Tuerxunyiming, Muhadasi; Xian, Feng; Zi, Jin; Yimamu, Yilihamujiang; Abuduwayite, Reshalaiti; Ren, Yan; Li, Qidan; Abudula, Abulizi; Liu, SiQi; Mohemaiti, Patamu

2018-01-05

Maturity-onset diabetes of the young (MODY) is an inherited monogenic type of diabetes. Genetic mutations in MODY often cause nonsynonymous changes that directly lead to the functional distortion of proteins and the pathological consequences. Herein, we proposed that the inherited mutations found in a MODY family could cause a disturbance of protein abundance, specifically in serum. The serum samples were collected from a Uyghur MODY family through three generations, and the serum proteins after depletion treatment were examined by quantitative proteomics to characterize the MODY-related serum proteins followed by verification using target quantification of proteomics. A total of 32 serum proteins were preliminarily identified as the MODY-related. Further verification test toward the individual samples demonstrated the 12 candidates with the significantly different abundance in the MODY patients. A comparison of the 12 proteins among the sera of type 1 diabetes, type 2 diabetes, MODY, and healthy subjects was conducted and revealed a protein signature related with MODY composed of the serum proteins such as SERPINA7, APOC4, LPA, C6, and F5.

Osteosarcoma tissues and cell lines from patients with differing serum alkaline phosphatase concentrations display minimal differences in gene expression patterns

PubMed Central

de Sá Rodrigues, L. C.; Holmes, K. E.; Thompson, V.; Piskun, C. M.; Lana, S. E.; Newton, M. A.; Stein, T. J.

2016-01-01

Serum alkaline phosphatase (ALP) concentration is a prognostic factor for osteosarcoma in multiple studies, although its biological significance remains incompletely understood. To determine whether gene expression patterns differed in osteosarcoma from patients with differing serum ALP concentrations, microarray analysis was performed on 18 primary osteosarcoma samples and six osteosarcoma cell lines from dogs with normal and increased serum ALP concentration. No differences in gene expression patterns were noted between tumours or cell lines with differing serum ALP concentration using a gene-specific two-sample t-test. Using a more sensitive empirical Bayes procedure, defective in cullin neddylation 1 domain containing 1 (DCUN1D1) was increased in both the tissue and cell lines of the normal ALP group. Using quantitative PCR (qPCR), differences in DCUN1D1 expression between the two groups failed to reach significance. The homogeneity of gene expression patterns of osteosarcoma associated differing serum ALP concentrations are consistent with previous studies suggesting serum ALP concentration is not associated with intrinsic differences of osteosarcoma cells. PMID:25643733

Direct detection and characterization of foot-and-mouth disease virus in East Africa using a field-ready real-time PCR platform.

PubMed

Howson, E L A; Armson, B; Lyons, N A; Chepkwony, E; Kasanga, C J; Kandusi, S; Ndusilo, N; Yamazaki, W; Gizaw, D; Cleaveland, S; Lembo, T; Rauh, R; Nelson, W M; Wood, B A; Mioulet, V; King, D P; Fowler, V L

2018-02-01

Effective control and monitoring of foot-and-mouth disease (FMD) relies upon rapid and accurate disease confirmation. Currently, clinical samples are usually tested in reference laboratories using standardized assays recommended by The World Organisation for Animal Health (OIE). However, the requirements for prompt and serotype-specific diagnosis during FMD outbreaks, and the need to establish robust laboratory testing capacity in FMD-endemic countries have motivated the development of simple diagnostic platforms to support local decision-making. Using a portable thermocycler, the T-COR™ 8, this study describes the laboratory and field evaluation of a commercially available, lyophilized pan-serotype-specific real-time RT-PCR (rRT-PCR) assay and a newly available FMD virus (FMDV) typing assay (East Africa-specific for serotypes: O, A, Southern African Territories [SAT] 1 and 2). Analytical sensitivity, diagnostic sensitivity and specificity of the pan-serotype-specific lyophilized assay were comparable to that of an OIE-recommended laboratory-based rRT-PCR (determined using a panel of 57 FMDV-positive samples and six non-FMDV vesicular disease samples for differential diagnosis). The FMDV-typing assay was able to correctly identify the serotype of 33/36 FMDV-positive samples (no cross-reactivity between serotypes was evident). Furthermore, the assays were able to accurately detect and type FMDV RNA in multiple sample types, including epithelial tissue suspensions, serum, oesophageal-pharyngeal (OP) fluid and oral swabs, both with and without the use of nucleic acid extraction. When deployed in laboratory and field settings in Tanzania, Kenya and Ethiopia, both assays reliably detected and serotyped FMDV RNA in samples (n = 144) collected from pre-clinical, clinical and clinically recovered cattle. These data support the use of field-ready rRT-PCR platforms in endemic settings for simple, highly sensitive and rapid detection and/or characterization of FMDV. © 2017 The Authors. Transboundary and Emerging Diseases Published by Blackwell Verlag GmbH.

Single chain variable fragment displaying M13 phage library functionalized magnetic microsphere-based protein equalizer for human serum protein analysis.

PubMed

Zhu, Guijie; Zhao, Peng; Deng, Nan; Tao, Dingyin; Sun, Liangliang; Liang, Zhen; Zhang, Lihua; Zhang, Yukui

2012-09-18

Single chain variable fragment (scFv) displaying the M13 phage library was covalently immobilized on magnetic microspheres and used as a protein equalizer for the treatment of human serum. First, scFv displaying M13 phage library functionalized magnetic microspheres (scFv@M13@MM) was incubated with a human serum sample. Second, captured proteins on scFv@M13@MM were eluted with 2 M NaCl, 50 mM glycine-hydrochloric acid (Gly-HCl), and 20% (v/v) acetonitrile with 0.5% (v/v) trifluoroacetic acid in sequence. Finally, the tightly bonded proteins were released by the treatment with thrombin. The eluates were first analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with silver staining. Results indicated that the difference of protein concentration was reduced obviously in NaCl and Gly-HCl fractions compared with untreated human serum sample. The eluates were also digested with trypsin, followed by online 2D-strong cation exchange (SCX)-RPLC-ESI-MS/MS analysis. Results demonstrated that the number of proteins identified from an scFv@M13@MM treated human serum sample was improved 100% compared with that from the untreated sample. In addition, the spectral count of 10 high abundance proteins (serum albumin, serotransferrin, α-2-macroglobulin, α-1-antitrypsin, apolipoprotein B-100, Ig γ-2 chain C region, haptoglobin, hemopexin, α-1-acid glycoprotein 1, and α-2-HS-glycoprotein) decreased evidently after scFv@M13@MM treatment. All these results demonstrate that scFv@M13@MM could efficiently remove high-abundance proteins, reduce the protein concentration difference of human serum, and result in more protein identification.

Comparison of human whole blood, plasma, and serum matrices for the determination of perfluorooctanesulfonate (PFOS), perfluorooctanoate (PFOA), and other fluorochemicals

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ehresman, David J.; Froehlich, John W.; Olsen, Geary W.

2007-02-15

Interest in human exposure to perfluorinated acids, including perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHS), perfluorooctanesulfonate (PFOS), and perfluorooctanoate (PFOA) has led to their measurement in whole blood, plasma and serum. Comparison of measurements in these different blood-based matrices, however, has not been rigorously investigated to allow for across-matrix comparisons. This research evaluated concentrations of PFBS, PFHS, PFOS, and PFOA in whole blood collected in heparin (lithium) and ethylenediamine tetraacetic acid (EDTA), plasma samples collected in heparin and EDTA, and serum (from whole blood allowed to clot). Blood samples were collected from 18 voluntary participants employed at 3M Company. Solid phase extraction methodsmore » were used for all analytical sample preparations, and analyses were completed using high-pressure liquid chromatography/tandem mass spectrometry methods. Serum concentrations ranged from: limit of quantitation (LOQ, 5 ng/mL) to 25 ng/mL for PFBS; LOQ (5 ng/mL) to 75 ng/mL for PFHS; LOQ (5 ng/mL) to 880 ng/mL for PFOS; and LOQ (5 or 10 ng/mL) to 7320 ng/mL for PFOA. Values less than the LOQ were not included in the statistical analyses of the mean of the ratios of individual values for the matrices. PFBS was not quantifiable in most samples. Serum to plasma ratios for PFHS, PFOS, and PFOA were 1:1 and this ratio was independent of the level of concentrations measured. Serum or plasma to whole blood ratios, regardless of the anticoagulant used, approximated 2:1. The difference between plasma and serum and whole blood corresponded to volume displacement by red blood cells, suggesting that the fluorochemicals are not found intracellularly or attached to the red blood cells.« less

Radioimmunoassays for the serum thymic factor (FTS)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Erickson, B.W.; Fok, K.F.; Incefy, G.S.

1987-01-06

This patent describes a radioimmunoassay of serum thymic factor (FTS) in a test sample, comprising: (a) contacting a first aliquot of the sample with anti-FTS antibody, with a known amount of FTS hormone standard and a known amount of radiolabeled FTS analogue; and (b) contacting a second aliquot of the sample with anti-FTS antibody and a known amount of radiolabeled FTS analogue; and (c) measuring the radioactivity of the antigen-antibody complex in each aliquot; and (d) calculating the amount of FTS in the test sample.

Field-Effect Biosensors for On-Site Detection: Recent Advances and Promising Targets.

PubMed

Choi, Jaebin; Seong, Tae Wha; Jeun, Minhong; Lee, Kwan Hyi

2017-10-01

There is an explosive interest in the immediate and cost-effective analysis of field-collected biological samples, as many advanced biodetection tools are highly sensitive, yet immobile. On-site biosensors are portable and convenient sensors that provide detection results at the point of care. They are designed to secure precision in highly ionic and heterogeneous solutions with minimal hardware. Among various methods that are capable of such analysis, field-effect biosensors are promising candidates due to their unique sensitivity, manufacturing scalability, and integrability with computational circuitry. Recent developments in nanotechnological surface modification show promising results in sensing from blood, serum, and urine. This report gives a particular emphasis on the on-site efficacy of recently published field-effect biosensors, specifically, detection limits in physiological solutions, response times, and scalability. The survey of the properties and existing detection methods of four promising biotargets, exosomes, bacteria, viruses, and metabolites, aims at providing a roadmap for future field-effect and other on-site biosensors. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Isolation of Coxiella burnetii from serum of patients with acute Q fever.

PubMed

Vincent, Gemma A; Graves, Stephen R; Robson, Jennifer M; Nguyen, Chelsea; Hussain-Yusuf, Hazizul; Islam, Aminul; Fenwick, Stanley G; Stenos, John

2015-12-01

Worldwide there are few isolate collections of the intracellular bacterium Coxiella burnetii, due to the difficulties associated with working with the organism and the scarcity of suitable samples from which to attempt isolation. Particularly lacking are isolates from acute Q fever patients. The aim of this study was to evaluate whether the serum samples taken from patients with confirmed acute Q fever during the early stage of their disease represented a potential source of viable C. burnetii. Isolation was attempted from 65 of these samples by inoculation of the serum into Vero cell culture and was successful in 36 cases (55%). This high success rate was likely due to extended incubation of up to twelve weeks of the inoculated cultures, allowing the growth of the organism to levels detectable by PCR. Retrospective analysis of the time the sera was stored prior to inoculation into culture demonstrated that C. burnetii remained viable for 224 days in samples stored refrigerated and 371 days in samples stored frozen at -20 °C. These results demonstrate that standard serum samples taken from acute Q fever patients are a valuable source of new isolates of C. burnetii, with no special handling of the specimens required to maintain the organism's viability. Copyright © 2015 Elsevier B.V. All rights reserved.

[Determination of ethylene glycol in biological fluids--propylene glycol interferences].

PubMed

Gomółka, Ewa; Cudzich-Czop, Sylwia; Sulka, Adrianna

2013-01-01

Many laboratories in Poland do not use gas chromatography (GC) method for determination of ethylene glycol (EG) and methanol in blood of poisoned patients, they use non specific spectrophotometry methods. One of the interfering substances is propylene glycol (PG)--compound present in many medical and cosmetic products: drops, air freshens, disinfectants, electronic cigarettes and others. In Laboratory of Analytical Toxicology and Drug Monitoring in Krakow determination of EG is made by GC method. The method enables to distinguish and make resolution of (EG) and (PG) in biological samples. In the years 2011-2012 in several serum samples from diagnosed patients PG was present in concentration from several to higher than 100 mg/dL. The aim of the study was to estimate PG interferences of serum EG determination by spectrophotometry method. Serum samples containing PG and EG were used in the study. The samples were analyzed by two methods: GC and spectrophotometry. Results of serum samples spiked with PG with no EG analysed by spectrophotometry method were improper ("false positive"). The results were correlated to PG concentration in samples. Calculated cross-reactivity of PG in the method was 42%. Positive results of EG measured by spectrophotometry method must be confirmed by reference GC method. Spectrophotometry method shouldn't be used for diagnostics and monitoring of patients poisoned by EG.

Detection of immunomagnetically captured 4',6-diamidino-2-phenyl-indole (DAPI)-labeled Escherichia coli 0157:H7 by fluorescent microscopic imaging

NASA Astrophysics Data System (ADS)

Tu, Shu-I.; Uknalis, Joseph; Patterson, Deidre; Gehring, Andrew G.

1999-01-01

Live cells of E. coliO157:H7 were captured by goat anti-E. coliO157 serum coated on the surface of polystyrene based immunomagnetic beads (IMB). The captured bacteria were labeled by 4',6-diamidino-2-phenylindole (DAPI), a nucleic acid stain, for observation by epifluorescent microscopy. The beads with captured bacteria were then concentrated by magnetic separators. The efficiency of this magnetic concentration step was less than that of using high speed centrifugation. The antibody-captured and IMB-immobilized bacteria were then applied on HF-treated, bovine serum albumin (BSA)-coated microscope slides mounted on an automated stage, and magnetically aligned before fluorescence distribution was measured by a cooled CCD attached to an inverted microscope. Since the beads were concentrated and linearly aligned along the edge of the magnetic field, image capture along the edge for a few field widths was sufficient to account for most of captured bacteria. We applied this approach to determine the bacterial counts in spiked beef hamburger patties. The results showed that after a 6-hour enrichment, sufficient number of the bacteria could be detected from the samples spiked with 1 CFU of E. coliO157:H7 per gram of the hamburger.

Spatial distribution of bisphenol S in surface water and human serum from Yangtze River watershed, China: Implications for exposure through drinking water.

PubMed

Wan, Yanjian; Xia, Wei; Yang, Shunyi; Pan, Xinyun; He, Zhenyu; Kannan, Kurunthachalam

2018-05-01

Bisphenol S (BPS) is an emerging environmental contaminant. The occurrence of this compound in humans and the environment is not well described. In this study, 120 surface water samples and 240 human serum samples were collected along the Yangtze River in 2015 for the determination of the occurrence of BPS. Surface water and human serum samples were extracted by solid phase extraction and liquid-liquid extraction, respectively, and analyzed by ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). BPS was detected in all river water samples at concentrations that ranged from 0.18 to 14.9 ng/L (median: 0.98 ng/L), with higher concentrations in spring than summer. The median estimated daily intake (EDI) of BPS through water ingestion by infants in spring and summer was 0.12 and 0.06 ng/kg body weight (bw)/day, respectively. BPS was detected in human serum with the highest concentrations in samples from Nanjing (median: 0.65 ng/mL, maximum: 169 ng/mL) among the four cities studied. No significant gender related difference in BPS concentrations was observed in human sera, while higher concentrations were found in younger individuals than elderly. The EDI of BPS calculated based on serum concentrations of adults in Nanjing was 22.8 ng/kg bw/day. Ingestion of water accounted for <1% of the total BPS intake by the Chinese population. This is the first report of the occurrence of BPS in water from the Yangtze River and human serum from several cities located along this river in China. Copyright © 2018 Elsevier Ltd. All rights reserved.

Chemometrics enhanced HPLC-DAD performance for rapid quantification of carbamazepine and phenobarbital in human serum samples.

PubMed

Vosough, Maryam; Ghafghazi, Shiva; Sabetkasaei, Masoumeh

2014-02-01

This paper describes development and validation of a simple and efficient bioanalytical procedure for simultaneous determination of phenobarbital and carbamazepine in human serum samples using high performance liquid chromatography with photodiode-array detection (HPLC-DAD) regarding a fast elution methodology in less than 5 min. Briefly, this method consisted of a simple deproteinization step of serum samples followed by HPLC analysis on a Bonus-RP column using an isocratic mode of elution with acetonitrile/K2HPO4 (pH=7.5) buffer solution (45:55). Due to the presence of serum endogenous components as non-calibrated components in the sample, second-order calibration based on multivariate curve resolution-alternating least squares (MCR-ALS), has been applied on a set of absorbance matrices collected as a function of retention time and wavelengths. Acceptable resolution and quantification results were achieved in the presence of matrix interferences and the second-order advantage was fully exploited. The average recoveries for carbamazepine and phenobarbital were 89.7% and 86.1% and relative standard deviation values were lower than 9%. Additionally, computed elliptical joint confidence region (EJCR) confirmed the accuracy of the proposed method and indicated the absence of both constant and proportional errors in the predicted concentrations. The developed method enabled the determination of the analytes in different serum samples in the presence of overlapped profiles, while keeping experimental time and extraction steps at minimum. Finally, the serum concentration levels of carbamazepine in three time intervals were reported for morphine-dependents who had received carbamazepine for treating their neuropathic pain. © 2013 Elsevier B.V. All rights reserved.

Metabolomic Quality Assessment of EDTA Plasma and Serum Samples.

PubMed

Malm, Linus; Tybring, Gunnel; Moritz, Thomas; Landin, Britta; Galli, Joakim

2016-10-01

Handling and processing of blood can significantly alter the molecular composition and consistency of biobank samples and can have a major impact on the identification of biomarkers. It is thus crucial to identify tools to determine the quality of samples to be used in biomarker discovery studies. In this study, a non-targeted gas chromatography/time-of-flight mass spectrometry (GC-TOFMS) metabolomic strategy was used with the aim of identifying quality markers for serum and plasma biobank collections lacking proper documentation of preanalytical handling. The effect of postcentrifugation delay was examined in serum stored in tubes with gel separation plugs and ethylenediaminetetraacetic acid (EDTA) plasma in tubes with or without gel separation plugs. The change in metabolic pattern was negligible in all sample types processed within 3 hours after centrifugation regardless of whether the samples were kept at 4°C or 22°C. After 8 and 24 hours postcentrifugation delay before aliquoting, there was a pronounced increase in the number of affected metabolites, as well as in the magnitude of the observed changes. No protective effect on the metabolites was observed in gel-separated EDTA plasma samples. In a separate series of experiments, lactate and glucose levels were determined in plasma to estimate the effect of precentrifugation delay. This separate experiment indicates that the lactate to glucose ratio may serve as a marker to identify samples with delayed time to centrifugation. Although our data from the untargeted GC-TOFMS analysis did not identify any specific markers, we conclude that plasma and serum metabolic profiles remain quite stable when plasma and serum are centrifuged and separated from the blood cells within 3 hours.

Identification of potential serum peptide biomarkers of biliary tract cancer using MALDI MS profiling

PubMed Central

2014-01-01

Background The aim of this discovery study was the identification of peptide serum biomarkers for detecting biliary tract cancer (BTC) using samples from healthy volunteers and benign cases of biliary disease as control groups. This work was based on the hypothesis that cancer-specific exopeptidases exist and that their activities in serum can generate cancer-predictive peptide fragments from circulating proteins during coagulation. Methods This case control study used a semi-automated platform incorporating polypeptide extraction linked to matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) to profile 92 patient serum samples. Predictive models were generated to test a validation serum set from BTC cases and healthy volunteers. Results Several peptide peaks were found that could significantly differentiate BTC patients from healthy controls and benign biliary disease. A predictive model resulted in a sensitivity of 100% and a specificity of 93.8% in detecting BTC in the validation set, whilst another model gave a sensitivity of 79.5% and a specificity of 83.9% in discriminating BTC from benign biliary disease samples in the training set. Discriminatory peaks were identified by tandem MS as fragments of abundant clotting proteins. Conclusions Serum MALDI MS peptide signatures can accurately discriminate patients with BTC from healthy volunteers. PMID:24495412

The hook effect in calcitonin immunoradiometric assay: a case report.

PubMed

Fangous, Marie-Sarah; Kerspern, Hélène; Moineau, Marie-Pierre; Kerlan, Véronique; Alavi, Zarrin; Carré, Jean-Luc

2012-12-01

The hook effect, which has long been detected and documented for immunoradiometric assays (IRMA) such as those measuring prolactin or thyroglobulin, occurs when the serum antigen level is extremely high, thus inducing a bias in the methodology of measurement. We report the case of an 80-year-old man with confirmed medullary thyroid carcinoma (MTC). In the case reported here, the clinical status of the patient contrasts with his tumor antigen, serum calcitonin (CT), concentrations. The measured increased CT concentrations revealed the presence of a hook effect. This phenomenon occurs due to an excess of antigen during the one-step IRMA where the signal antibodies, bound to the non-captured antigens, are washed out during the measurement, inducing the loss of signal. Aiming to prevent the "hook effect", successive dilutions of the same sample of serum were done. Previous studies have shown when one-step IRMA reveals high concentrations of a tumor serum antigen (i.e. prolactin or thyroglobulin), a two-step IRMA or a systematic 1:10 dilution of the serum sample prevents the formation of the "hook effect". In our case report, the CT "hook effect" formation was prevented by performing serial dilutions of the serum sample. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Protein microarray analysis reveals BAFF-binding autoantibodies in systemic lupus erythematosus

PubMed Central

Price, Jordan V.; Haddon, David J.; Kemmer, Dodge; Delepine, Guillaume; Mandelbaum, Gil; Jarrell, Justin A.; Gupta, Rohit; Balboni, Imelda; Chakravarty, Eliza F.; Sokolove, Jeremy; Shum, Anthony K.; Anderson, Mark S.; Cheng, Mickie H.; Robinson, William H.; Browne, Sarah K.; Holland, Steven M.; Baechler, Emily C.; Utz, Paul J.

2013-01-01

Autoantibodies against cytokines, chemokines, and growth factors inhibit normal immunity and are implicated in inflammatory autoimmune disease and diseases of immune deficiency. In an effort to evaluate serum from autoimmune and immunodeficient patients for Abs against cytokines, chemokines, and growth factors in a high-throughput and unbiased manner, we constructed a multiplex protein microarray for detection of serum factor–binding Abs and used the microarray to detect autoantibody targets in SLE. We designed a nitrocellulose-surface microarray containing human cytokines, chemokines, and other circulating proteins and demonstrated that the array permitted specific detection of serum factor–binding probes. We used the arrays to detect previously described autoantibodies against cytokines in samples from individuals with autoimmune polyendocrine syndrome type 1 and chronic mycobacterial infection. Serum profiling from individuals with SLE revealed that among several targets, elevated IgG autoantibody reactivity to B cell–activating factor (BAFF) was associated with SLE compared with control samples. BAFF reactivity correlated with the severity of disease-associated features, including IFN-α–driven SLE pathology. Our results showed that serum factor protein microarrays facilitate detection of autoantibody reactivity to serum factors in human samples and that BAFF-reactive autoantibodies may be associated with an elevated inflammatory disease state within the spectrum of SLE. PMID:24270423

Application of Ultrasound-Assisted Surfactant-Enhanced Emulsification Microextraction Based on Solidification of Floating Organic Droplets and High Performance Liquid Chromatography for Preconcentration and Determination of Alprazolam and Chlordiazepoxide in Human Serum Samples.

PubMed

Goudarzi, Nasser; Amirnavaee, Monavar; Arab Chamjangali, Mansour; Farsimadan, Sahar

2017-07-01

An improved microextraction method is proposed on the basis of ultrasound-assisted surfactant-enhanced emulsification and solidification of a floating organic droplet procedure combined with high performance liquid chromatography for the preconcentration and quantification of alprazolam (ALP) and chlordiazepoxide (CHL) present in a number of human serum samples. Several parameters affecting the extraction efficiency were investigated by the Plackett -Burman factorial design as the screening design. Then the response surface methodology based on the Box-Behnken design was used to optimize the effective parameters in the proposed procedure. The limits of detection for the proposed method were found to be 3.0 and 3.1 ng mL-1 for CHL and ALP, respectively. The calibration curves obtained for the method were linear in the ranges of 10.0-3,500.0 and 10.0-3,000.0 ng mL-1 for CHL and ALP, respectively, with a good determination coefficient. The recoveries of the drugs in the spiked human serum samples were above 93.0%. The developed method was successfully applied to the analysis of these studied drugs in human serum samples. The pre-treatment of the serum samples was performed using acetonitrile to remove the proteins. The proposed procedure was an accurate and reliable one for the determination and preconcentration of these drugs in blood samples. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Polychlorinated biphenyls and polybrominated biphenyl ethers in adipose tissue and matched serum from an E-waste recycling area (Wenling, China).

PubMed

Lv, Quan-Xia; Wang, Wenyue; Li, Xing-Hong; Yu, Lianlian; Zhang, Yun; Tian, Yuan

2015-04-01

To Date, the knowledge on relationship between PCBs/PBDEs exposure and thyroid hormones (THs) levels during pregnancy still needs to be extended. Meanwhile, studies on congener-specific adipose-serum ratios for PCBs/PBDEs were limited. This study reports the levels of PCBs/PBDEs in serum-adipose tissue samples (n = 64) from expectant women living surrounding e-waste recycling sites in Wenling, China. Their concentrations varied from several to hundreds of ng g(-1) lipid. Maternal exposure to PCBs was associated with lower TSH during pregnancy, suggesting possible implication for maternal health and fetal development. The compound levels between the adipose tissue and matched serum samples were highly correlated (p < 0.001), generating a predicted adipose-serum partitioning relationship for individual PCB congener and PBDE congener. Molecular characteristics, such as Kow value, molecular weight and molecular volume, may play a key role in the variable partitioning of some compounds between serum and adipose tissue. Copyright © 2015 Elsevier Ltd. All rights reserved.

Reference intervals for acute phase protein and serum protein electrophoresis values in captive Asian elephants (Elephas maximus).

PubMed

Isaza, Ramiro; Wiedner, Ellen; Hiser, Sarah; Cray, Carolyn

2014-09-01

Acute phase protein (APP) immunoassays and serum protein electrophoresis (SPEP) are assays for evaluating the inflammatory response and have use as diagnostic tools in a variety of species. Acute phase proteins are markers of inflammation that are highly conserved across different species while SPEP separates and quantifies serum protein fractions based on their physical properties. In the current study, serum samples from 35 clinically healthy Asian elephants (Elephas maximus) were analyzed using automated assays for C-reactive protein, serum amyloid A, and haptoglobin and SPEP. Robust methods were used to generate reference intervals for the APPs: C-reactive protein (1.3-12.8 mg/l), serum amyloid A (0-47.5 mg/l), and haptoglobin (0-1.10 mg/ml). In addition, SPEP was performed on these samples to establish reference intervals for each protein fraction. A combination of APPs and SPEP measurements are valuable adjunctive diagnostic tools in elephant health care. © 2014 The Author(s).

MicroRNA Expression Profiling of the Armed Forces Health Surveillance Branch Cohort for Identification of "Enviro-miRs" Associated With Deployment-Based Environmental Exposure.

PubMed

Dalgard, Clifton L; Polston, Keith F; Sukumar, Gauthaman; Mallon, Col Timothy M; Wilkerson, Matthew D; Pollard, Harvey B

2016-08-01

The aim of this study was to identify serum microRNA (miRNA) biomarkers that indicate deployment-associated exposures in service members at military installations with open burn pits. Another objective was to determine detection rates of miRNAs in Department of Defense Serum Repository (DoDSR) samples with a high-throughput methodology. Low-volume serum samples (n = 800) were profiled by miRNA-capture isolation, pre-amplification, and measurement by a quantitative PCR-based OpenArray platform. Normalized quantitative cycle values were used for differential expression analysis between groups. Assay specificity, dynamic range, reproducibility, and detection rates by OpenArray passed target desired specifications. Serum abundant miRNAs were consistently measured in study specimens. Four miRNAs were differentially expressed in the case deployment group subjects. miRNAs are suitable RNA species for biomarker discovery in the DoDSR serum specimens. Serum miRNAs are candidate biomarkers for deployment and environmental exposure in military service members.

Impact of Carrier Fluid Composition on Recovery of Nanoparticles and Proteins in Flow Field Flow Fractionation

PubMed Central

Schachermeyer, Samantha; Ashby, Jonathan; Kwon, MinJung; Zhong, Wenwan

2012-01-01

Flow field flow fractionation (F4) is an invaluable separation tool for large analytes, including nanoparticles and biomolecule complexes. However, sample loss due to analyte-channel membrane interaction limits extensive usage of F4 at present, which could be strongly affected by the carrier fluid composition. This work studied the impacts of carrier fluid (CF) composition on nanoparticle (NP) recovery in F4, with focus on high ionic strength conditions. Successful analysis of NPs in a biomolecules-friendly environment could expand the applicability of F4 to the developing field of nanobiotechnology. Recovery of the unfunctionalized polystyrene NPs of 199-, 102-, and 45-nm in CFs with various pH (6.2, 7.4 and 8.2), increasing ionic strength (0–0.1 M), and different types of co- and counter-ions, were investigated. Additionally, elution of the 85-nm carboxylate NPs and two proteins, human serum albumin (HSA) and immunoglobulin (IgG), at high ionic strengths (0–0.15 M) was investigated. Our results suggested that; 1) Electrostatic repulsion between the negatively charged NPs and the regenerated cellulose membrane was the main force to avoid particle adsorption on the membrane; 2) Larger particles experienced higher attractive force and thus were influenced more by variation in CF composition; and 3) Buffers containing weak anions or NPs with weak anion as the surface functional groups provided higher tolerance to the increase in ionic strength, owing to more anions being trapped inside the NP porous structure. Protein adsorption onto the membrane was also briefly investigated in salted CFs, using human serum albumin and immunoglobulin. We believe our findings could help to identify the basic carrier fluid composition for higher sample recovery in F4 analysis of nanoparticles in a protein-friendly environment, which will be useful for applying F4 in bioassays and in nanotoxicology studies. PMID:23058938

Increased Serum Al Levels in Hemodialysis Patients Kept Enhanced during a 2-Year Prospective Study.

PubMed

Reina de la Torre, Maria L; Lopez García de la Serrana, Herminia; Martí Del Moral, Loreto; Palomares Bayo, Magdalena; Navarro-Alarcón, Miguel

2017-07-01

The regulation of mineral homeostasis is altered in hemodialysis patients with renal insufficiency, producing increased risk for secondary diseases like cardiovascular ones. We hypothesized that risen serum aluminum (Al) concentration in hemodialysis patients kept enhanced during a 2-year longitudinal study is associated with enhanced cardiovascular risk and influenced by medical treatments. This study reports the prospective monitoring of serum Al levels in six-monthly samplings over 2 years in 116 hemodialysis patients and a control group of 50 healthy adults. The influence of other factors like sex, age, kidney transplant, disease etiology, and drug consumption was also considered. At each sampling, serum Al levels were significantly higher in the patients than in the healthy controls (P < 0.05). Levels in the patient group were statistically significantly lower at the third and fourth versus first and second samplings, which may be related to Al accumulation in tissues. Increased Al levels in patients were positively and significantly related to serum calcium (Ca) and uric acid levels. Serum Al concentrations were significantly lower in patients receiving vasodilators and diuretics. Higher serum Al levels in hemodialyzed patients administered with phosphate binders or anti-hyperkalemics are attributable to their usual Al salt content. The consumption of antianemic drugs increases Al absorption by forming more bioavailable complexes with the compounds in these drugs. In conclusion, this is the first study to indicate that cardiovascular problems associated with elevated serum Al levels in hemodialysis patients may be in part mitigated by administrating vasodilators and diuretics, which reduce these levels.

Organochlorine pesticide residue levels in blood serum of inhabitants from Veracruz, Mexico.

PubMed

Waliszewski, Stefan M; Caba, M; Herrero-Mercado, M; Saldariaga-Noreña, H; Meza, E; Zepeda, R; Martínez-Valenzuela, C; Gómez Arroyo, S; Villalobos Pietrini, R

2012-09-01

The objective of the present study was to monitor the levels of organochlorine pesticides HCB; α-, β-, γ-HCH; pp'DDE; op'DDT; and pp'DDT in blood serum of Veracruz, Mexico inhabitants. Organochlorine pesticides were analyzed in 150 blood serum samples that constituted that which remained after clinical analyses, using gas chromatography-electron-capture detection (GC-ECD). The results were expressed as milligrams per kilogram on fat basis and micrograms per liter on wet weight. Only the following pesticides were detected: p,p'-DDE was the major organochlorine component, detected in 100% of samples at mean 15.8 mg/kg and 8.4 μg/L; p,p'-DDT was presented in 41.3.% of monitored samples at mean 3.1 mg/kg and 1.4 μg/L; β-HCH was found in 48.6% of the samples at mean 4.9 mg/kg and 2.7 μg/L; op'DDT was determined to be in only 3.3% of monitored samples at mean 2.7 mg/kg and 1.4 μg/L. The pooled samples divided according to sex showed significant differences of β-HCH and pp'DDE concentrations in females. The samples grouped according to age presented the third tertile as more contaminated in both sexes, indicating age as a positively associated factor with serum organochlorine pesticide levels in Veracruz inhabitants.

Lyssavirus Surveillance in Bats, Bangladesh

PubMed Central

Niezgoda, Michael; Carroll, Darin S.; Keeler, Natalie; Hossain, Mohammed Jahangir; Breiman, Robert F.; Ksiazek, Thomas G.; Rupprecht, Charles E.

2006-01-01

Lyssavirus surveillance in bats was performed in Bangladesh during 2003 and 2004. No virus isolates were obtained. Three serum samples (all from Pteropus giganteus, n = 127) of 288 total serum samples, obtained from bats in 9 different taxa, neutralized lyssaviruses Aravan and Khujand. The infection occurs in bats in Bangladesh, but virus prevalence appears low. PMID:16704789

Determination of novel brominated flame retardants and polybrominated diphenyl ethers in serum using gas chromatography-mass spectrometry with two simplified sample preparation procedures.

PubMed

Gao, Le; Li, Jian; Wu, Yandan; Yu, Miaohao; Chen, Tian; Shi, Zhixiong; Zhou, Xianqing; Sun, Zhiwei

2016-11-01

Two simple and efficient pretreatment procedures have been developed for the simultaneous extraction and cleanup of six novel brominated flame retardants (NBFRs) and eight common polybrominated diphenyl ethers (PBDEs) in human serum. The first sample pretreatment procedure was a quick, easy, cheap, effective, rugged, and safe (QuEChERS)-based approach. An acetone/hexane mixture was employed to isolate the lipid and analytes from the serum with a combination of MgSO 4 and NaCl, followed by a dispersive solid-phase extraction (d-SPE) step using C18 particles as a sorbent. The second sample pretreatment procedure was based on solid-phase extraction. The sample extraction and cleanup were conducted directly on an Oasis HLB SPE column using 5 % aqueous isopropanol, concentrated sulfuric acid, and 10 % aqueous methanol, followed by elution with dichloromethane. The NBFRs and PBDEs were then detected using gas chromatography-negative chemical ionization mass spectrometry (GC-NCI MS). The methods were assessed for repeatability, accuracy, selectivity, limits of detection (LODs), and linearity. The results of spike recovery experiments in fetal bovine serum showed that average recoveries ranged from 77.9 % to 128.8 % with relative standard deviations (RSDs) from 0.73 % to 12.37 % for most of the analytes. The LODs for the analytes in fetal bovine serum ranged from 0.3 to 50.8 pg/mL except for decabromodiphenyl ethane. The proposed method was successfully applied to the determination of the 14 brominated flame retardants in human serum. The two pretreatment procedures described here are simple, accurate, and precise, and are suitable for the routine analysis of human serum. Graphical Abstract Workflow of a QuEChERS-based approach (top) and an SPE-based approach (bottom) for the detection of PBDEs and NBFRs in serum.

Fabrication of Fe3O4 Nanoparticle-coalesced Hydroxylated Multi-walled Carbon Nanotubes for the Analysis of Strychnine in Human Serum.

PubMed

Feng, Zufei; Xu, Yuehong; Wei, Shuguang; Zhang, Bao; Guan, Fanglin; Li, Shengbin

2015-01-01

A magnetic carbon nanomaterial for Fe3O4-modified hydroxylated multi-walled carbon nanotubes (Fe3O4-MWCNTs-OH) was prepared by the aggregating effect of Fe3O4 nanoparticles on MWCNTs-OH, and this material was combined with high-performance liquid chromatography (HPLC)/photodiode array detector (PAD) to determine strychnine in human serum samples. Some important parameters that could influence the extraction efficiency of strychnine were optimized, including the extraction time, amounts of Fe3O4-MWCNTs-OH, pH of sample solution, desorption solvent and desorption time. Under optimal conditions, the recoveries of spiked serum samples were between 98.3 and 102.7%, and the relative standard deviations (RSDs) ranged from 0.9 to 5.3%. The correlation coefficient was 0.9997. The LODs and LOQs of strychnine were 6.2 and 20.5 ng mL(-1), at signal-to-noise ratios of 3 and 10, respectively. These experimental results showed that the proposed method is feasible for the analysis of strychnine in serum samples.

Preanalytical considerations in detection of colorectal cancer in blood serum using Raman molecular imaging (Conference Presentation)

NASA Astrophysics Data System (ADS)

Treado, Patrick J.; Stewart, Shona D.; Smith, Aaron; Kirschner, Heather; Post, Christopher; Overholt, Bergein F.

2016-03-01

Colorectal cancer (CRC) is the third most common cancer in men and women in the United States. Raman Molecular Imaging (RMI) is an effective technique to evaluate human tissue, cells and bodily fluids, including blood serum for disease diagnosis. ChemImage Corporation, in collaboration with clinicians, has been engaged in development of an in vitro diagnostic Raman assay focused on CRC detection. The Raman Assay for Colorectal Cancer (RACC) exploits the high specificity of Raman imaging to distinguish diseased from normal dried blood serum droplets without additional reagents. Pilot Study results from testing of hundreds of biobank patient samples have demonstrated that RACC detects CRC with high sensitivity and specificity. However, expanded clinical trials, which are ongoing, are revealing a host of important preanalytical considerations associated with sample collection, sample storage and stability, sample shipping, sample preparation and sample interferents, which impact detection performance. Results from recent clinical studies will be presented.

Evaluation and optimization of a commercial blocking ELISA for detecting antibodies to influenza A virus for research and surveillance of mallards.

PubMed

Shriner, Susan A; VanDalen, Kaci K; Root, J Jeffrey; Sullivan, Heather J

2016-02-01

The availability of a validated commercial assay is an asset for any wildlife investigation. However, commercial products are often developed for use in livestock and are not optimized for wildlife. Consequently, it is incumbent upon researchers and managers to apply commercial products appropriately to optimize program outcomes. We tested more than 800 serum samples from mallards for antibodies to influenza A virus with the IDEXX AI MultiS-Screen Ab test to evaluate assay performance. Applying the test per manufacturer's recommendations resulted in good performance with 84% sensitivity and 100% specificity. However, performance was improved to 98% sensitivity and 98% specificity by increasing the recommended cut-off. Using this alternative threshold for identifying positive and negative samples would greatly improve sample classification, especially for field samples collected months after infection when antibody titers have waned from the initial primary immune response. Furthermore, a threshold that balances sensitivity and specificity reduces estimation bias in seroprevalence estimates. Published by Elsevier B.V.

Comparison of serum, EDTA plasma and P100 plasma for luminex-based biomarker multiplex assays in patients with chronic obstructive pulmonary disease in the SPIROMICS study.

PubMed

O'Neal, Wanda K; Anderson, Wayne; Basta, Patricia V; Carretta, Elizabeth E; Doerschuk, Claire M; Barr, R Graham; Bleecker, Eugene R; Christenson, Stephanie A; Curtis, Jeffrey L; Han, Meilan K; Hansel, Nadia N; Kanner, Richard E; Kleerup, Eric C; Martinez, Fernando J; Miller, Bruce E; Peters, Stephen P; Rennard, Stephen I; Scholand, Mary Beth; Tal-Singer, Ruth; Woodruff, Prescott G; Couper, David J; Davis, Sonia M

2014-01-08

As a part of the longitudinal Chronic Obstructive Pulmonary Disease (COPD) study, Subpopulations and Intermediate Outcome Measures in COPD study (SPIROMICS), blood samples are being collected from 3200 subjects with the goal of identifying blood biomarkers for sub-phenotyping patients and predicting disease progression. To determine the most reliable sample type for measuring specific blood analytes in the cohort, a pilot study was performed from a subset of 24 subjects comparing serum, Ethylenediaminetetraacetic acid (EDTA) plasma, and EDTA plasma with proteinase inhibitors (P100). 105 analytes, chosen for potential relevance to COPD, arranged in 12 multiplex and one simplex platform (Myriad-RBM) were evaluated in duplicate from the three sample types from 24 subjects. The reliability coefficient and the coefficient of variation (CV) were calculated. The performance of each analyte and mean analyte levels were evaluated across sample types. 20% of analytes were not consistently detectable in any sample type. Higher reliability and/or smaller CV were determined for 12 analytes in EDTA plasma compared to serum, and for 11 analytes in serum compared to EDTA plasma. While reliability measures were similar for EDTA plasma and P100 plasma for a majority of analytes, CV was modestly increased in P100 plasma for eight analytes. Each analyte within a multiplex produced independent measurement characteristics, complicating selection of sample type for individual multiplexes. There were notable detectability and measurability differences between serum and plasma. Multiplexing may not be ideal if large reliability differences exist across analytes measured within the multiplex, especially if values differ based on sample type. For some analytes, the large CV should be considered during experimental design, and the use of duplicate and/or triplicate samples may be necessary. These results should prove useful for studies evaluating selection of samples for evaluation of potential blood biomarkers.

Comparison of serum, EDTA plasma and P100 plasma for luminex-based biomarker multiplex assays in patients with chronic obstructive pulmonary disease in the SPIROMICS study

PubMed Central

2014-01-01

Background As a part of the longitudinal Chronic Obstructive Pulmonary Disease (COPD) study, Subpopulations and Intermediate Outcome Measures in COPD study (SPIROMICS), blood samples are being collected from 3200 subjects with the goal of identifying blood biomarkers for sub-phenotyping patients and predicting disease progression. To determine the most reliable sample type for measuring specific blood analytes in the cohort, a pilot study was performed from a subset of 24 subjects comparing serum, Ethylenediaminetetraacetic acid (EDTA) plasma, and EDTA plasma with proteinase inhibitors (P100™). Methods 105 analytes, chosen for potential relevance to COPD, arranged in 12 multiplex and one simplex platform (Myriad-RBM) were evaluated in duplicate from the three sample types from 24 subjects. The reliability coefficient and the coefficient of variation (CV) were calculated. The performance of each analyte and mean analyte levels were evaluated across sample types. Results 20% of analytes were not consistently detectable in any sample type. Higher reliability and/or smaller CV were determined for 12 analytes in EDTA plasma compared to serum, and for 11 analytes in serum compared to EDTA plasma. While reliability measures were similar for EDTA plasma and P100 plasma for a majority of analytes, CV was modestly increased in P100 plasma for eight analytes. Each analyte within a multiplex produced independent measurement characteristics, complicating selection of sample type for individual multiplexes. Conclusions There were notable detectability and measurability differences between serum and plasma. Multiplexing may not be ideal if large reliability differences exist across analytes measured within the multiplex, especially if values differ based on sample type. For some analytes, the large CV should be considered during experimental design, and the use of duplicate and/or triplicate samples may be necessary. These results should prove useful for studies evaluating selection of samples for evaluation of potential blood biomarkers. PMID:24397870

Improvement in the stability of serum samples stored in an automated refrigerated module.

PubMed

Parra-Robert, Marina; Rico-Santana, Naira; Alcaraz-Quiles, José; Sandalinas, Silvia; Fernández, Esther; Falcón, Isabel; Pérez-Riedweg, Margarita; Bedini, Josep Lluís

2016-12-01

In clinical laboratories it is necessary to know for how long the analytes are stable in the samples with specific storage conditions. Our laboratory has implemented the new Aptio Automation System (AAS) (Siemens Healthcare Diagnostics) where the analyzed samples are stored in a refrigerated storage module (RSM) after being sealed. The aim of the study was to evaluate the stability of serum samples with the AAS and comparing the results with a previous study using a conventional refrigerated system. Serum samples from a total of 50 patients were collected and for each of them 27 biochemical analytes were analyzed. The samples were divided in 5 sets of 10 samples. Each set was re-analyzed at one of the following times: 24, 48, 72, 96 and 120h. Stability was evaluated according to the Total Limit of Change (TLC) criteria, which combine both analytical and biologic variation. A total of 26 out of 27 analytes were stable at the end of the study according to TLC criteria. Lactate dehydrogenase was not stable at 48h observing a decrease in its concentration until the end of the study. In the previous study (conventional storage system) 9 biochemical analytes were not stable with an increase of their levels due to the evaporation process. The RSM connected to the AAS improves the stability of serum samples. This system avoids the evaporation process due to the sealing of samples and allows better control of the samples during their storage. Copyright © 2016 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Detection of trace fluoride in serum and urine by online membrane-based distillation coupled with ion chromatography.

PubMed

Lou, Chaoyan; Guo, Dandan; Wang, Nani; Wu, Shuchao; Zhang, Peimin; Zhu, Yan

2017-06-02

An online membrane-based distillation (MBD) coupled with ion chromatography (IC) method was proposed for automatic detection of trace fluoride (F - ) in serum and urine samples. The system consisted of a sample vessel, a lab-made membrane module and an ion chromatograph. Hydrophobic polytetrafluoroethylene (PTFE) hollow fiber membrane was used in MBD which was directly performed in serum and urine samples to eliminate the matrix interferences and enrich fluoride, while enabling automation. The determination of fluoride in biological samples was carried out by IC with suppressed conductometric detection. The proposed method feasibly determined trace fluoride in serum and urine matrices with the optimized parameters, such as acid concentration, distillation temperature, and distillation time, etc. Fluoride exhibited satisfactory linearity in the range of 0.01-5.0mg/L with a correlation coefficient of 0.9992. The limit of detection (LOD, S/N=3) and limit of quantification (LOQ, S/N=10) were 0.78μg/L and 2.61μg/L, respectively. The relative standard deviations of peak area and peak height were all less than 5.15%. The developed method was validated for the determination of fluoride in serum and urine with good spiked recoveries ranging between 97.1-101.9%. This method also can be proposed as a suitable alternative for the analysis of fluoride in other complex biological samples. Copyright © 2017. Published by Elsevier B.V.

Stability of Chronic Hepatitis-Related Parameters in Serum Samples After Long-Term Storage.

PubMed

Yu, Rentao; Dan, Yunjie; Xiang, Xiaomei; Zhou, Yi; Kuang, Xuemei; Yang, Ge; Tang, Yulan; Liu, Mingdong; Kong, Weilong; Tan, Wenting; Deng, Guohong

2017-06-01

Serum samples are widely used in clinical research, but a comprehensive research of the stability of parameters relevant to chronic hepatitis and the effect of a relatively long-term (up to 10 years) storage on the stability have rarely been studied. To investigate the stability of chronic hepatitis-related parameters in serum samples after long-term storage. The storage stability of common clinical parameters such as total bile acid (TBA), total bilirubin (TBIL), potassium, cholesterol, and protein parameters such as alanine aminotransferase (ALT), creatine kinase (CK), γ-glutamyltransferase (GGT), albumin, high-density lipoprotein (HDL) and also hepatitis B virus (HBV) DNA, hepatitis C virus (HCV) RNA, hepatitis B surface antigen (HBsAg), and chemokine (C-X-C motif) ligand 10 (CXCL10) were tested in serum samples after storing at -20°C or -70°C for 1, 2, 3, 7, 8, and 10 years. Levels of TBA, TBIL, and protein parameters such as ALT, CK, GGT, HDL, and HBsAg decreased significantly, but levels of potassium and cholesterol increased significantly after long-term storage, whereas blood glucose and triglycerides were stable during storage. HBV DNA remained stable at -70°C but changed at -20°C, whereas HCV RNA was stable after 1-, 2-, and 3-year storage. CXCL10 was still detectable after 8-year storage. Low temperatures (-70°C/80°C) are necessary for storage of serum samples in chronic hepatitis B research after long-term storage.

Amyloid Beta and Tau as Alzheimer's Disease Blood Biomarkers: Promise From New Technologies.

PubMed

Lue, Lih-Fen; Guerra, Andre; Walker, Douglas G

2017-07-01

The utility of the levels of amyloid beta (Aβ) peptide and tau in blood for diagnosis, drug development, and assessment of clinical trials for Alzheimer's disease (AD) has not been established. The lack of availability of ultra-sensitive assays is one critical issue that has impeded progress. The levels of Aβ species and tau in plasma and serum are much lower than levels in cerebrospinal fluid. Furthermore, plasma or serum contain high levels of assay-interfering factors, resulting in difficulties in the commonly used singulex or multiplex ELISA platforms. In this review, we focus on two modern immune-complex-based technologies that show promise to advance this field. These innovative technologies are immunomagnetic reduction technology and single molecule array technology. We describe the technologies and discuss the published studies using these technologies. Currently, the potential of utilizing these technologies to advance Aβ and tau as blood-based biomarkers for AD requires further validation using already collected large sets of samples, as well as new cohorts and population-based longitudinal studies.

Measurements of C-reactive protein (CRP) and nerve-growth-factor (NGF) concentrations in serum and urine samples of dogs with neurologic disorders.

PubMed

Kordass, Ulrike; Carlson, Regina; Stein, Veronika Maria; Tipold, Andrea

2016-01-08

The purpose of this study was to prove the hypothesis that C-reactive protein (CRP) and nerve growth factor (NGF) may be potential biomarkers for lower urinary tract disorders and may be able to distinguish between micturition dysfunctions of different origin in dogs with spinal cord diseases. NGF- and CRP- concentrations were measured in serum and urine samples using specific ELISA-Kits. Results in urine were standardized by urine-creatinine levels. CRP in serum was detectable in 32/76 and in urine samples in 40/76 patients. NGF could be measured in all serum and in 70/76 urine samples. Urinary CRP concentrations were significantly higher in dogs with micturition dysfunction (p = 0.0009) and in dogs with different neurological diseases (p = 0.0020) compared to the control group. However, comparing dogs with spinal cord disorders with and without associated micturition dysfunction no significant difference could be detected for NGF and CRP values in urine or serum samples. Additionally, levels did not decrease significantly, when measured at the time when the dogs regained the ability to urinate properly (urinary NGF p = 0.7962; urinary CRP p = 0.078). Urine samples with bacteria and/or leukocytes had no significant increase in urinary NGF (p = 0.1112) or CRP (p = 0.0534) concentrations, but higher CRP-levels in urine from dogs with cystitis were found compared to dogs without signs of cystitis. From these data we conclude that neither CRP nor NGF in urine or serum can be considered as reliable biomarkers for micturition disorders in dogs with spinal cord disorders in a clinical setting, but their production might be part of the pathogenesis of such disorders. Significantly higher levels of CRP could be found in the urine of dogs with micturition dysfunctions compared to control dogs. This phenomenon could potentially be explained by unspecific extrahepatic CRP production by smooth muscle cells in the dilated bladder.

Pharmacokinetic-pharmacodynamic study of subcutaneous injection of depot nandrolone decanoate using dried blood spots sampling coupled with ultrapressure liquid chromatography tandem mass spectrometry assays.

PubMed

Singh, Gurmeet K S; Turner, Leo; Desai, Reena; Jimenez, Mark; Handelsman, David J

2014-07-01

Testosterone (T) and nandrolone (N) esters require deep im injections by medical personnel but these often deposit injectate into sc fat so that more convenient sc self-administration may be feasible. To investigate the feasibility and pharmacology of sc injection of N decanoate in healthy men using dried blood spot (DBS) for frequent blood sampling without clinic visits. Healthy male volunteers received 100 mg N decanoate by a single sc injection. Finger-prick capillary blood was spotted onto filter paper before injection daily at home for 21 d and stored at room temperature. Venous whole blood was also spotted onto filter paper before and weekly for 3 wk after injection. DBS were extracted for assay of N and T by liquid chromatography tandem mass spectrometry in a single batch with serum concentrations estimated with adjustment for capillary blood sample volume and hematocrit to define peak (N) or nadir (T) time and concentration from individual daily measurements. Daily serum N peaked 2.50 ± 0.25 (SEM) ng/mL at a median (range) of 6 (4-13) days causing a reduction in serum T from 3.50 ± 0.57 ng/mL at baseline to a nadir of 0.38 ± 0.13 (SEM) ng/mL (89 ± 3% suppression) at a median (range) of 8 (5-16) days. Simultaneously sampled capillary, venous whole blood, and serum gave almost identical results for serum T and N. Finger-pricks and sc injections were well tolerated. This study demonstrates that A) DBS sampling with liquid chromatography mass spectrometry steroid analysis achieves frequent time sampling in the community without requiring clinic visits, venesection, or frozen serum storage, and B) androgen esters in an oil vehicle can be delivered effectively by sc injection, thus avoiding the need for medically supervised deep-im injections.

Polybrominated Diphenyl Ethers in Human Milk and Serum from the U.S. EPA MAMA Study: Modeled Predictions of Infant Exposure and Considerations for Risk Assessment

PubMed Central

Marchitti, Satori A.; Fenton, Suzanne E.; Mendola, Pauline; Kenneke, John F.; Hines, Erin P.

2016-01-01

Background: Serum concentrations of polybrominated diphenyl ethers (PBDEs) in U.S. women are believed to be among the world’s highest; however, little information exists on the partitioning of PBDEs between serum and breast milk and how this may affect infant exposure. Objectives: Paired milk and serum samples were measured for PBDE concentrations in 34 women who participated in the U.S. EPA MAMA Study. Computational models for predicting milk PBDE concentrations from serum were evaluated. Methods: Samples were analyzed using gas chromatography isotope-dilution high-resolution mass spectrometry. Observed milk PBDE concentrations were compared with model predictions, and models were applied to NHANES serum data to predict milk PBDE concentrations and infant intakes for the U.S. population. Results: Serum and milk samples had detectable concentrations of most PBDEs. BDE-47 was found in the highest concentrations (median serum: 18.6; milk: 31.5 ng/g lipid) and BDE-28 had the highest milk:serum partitioning ratio (2.1 ± 0.2). No evidence of depuration was found. Models demonstrated high reliability and, as of 2007–2008, predicted U.S. milk concentrations of BDE-47, BDE-99, and BDE-100 appear to be declining but BDE-153 may be rising. Predicted infant intakes (ng/kg/day) were below threshold reference doses (RfDs) for BDE-99 and BDE-153 but above the suggested RfD for BDE-47. Conclusions: Concentrations and partitioning ratios of PBDEs in milk and serum from women in the U.S. EPA MAMA Study are presented for the first time; modeled predictions of milk PBDE concentrations using serum concentrations appear to be a valid method for estimating PBDE exposure in U.S. infants. Citation: Marchitti SA, Fenton SE, Mendola P, Kenneke JF, Hines EP. 2017. Polybrominated diphenyl ethers in human milk and serum from the U.S. EPA MAMA Study: modeled predictions of infant exposure and considerations for risk assessment. Environ Health Perspect 125:706–713; http://dx.doi.org/10.1289/EHP332 PMID:27405099

Evaluation of the use of nonesterified fatty acids and β-hydroxybutyrate concentrations in pooled serum samples for herd-based detection of subclinical ketosis in dairy cows during the first week after parturition.

PubMed

Borchardt, Stefan; Staufenbiel, Rudolf

2012-04-15

To evaluate the use of nonesterified fatty acids (NEFA) and β-hydroxybutyrate (BHBA) concentrations in pooled serum samples for herd-based detection of subclinical ketosis (SCK) in dairy cows after calving. Cross-sectional study. 1,100 dairy cows from 110 herds. Blood samples were collected from 10 healthy cows/herd in the first week after parturition. Aliquots of serum were mixed to create a pooled sample. Concentrations of NEFA and BHBA were measured to estimate prevalence of SCK. Pooled sample test results were compared with those obtained for individual samples. Linear regression and receiver-operating characteristic curve analysis were performed; Bland-Altman plots were used to evaluate agreement between methods. Overall prevalence of SCK was 30.7%, 19.3%, and 13.6%, as determined by use of BHBA threshold concentrations of 1,000, 1,200, and 1,400 μmol/L, respectively. Pooled sample concentrations of NEFA and BHBA were significantly correlated (r = 0.98 and 0.97, respectively) with individual sample means and with the number of cows that had NEFA (R(2) range, 0.81 to 0.84) or BHBA (R(2) range, 0.65 to 0.76) concentrations above predefined thresholds. Pooled sample concentrations of NEFA and BHBA were very accurate to highly accurate for herd-based detection of SCK. Analysis of NEFA and BHBA concentrations in pooled serum samples was useful for herd-based detection of SCK. A sample size of 10 cows/herd was deemed adequate for monitoring dairy herds for SCK. Reference criteria specific to pooled samples should be used for this type of herd-based testing.

Yellow Fever Outbreak, Imatong, Southern Sudan

PubMed Central

Ofula, Victor O.; Sang, Rosemary C.; Konongoi, Samson L.; Sow, Abdourahmane; De Cock, Kevin M.; Tukei, Peter M.; Okoth, Fredrick A.; Swanepoel, Robert; Burt, Felicity J.; Waters, Norman C.; Coldren, Rodney L.

2004-01-01

In May 2003, the World Health Organization received reports about a possible outbreak of a hemorrhagic disease of unknown cause in the Imatong Mountains of southern Sudan. Laboratory investigations were conducted on 28 serum samples collected from patients in the Imatong region. Serum samples from 13 patients were positive for immunoglobulin M antibody to flavivirus, and serum samples from 5 patients were positive by reverse transcription–polymerase chain reaction with both the genus Flavivirus–reactive primers and yellow fever virus–specific primers. Nucleotide sequencing of the amplicons obtained with the genus Flavivirus oligonucleotide primers confirmed yellow fever virus as the etiologic agent. Isolation attempts in newborn mice and Vero cells from the samples yielded virus isolates from five patients. Rapid and accurate laboratory diagnosis enabled an interagency emergency task force to initiate a targeted vaccination campaign to control the outbreak. PMID:15207058

Biomarker-Based Prediction Models for Response to Treatment in Systemic Sclerosis-Related Interstitial Lung Disease

DTIC Science & Technology

2017-10-01

in the baseline samples of the Scleroderma Lung Study II (SLS II). We are currently analyzing whether these serum proteins have predictive...In this project, we use the valuable samples collected in the Scleroderma Lung Study II (SLSII) clinical trial and the observational cohort, GENISOS...determine key serum protein levels and transcript signatures in whole blood and skin samples collected in the SLSII study . The identified candidate

Comparison of plasma and tissue disposition of enrofloxacin in rainbow trout (Oncorhynchus mykiss) and common carp (Cyprinus carpio) after a single oral administration.

PubMed

Kyuchukova, Ralica; Milanova, Aneliya; Pavlov, Alexander; Lashev, Lubomir

2015-01-01

The aim of the study was to investigate the serum and tissue disposition of enrofloxacin and its active metabolite ciprofloxacin in rainbow trout (Oncorhynchus mykiss) and common carp (Cyprinus carpio) after a single oral administration at a dose of 10 mg kg(-1). Concentrations of enrofloxacin in the serum of rainbow trout showed high variability with two peaks at the third and 24th hour after administration. The highest concentrations were found in the liver. The curves of liver levels showed similar changes to the respective serum samples. In the muscles, enrofloxacin concentrations were also higher compared with the respective serum samples. Ciprofloxacin concentrations were lower and showed smaller variations in all investigated tissues. The serum and tissue concentrations of enrofloxacin and ciprofloxacin in common carp showed two peaks, with the first Cmax at the third hour after drug administration as in rainbow trout. Concentrations of both investigated substances were higher in the liver than in the serum. The differences in common carp were less pronounced in comparison with rainbow trout. Relatively high levels of both substances were found in the muscles. Seven days after treatment enrofloxacin concentrations in the serum and tissues were within the therapeutic levels for most of the sensitive microorganisms in trout. Lower concentrations of its metabolite ciprofloxacin were found in the investigated tissues at the last sampling point. Lower levels of both substances were found in carp.

Technical note: Evaluation of digital refractometers to estimate serum immunoglobulin G concentration and passive transfer in Jersey calves.

PubMed

McCracken, M M; Morrill, K M; Fordyce, A L; Tyler, H D

2017-10-01

Previous data have demonstrated that refractometers can be used to estimate serum IgG, and that a cut-point of 7.8% Brix should be used to identify failure of passive transfer (FPT) in 1-d-old Holstein calves. The objective of the present study was to validate the use of refractometry to estimate serum IgG concentrations and evaluate FPT in Jersey calves. Blood samples (n = 97) were obtained from 1- to 3-d-old Jersey calves and centrifuged at 3,300 × g for 20 min at 25°C. Serum was analyzed for % Brix, total protein (TP), and refractive index (nD) using a Sper Scientific Digital Refractometer (model #300036, Sper Scientific, Scottsdale, AZ) within 12 h of sampling. Samples were then frozen and later analyzed in the laboratory for IgG by radial immunodiffusion. The mean serum IgG concentration for all calves was 23.7 mg/mL (SD = 12.5), with a range of 2.3 to 65.5 mg/mL. Mean serum % Brix was 8.9 (SD = 1.1; range 6.5 to 12.0). Serum % Brix was moderately correlated with IgG concentration (r = 0.77). Total protein and IgG were moderately correlated (r = 0.790). Regression was used to determine cut-points for approximately 10, 12, and 14 mg of IgG/mL and to determine the sensitivity and specificity of refractometry to identify FPT (serum IgG <10 mg/mL at 24 h of life). Brix cut-points analyzed were 7.1, 7.3, and 7.6%; TP cut-points were 4.6, 5.0, and 5.5 g/dL; and nD cut-points were 1.34332, 1.34271, and 1.3448, respectively, for 10, 12, and 14 mg of IgG/mL. The 7.3% Brix and 4.6 g/dL TP cut-points resulted in the greatest percentage of samples being correctly classified. These data suggest that digital refractometry is an acceptable and rapid method to estimate immunoglobulin G in Jersey calf serum. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Serum prolactin in coeliac disease: a marker for disease activity

PubMed Central

Reifen, R.; Buskila, D.; Maislos, M.; Press, J.; Lerner, A.

1997-01-01

Accepted 21 April 1997
 Prolactin, a polypeptide hormone of anterior pituitary origin, has pronounced physiological effects on growth, reproduction, and osmoregulation. Increasing evidence indicates that prolactin also has an immunomodulatory influence on the immune system. The status of prolactin in patients with coeliac disease was investigated by obtaining serum samples from 48 patients with active and non-active coeliac disease. These were compared with samples from 20 children with familial Mediterranean fever and 65 normal controls. Serum prolactin in patients with active coeliac disease was significantly higher than in the other groups studied and reference values. Serum prolactin correlated well with the degree of mucosal atrophy and with the serum concentration of antiendomysial antibodies. Prolactin may play a part in immune modulation in the intestinal damage of coeliac disease and serve as a potential marker for disease activity.

 PMID:9301358

Accuracy of cotinine serum test to detect the smoking habit and its association with periodontal disease in a multicenter study.

PubMed

Duque, A; Martínez, P-J; Giraldo, A; Gualtero, D-F; Ardila, C-M; Contreras, A; Duarte, S; Lafaurie, G-I

2017-07-01

The validity of the surveys on self-reported smoking status is often questioned because smokers underestimate cigarette use and deny the habit. It has been suggested that self-report should be accompanied by cotinine test. This report evaluates the usefulness of serum cotinine test to assess the association between smoking and periodontal status in a study with a large sample population to be used in studies with other serum markers in epidemiologic and periodontal medicine researches. 578 patients who were part of a multicenter study on blood biomarkers were evaluated about smoking and its relation to periodontal disease. Severity of periodontal disease was determinate using clinical attachment loss (CAL). Smoking was assessed by a questionnaire and a blood sample drawn for serum cotinine determination. The optimal cut-off point for serum cotinine was 10 ng/ml. Serum cotinine showed greater association with severity of CAL than self-report for mild-moderate CAL [OR 2.03 (CI95% 1.16-3.53) vs. OR 1.08 (CI95% 0.62-1.87) ] advanced periodontitis [OR 2.36 (CI95% 1.30- 4.31) vs. OR 2.06 (CI95% 0.97-4.38) ] and extension of CAL > 3 mm [ OR 1.78 (CI95% 1.16-1.71) vs. 1.37 (CI95% 0.89-2.11)]. When the two tests were evaluated together were not shown to be better than serum cotinine test. Self-reported smoking and serum cotinine test ≥ 10ng/ml are accurate ,complementary and more reliable methods to assess the patient's smoking status and could be used in studies evaluating serum samples in large population and multicenter studies. The serum cotinine level is more reliable to make associations with the patient's periodontal status than self-report questionnaire and could be used in multicenter and periodontal medicine studies.

Transfer of perfluoroalkyl substances from mother to fetus in a Spanish birth cohort.

PubMed

Manzano-Salgado, Cyntia B; Casas, Maribel; Lopez-Espinosa, Maria-Jose; Ballester, Ferran; Basterrechea, Mikel; Grimalt, Joan O; Jiménez, Ana-María; Kraus, Thomas; Schettgen, Thomas; Sunyer, Jordi; Vrijheid, Martine

2015-10-01

Prenatal exposure to perfluoroalkyl substances (PFAS) might affect child health; thus estimating PFAS fetal burden is relevant. PFAS fetal burden is best estimated in cord samples; previous studies have used either maternal plasma or serum during pregnancy as proxy, but their validity is not clear. We aimed to evaluate PFAS transfer between mother and fetus and determine its predictors in a Spanish birth cohort. We measured perfluorobutane sulfonate (PFBS), perfluorohexane sulfonate (PFHxS), perfluorooctane sulfonate (PFOS), perfluorooctanoate (PFOA), and perfluorononanoate (PFNA) in maternal blood and cord serum from 66 mother-child pairs. We used Spearman's rank coefficients to correlate PFAS concentrations in first trimester maternal plasma and serum, with cord serum samples. We assessed PFAS placental transfer by calculating maternal to cord ratios and examined their association with maternal socio-demographic characteristics and child sex using linear regression models. Median concentrations of PFAS (ng/mL) of PFHxS, PFOS, PFOA, and PFNA in maternal plasma (0.79, 6.18, 2.85 and 0.84, respectively) and serum (0.84, 6.99, 2.97 and 0.85) were higher than in cord serum (0.40, 1.86, 1.90 and 0.32). PFBS was not detected. Positive Spearman's correlations (p-values<0.001) were found between maternal plasma and serum (⍴≥0.80), maternal plasma and cord (⍴≥0.66), and maternal serum and cord samples (⍴≥0.67). Maternal plasma to cord ratios were above 1 (PFHxS: 2.35 [95%CI: 2.05, 2.70], PFOS: 3.33 [3.05, 3.62], PFOA: 1.37 [1.27, 1.48], PFNA: 2.39 [2.18, 2.63]); maternal serum to cord ratios were similar. Maternal to cord ratios decreased with maternal age, but not with other socio-demographic factors. Our results suggest that PFAS fetal body burden can be assessed using as proxy maternal plasma or serum collected early in pregnancy. Maternal age might influence PFAS placental transfer. Copyright © 2015 Elsevier Inc. All rights reserved.

Inverse association linking serum levels of potential antioxidant vitamins with C-reactive protein levels using a novel analytical approach.

PubMed

Cheng, Hui G; Alshaarawy, Omayma; Cantave, Marven D; Anthony, James C

2016-10-01

Exposures to antioxidants (AO) are associated with levels of C-reactive protein (CRP), but the pattern of evidence is mixed, due in part to studying each potential AO, one at a time, when multiple AO exposures might affect CRP levels. By studying multiple AO via a composite indicator approach, we estimate the degree to which serum CRP level is associated with serum AO level. Standardised field survey protocols for the US National Health and Nutrition Examination Survey (NHANES) 2003-2006 yielded nationally representative cross-sectional samples of adults aged 20 years and older (n 8841). NHANES latex-enhanced nephelometry quantified serum CRP levels. Liquid chromatography quantified serum concentrations of vitamins A, E and C and carotenoids. Using structural equations, we regressed CRP level on AO levels, and derived a summary estimate for a composite of these potential antioxidants (CPA), with covariates held constant. The association linking CPA with CRP was inverse, stronger for slightly elevated CRP (1·8≤CRP<10 mg/l; slope= -1·08; 95 % CI -1·39, -0·77) and weaker for highly elevated CRP (≥10 mg/l; slope= -0·52; 95 % CI -0·68, -0·35), with little change when covariates were added. Vitamins A and C, as well as lutein+zeaxanthin, were prominent contributors to the composite. In these cross-sectional data studied via a composite indicator approach, the CPA level and the CRP level were inversely related. The stage is set for more confirmatory longitudinal or intervention research on multiple vitamins. The composite indicator approach might be most useful in epidemiology when several exposure constructs are too weakly inter-correlated to be studied via formal measurement models for underlying latent dimensions.

Simultaneous determination of phosphatidylcholine-derived quaternary ammonium compounds by a LC-MS/MS method in human blood plasma, serum and urine samples.

PubMed

Steuer, Christian; Schütz, Philipp; Bernasconi, Luca; Huber, Andreas R

2016-01-01

The determination of circulating trimethylamine-N-oxide (TMAO), choline, betaine, l-carnitine and O-acetyl-l-carnitine concentration in different human matrices is of great clinical interest. Recent results highlighted the prognostic value of TMAO and quaternary ammonium containing metabolites in the field of cardiovascular and kidney diseases. Herein, we report a method for the rapid and simultaneous measurement of closely related phosphatidylcholine-derived metabolites in three different biological matrices by stable isotope dilution assay. Plasma, serum and urine samples were simply deproteinized and separated by HILIC-chromatography. Detection and quantification were performed using LC-MS/MS with electrospray ionization in positive mode. For accuracy and precision, full calibration was performed covering more than the full reference range. Assay performance metrics include intra- and interday imprecision were below 10% for all analytes. To exclude matrix effects standard addition methods were applied for all matrices. It was shown that calibration standards and quality control prepared in water can be used instead of matrix-matched calibration and controls. The LC/MS/MS-based assay described in this article may improve future clinical studies evaluating TMAO and related substances as prognostic markers for cardiovascular risk and all-cause mortality in different patient populations. Copyright © 2015 Elsevier B.V. All rights reserved.

Determinants of serum zinc concentrations in a population of French middle-age subjects (SU.VI.MAX cohort).

PubMed

Arnaud, J; Touvier, M; Galan, P; Andriollo-Sanchez, M; Ruffieux, D; Roussel, A-M; Hercberg, S; Favier, A

2010-10-01

Few studies have been conducted on determinants of serum zinc concentration, especially in France. The aim of this study was to investigate the relationships between serum zinc concentrations, and age, education, living area and life style in a large sample cohort of French adults. Blood samples were collected after 12-h fasting in French adults (7448 women, 35-65 years old and 4926 men, 45-65 years old) participating to the SUpplementation in AntioXidant VItamins and Minerals cohort at enrolment. Serum zinc was determined by flame atomic absorption. Serum zinc concentration was lower in women (mean: 12.9 μmol/l, 95% confidence interval: 9.2-16.6 μmol/l) compared with men (mean: 13.4 μmol/l, 95% confidence interval: 9.7-17.1 μmol/l, P<0.0001). In total, 10% of the women showed serum zinc values <10.7 μmol/l and 10% of the men showed serum zinc values <11.3 μmol/l. Age was negatively associated with serum zinc only in men (P=0.001). In women, a positive association between smoking and serum zinc concentration (P=0.0003), and a negative relationship between highest education level and serum zinc concentration (P=0.01) were observed. With regard to geographical areas, the highest serum zinc concentrations were found in the Center, and the lowest in the Southwest of France for both men and women. The association between serum zinc concentrations and food categories or macronutrient intake disappeared after stratifying by gender. Age, gender and geographic area seem the main determinants of serum zinc concentrations in this study.

Thyroglobulin is a more sensitive indicator of iodine deficiency than thyrotropin: development and evaluation of dry blood spot assays for thyrotropin and thyroglobulin in iodine-deficient geographical areas.

PubMed

Missler, U; Gutekunst, R; Wood, W G

1994-03-01

Immunometric assays were developed for thyrotropin and thyroglobulin using time-resolved fluorescence as the measurement signal. The assays were suitable for measurements in serum/plasma or in dry blood spots (3 mm diameter). Both assays have acceptable coefficients of variation for dry blood spots (intra-assay median CV < 10%, interassay CV < 15%) in the concentration range of interest (thyrotropin 3-250 mU/l, thyroglobulin 10-500 micrograms/l). The relatively high CV values are not only due to the assay design but also the inhomogeneity of the samples used. For serum samples the median intra-assay CV was < 3% for thyrotropin in the range 0.1-50 mU/l and for thyroglobulin between 2 and 500 micrograms/l. The corresponding inter-assay CV were less than 5%. The assays were evaluated in field studies carried out under auspices of International Council for Control of Iodine Deficiency Disorders (ICCIDD) with the support of UNICEF in Algeria, Peru, India and Zimbabwe, and were found to be practical inasmuch as dry blood spot samples could be transported without special precautions for up to 5-6 weeks without significant loss in immunoreactivity. This agrees with other findings. The results showed that serum thyroglobulin levels are a more sensitive indicator of iodine deficiency than thyrotropin; elevated thyroglobulin levels were found in 182/304 children in Zimbabwe compared with elevated thyrotropin level in 28/304 cases. 213/304 children had enlarged thyroid glands. The cut-off levels used here were 4.5 mU/l thyrotropin and 20 micrograms/l for thyroglobulin, both in whole blood. The assays proved useful for assessing the efficacy of iodine therapy, either by oral dosage or intramuscularly (iodised oil).(ABSTRACT TRUNCATED AT 250 WORDS)

Presence of antigen and antibodies in serum and genital discharges of cows from dairy herds naturally infected with Leptospira interrogans serovar hardjo.

PubMed

Dhaliwal, G S; Murray, R D; Dobson, H; Montgomery, J; Ellis, W A

1996-03-01

Samples of cervico-vaginal mucus from 163 bulling cows (group 1) and post calving discharges from 59 newly calved cows (group 2) in five dairy herds naturally infected with Leptospira interrogans serovar hardjo were examined for the presence of antigen and IgG and IgA antibodies by using two ELISA systems which were protein or carbohydrate based. Corresponding serum samples were examined for systemic immune responses by using a microscopic agglutination test (MAT) and IgG-ELISA tests. Antigen was detected by direct immunofluorescence in six of the 163 samples of cervico-vaginal mucus. Both IgG and IgA antibodies were detected by ELISA in the genital discharges with a prevalence much higher than that obtained by the MAT but lower than that observed with the serum IgG-ELISA. Combining both groups, none of the MAT-positive cattle was negative by serum-ELISA. By using the protein or carbohydrate fraction serum IgG-ELISA assays, respectively, 29 or 41 per cent of the MAT-negative cows were positive at a titre of at least 1:40. Similarly, eight or 23 samples (10 or 27 per cent) had titres of at least 1:20 in the genital discharge ELISA for IgG and IgA antibodies, respectively. The serum IgG-ELISA was the most efficient in detecting hardjo antibodies, but in group 2 the IgG- and IgA-ELISA of the post calving discharge proved to be equally effective.

Investigation of effects of 60-Hz electric and magnetic fields on operant and social behavior and on the neuroendocrine system of nonhuman primates: Neuroendocrine portion of Experiment IV

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rogers, W.R.; Rhodes, J.W.

1992-08-31

This quarterly report covers the neuroendocrine Portion of Experiment IV. Serum melatonin concentration was measured in individual baboons, each implanted with a chronically indwelling venous cannula. As in Experiment III the system of six automatic blood samplers was used to achieve undisturbed, 24 hr per day, simultaneous blood sampling from six individual subjects. The objective of the neuroendocrine portion of Experiment IV was to determine if 30 kV/m electric and 1.0 G magnetic field (E/MF) exposure produced a 50% decline in nocturnal serum melatonin concentration. Other groups of subjects were tested concurrently during Experiment IV to assess E/MF effects onmore » group social and individual operant behavior. The results of these experiments will be covered respectively in the next two quarterly reports. The results of Experiment IV, as was the case with the result of Experiments III and IIIA, provide little or no evidence that E/MF exposure, under the conditions of these experiments, affects nocturnal serum melatonin concentrations of nonhuman primates. Together the negative results of Experiments III, IIA and IV indicate that day-time exposure of primates to slow-onset/offset, regularly-scheduled E/MF does not produce melatonin suppression, strongly suggesting that such exposure would not affect human melatonin either. However, before concluding that E/MF exposure in general has no effect on primate melatonin, nightime exposure needs to be examined, and the possibility, suggested by the Pilot Experiment, that fast onset/offset, irregularly-scheduled E/MF can completely suppress melatonin needs to be investigated.« less

Investigation of effects of 60-Hz electric and magnetic fields on operant and social behavior and on the neuroendocrine system of nonhuman primates: Neuroendocrine portion of Experiment IV. Quarterly report No. 38

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rogers, W.R.; Rhodes, J.W.

1992-08-31

This quarterly report covers the neuroendocrine Portion of Experiment IV. Serum melatonin concentration was measured in individual baboons, each implanted with a chronically indwelling venous cannula. As in Experiment III the system of six automatic blood samplers was used to achieve undisturbed, 24 hr per day, simultaneous blood sampling from six individual subjects. The objective of the neuroendocrine portion of Experiment IV was to determine if 30 kV/m electric and 1.0 G magnetic field (E/MF) exposure produced a 50% decline in nocturnal serum melatonin concentration. Other groups of subjects were tested concurrently during Experiment IV to assess E/MF effects onmore » group social and individual operant behavior. The results of these experiments will be covered respectively in the next two quarterly reports. The results of Experiment IV, as was the case with the result of Experiments III and IIIA, provide little or no evidence that E/MF exposure, under the conditions of these experiments, affects nocturnal serum melatonin concentrations of nonhuman primates. Together the negative results of Experiments III, IIA and IV indicate that day-time exposure of primates to slow-onset/offset, regularly-scheduled E/MF does not produce melatonin suppression, strongly suggesting that such exposure would not affect human melatonin either. However, before concluding that E/MF exposure in general has no effect on primate melatonin, nightime exposure needs to be examined, and the possibility, suggested by the Pilot Experiment, that fast onset/offset, irregularly-scheduled E/MF can completely suppress melatonin needs to be investigated.« less

Influence of storage conditions on aluminum concentrations in serum, dialysis fluid, urine, and tap water.

PubMed

Wilhelm, M; Ohnesorge, F K

1990-01-01

The influence of storage temperature, vessel type, and treatment on alterations of aluminum (Al) concentrations in serum, urine, and dialysis fluid samples was studied at three different concentrations for each sample over an 18-month period. Furthermore, the influence of acidification on Al levels in tap water, urine, and dialysis fluid samples was studied over a four-month period. Al was measured by atomic absorption spectrometry. Sample storage in glass vessels was unsuitable, whereas only minor alterations of Al levels were observed with storage in polypropylene tubes, polystyrene tubes, and Monovettes. By using appropriate plastic containers, acid washing of the vessels showed no improvement. Frozen storage was superior compared with 4 degrees C, whereas storage at -80 degrees C offered no advantage compared with storage at -20 degrees C. Acidification of tap water samples was necessary to stabilize Al levels during storage. No striking effect of acidification on Al levels in urine and dialysis fluid samples was found. It is concluded that longterm storage of serum, urine, tap water, and dialysis fluid samples is possible if appropriate conditions are used.

Serologic Evidence of Lyssavirus Infection in Bats, Cambodia

PubMed Central

Molia, Sophie; Audry, Laurent; Hout, Sotheara; Ngin, Sopheak; Walston, Joe; Bourhy, Hervé

2004-01-01

In Cambodia, 1,303 bats of 16 species were tested for lyssavirus. No lyssavirus nucleocapsid was detected in 1,283 brains tested by immunofluorescence assay. Antibodies against lyssaviruses were detected by enzyme-linked immunosorbent assay in 144 (14.7%) of 981 serum samples. Thirty of 187 serum samples contained neutralizing antibodies against different lyssaviruses. PMID:15663870

Prevalence of equine Piroplasmosis and its association with tick infestation in the State of Sao Paulo, Brazil

USDA-ARS?s Scientific Manuscript database

Serum samples were collected from 582 horses from 40 stud farms in the State of São Paulo and tick (Acari: Ixodidae) infestations were evaluated on them. Serum samples were subjected to the complement fixation test (CFT) and a competitive inhibition ELISA (cELISA) for Babesia caballi and Theileria e...

Sequences of Zika Virus Genomes from a Pediatric Cohort in Nicaragua.

PubMed

Oldfield, Lauren M; Fedorova, Nadia; Puri, Vinita; Shrivastava, Susmita; Amedeo, Paolo; Durbin, Alan; Rocchi, Iara; Williams, Torrey; Shabman, Reed S; Tan, Gene S; Balmaseda, Angel; Kuan, Guillermina; Saborio, Saira; Gordon, Aubree; Harris, Eva; Pickett, Brett E

2018-06-14

We report here the whole-genome sequence of 11 Zika virus (ZIKV) samples from six pediatric patients in Nicaragua. Serum samples were collected, and ZIKV was isolated in tissue culture. Both serum and virus isolates were sequenced. The consensus ZIKV genomes are greater than 99% identical to each other. Copyright © 2018 Oldfield et al.

Clinical Validation of a miRNA Blood Test to Identify High-Risk Individuals Eligible for Low-Dose Computed Tomography Screening for Lung Cancer Early Detection

DTIC Science & Technology

2015-10-01

screening. The project requires the use of serum samples obtained from at-risk subjects (heavy smokers with defined characteristics) undergoing...screening. The project requires the use of serum samples obtained from at-risk subjects (heavy smokers with defined characteristics) that were collected

[Applying competitive polymerase chain reaction to the detection of hepatitis B virus DNA].

PubMed

Wang, Ling; Yang, Peng; Li, Shuang-qing; Xu, Shu-hui; Cao, Gui-qun; Zhang, Fa-qiang; Zhang, Mei-xia; Chen, Qing-ying; Xia, Qing-jie; Liu, Kai; Tang, Fang; Zhang, Yuan-zheng

2004-11-01

To reduce the rate of accidental false negative result in the HBV DNA PCR test on clinical serum samples. A competitive polymerase chain reaction (C-PCR) was used to decrease the false negative ratio. In the C-PCR, a constructed inner control DNA was added for co-amplification with the HBV target DNA. In a 20 microl C-PCR system, about 60 to 200 copies of inner control DNA could give apparent co-amplification signal band after electrophoresis on a 2% agarose gel. Five of 120 samples of clinical serum (4.2%) could not be amplified. C-PCR has the advantage of yielding information on false negative in the HBV DNA PCR assay of clinical serum samples.

A 3D-Printed, Portable, Optical-Sensing Platform for Smartphones Capable of Detecting the Herbicide 2,4-Dichlorophenoxyacetic Acid

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Yijia; Zeinhom, Mohamed M. A.; Yang, Mingming

Onsite rapid detection of herbicide and herbicide residuals in environmental and biological specimens is important for agriculture, environment, food safety, and health care. Traditional method for herbicide detection requires expensive laboratory equipment and a long turn-round time. In this work, we developed a single-stripe microliter plate smartphone colorimetric device for rapid and low-cost in-field test. This portable smartphone platform is capable of screening 8 samples in a microplate single-stripe. The device combined the advantages of small size (50×100×160 mm3) and low cost ($10). The platform was calibrated by using two different dye solutions, i.e. methyl blue (MB) and Rhodamine B,more » for green and red channels. The results showed good correlation with results attained from a traditional laboratory reader. We demonstrated the application of this platform for an herbicide, 2,4-Dichlorophenoxyacetic acid detection in the range of 1 ppb to 80 ppb. Spiked samples of tap water, rat serum, plasma and human serum were tested by our device. Recoveries obtained varied from 95.6% to 105.2% for all spiked samples using the microplate reader and from 93.7% to 106.9% using the smartphone device. This work validated that the smartphone optical sensing platform is comparable to the commercial microplate reader, it is eligible for onsite rapid and low-cost detection of herbicide for environmental evaluation and biological monitoring.« less

p16 promoter hypermethylation: A useful serum marker for early detection of gastric cancer

PubMed Central

Abbaszadegan, Mohammad Reza; Moaven, Omeed; Sima, Hamid Reza; Ghafarzadegan, Kamran; A'rabi, Azadeh; Forghani, Mohammad Naser; Raziee, Hamid Reza; Mashhadinejad, Ali; Jafarzadeh, Mostafa; Esmaili-Shandiz, Ehsan; Dadkhah, Ezzat

2008-01-01

AIM: To determine p16 promoter hypermethylation in gastric tumoral tissue and serum samples, its impact on p16-protein expression, and correlation with clinical and histological features. METHODS: Samples were obtained from 52 histologically confirmed cases of gastric adenocarcinoma. Gastric tissue and serum of 50 age- and sex-matched individuals with normal gastroscopy and biopsy were obtained as control samples. Methylation-specific polymerase chain reaction (MSP) was used to evaluate methylation status of p16 promoter. p16-protein expression was analyzed by immunohistochemical staining on paraffin-embedded sections. RESULTS: Methylation was detected in 44.2% (23/52) of tumoral tissues. 60.9% of them were also methylated in serum, i.e., 26.9% of all patients (14/52). Methylation was not detected in tissue and sera of control samples. p16-protein expression was decreased in 61.5% of cases (32/52), and was significantly associated with promoter hypermethylation (P < 0.001). Methylation was significantly more frequent in higher pathological grades (P < 0.05). Methylation was not associated with other clinicopathological features and environmental factors including H pylori infection and smoking. CONCLUSION: p16 promoter hypermethylation is an important event in gastric carcinogenesis. It is the principle mechanism of p16 gene silencing. It is related to malignant tumor behavior. Detection of DNA methylation in serum may be a biomarker for early detection of gastric cancer. PMID:18395906

Application of Fourier-transform infrared (FT-IR) spectroscopy for simple and easy determination of chylomicron-triglyceride and very low density lipoprotein-triglyceride.

PubMed

Sato, Kenichi; Seimiya, Masanori; Kodera, Yoshio; Kitamura, Akihide; Nomura, Fumio

2010-02-01

Fourier-transform infrared (FT-IR) spectroscopy is a simple and reagent-free physicochemical analysis method, and is a potential alternative to more time-consuming and labor-intensive procedures. In this study, we aimed to use FT-IR spectroscopy to determine serum concentrations of chylomicron-triglyceride (TG) and very low density lipoprotein (VLDL)-TG. We analyzed a chylomicron fraction and VLDL fraction, which had been obtained by ultracentrifugation, to search for wavelengths to designate to each fraction. Then, partial least square (PLS) calibrations were developed using a training set of samples, for which TG concentrations had been determined by conventional procedures. Validation was conducted with another set of samples using the PLS model to predict serum TG concentrations on the basis of the samples' IR spectra. We analyzed a total of 150 samples. Serum concentrations of chylomicron-TG and VLDL-TG estimated by FT-IR spectroscopy agreed well with those obtained by the reference method (r=0.97 for both lipoprotein fractions). FT-IR spectrometric analysis required 15mul of serum and was completed within 1min. Serum chylomicron-TG and VLDL-TG concentrations can be determined with FT-IR spectroscopy. This rapid and simple test may have a great impact on the management of patients with dyslipidemia. Copyright 2009. Published by Elsevier B.V.

Development, validation and application of an ICP-MS/MS method to quantify minerals and (ultra-)trace elements in human serum.

PubMed

Meyer, Sören; Markova, Mariya; Pohl, Gabriele; Marschall, Talke A; Pivovarova, Olga; Pfeiffer, Andreas F H; Schwerdtle, Tanja

2018-09-01

Multi-element determination in human samples is very challenging. Especially in human intervention studies sample volumes are often limited to a few microliters and due to the high number of samples a high-throughput is indispensable. Here, we present a state-of-the-art ICP-MS/MS-based method for the analysis of essential (trace) elements, namely Mg, Ca, Fe, Cu, Zn, Mo, Se and I, as well as food-relevant toxic elements such as As and Cd. The developed method was validated regarding linearity of the calibration curves, method LODs and LOQs, selectivity and trueness as well as precision. The established reliable method was applied to quantify the element serum concentrations of participants of a human intervention study (LeguAN). The participants received isocaloric diets, either rich in plant protein or in animal protein. While the serum concentrations of Mg and Mo increased in participants receiving the plant protein-based diet (above all legumes), the Se concentration in serum decreased. In contrast, the animal protein-based diet, rich in meat and dairy products, resulted in an increased Se concentration in serum. Copyright © 2018 Elsevier GmbH. All rights reserved.

Fast detection of Piscirickettsia salmonis in Salmo salar serum through MALDI-TOF-MS profiling.

PubMed

Olate, Verónica R; Nachtigall, Fabiane M; Santos, Leonardo S; Soto, Alex; Araya, Macarena; Oyanedel, Sandra; Díaz, Verónica; Marchant, Vanessa; Rios-Momberg, Mauricio

2016-03-01

Piscirickettsia salmonis is a pathogenic bacteria known as the aetiological agent of the salmonid rickettsial syndrome and causes a high mortality in farmed salmonid fishes. Detection of P. salmonis in farmed fishes is based mainly on molecular biology and immunohistochemistry techniques. These techniques are in most of the cases expensive and time consuming. In the search of new alternatives to detect the presence of P. salmonis in salmonid fishes, this work proposed the use of MALDI-TOF-MS to compare serum protein profiles from Salmo salar fish, including experimentally infected and non-infected fishes using principal component analysis (PCA). Samples were obtained from a controlled bioassay where S. salar was challenged with P. salmonis in a cohabitation model and classified according to the presence or absence of the bacteria by real time PCR analysis. MALDI spectra of the fish serum samples showed differences in its serum protein composition. These differences were corroborated with PCA analysis. The results demonstrated that the use of both MALDI-TOF-MS and PCA represents a useful tool to discriminate the fish status through the analysis of salmonid serum samples. Copyright © 2016 John Wiley & Sons, Ltd.

Serum cation profile of broilers at various stages of exposure to deoxynivalenol.

PubMed

Yunus, Agha Waqar; Böhm, Josef

2013-05-01

The present experiment was carried out to investigate if levels of serum cations in broilers are modulated differently at various stages of exposure to deoxynivalenol (DON). Male broiler chicks at 7 days of age were fed a basal diet (0.27 mg of DON; 0.01 mg of zearalenone/kg), or either a low DON diet (1.68 mg of DON; 0.15 mg of zearalenone/kg) or a high DON diet (12.21 mg of DON; 1.09 mg of zearalenone/kg) produced using extracts from Fusarium graminearum cultures. Blood samples from the birds were collected during weeks 2, 4, and 5 of exposure. The high DON diet resulted in lower serum calcium levels compared to the basal diet at all the 3 sampling stages, while the low DON diet resulted in lower serum calcium levels only during weeks 2 and 5. Serum potassium levels were reduced under both the DON diets during weeks 2 and 5, while no diet-associated changes were found for serum levels of magnesium, sodium, and zinc. Under the present experimental conditions, the serum levels of calcium were consistently modulated in the broilers exposed to the DON-contaminated diets. The modulation of serum levels of potassium was, however, dependent upon the stage of exposure to DON.

Avian influenza A (H5N1) virus antibodies in pigs and residents of swine farms, southern China.

PubMed

Cao, Nan; Zhu, Wanjun; Chen, Ye; Tan, Likai; Zhou, Pei; Cao, Zhenpeng; Ke, Changwen; Li, Yugu; Wu, Jie; Qi, Wenbao; Jiao, Peirong; Zhang, Guihong

2013-12-01

Since 1997, the H5 avian influenza viruses (AIVs) circulating in China have become an international concern. Clade 2.3.2 of H5N1 AIVs is genetically distinct from the viruses isolated before 2007 and antigenically different from the vaccine strains widely used in China. Swine farms in rural China are thought to play an important role in AIVs ecology. A seroepidemiological study was undertaken among swine farm residents and pigs to understand the prevalence of antibodies against H5N1 AIVs in southern China. During the period March 24, 2008 to December 25, 2012,serum samples were collected from 1606 swine farm residents on 40 swine farms in southern China. A total of 1980 pigs' serum samples were collected in the same swine farms where swine workers' serum samples were collected from March 2009 to March 2013. For a control group, 104 serum samples were collected from healthy city residents in Nanchang. All the serum samples were collected to perform hemagglutination inhibition (HI) and (neutralization) NT assays to investigate the prevalence of H5N1 AIV infections in southern China. Sixteen human samples were positive by HI assay and 10 of these were also positive by NT assay against H5N1. No serum samples from human control and pigs were HI positive for H5N1 AIV. Our results demonstrate minimal transmission H5N1 AIV from birds to pigs in the swine farms studied and the risk of poultry-to-human and poultry-to-pig transmission for at least clades 2.3.2 seemed very low. This study provides the first data regarding antibodies against H5N1 AIV in humans and pigs on swine farms in China. The findings of this study can serve as a baseline for additional serologic studies to assess transmission of H5N1 viruses between avian species, pigs and swine workers. Copyright © 2013 Elsevier B.V. All rights reserved.

Use of cerebrospinal fluid and serum samples impregnated on FTATM Elute filter paper for the diagnosis of infections caused by Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae.

PubMed

Fukasawa, Lucila Okuyama; Gonçalves, Maria Gisele; Higa, Fábio Takenori; Castilho, Euclides Ayres; Ibarz-Pavón, Ana Belén; Sacchi, Claudio Tavares

2017-01-01

The lack of information regarding the burden of acute bacterial meningitis in Latin America leads to a reduction in the estimated incidence rates of the disease, and impairs public health decisions on the use and follow-up of preventive interventions, particularly, the evaluation of existing vaccination policies. The use of the real-time PCR in diagnostic routine procedures has resulted in a substantial increase in confirmed bacterial meningitis cases. However, in resource-poor countries, these assays are only available in reference laboratories. Sample transportation to these laboratories is a critical constraint, as it requires specialized, high cost courier services. To overcome this barrier we evaluated the use of FTATM Elute filter paper cards for the conservation and processing of samples under normal environmental conditions, as they would be when transported from remote and under-equipped healthcare facilities to the reference centers. A total of 401 samples received in 2015 as part of Sao Paulo's national surveillance for routine diagnosis were selected for this study. The sensitivity and specificity of real-time PCR were evaluated using fresh serum and cerebrospinal fluid (CSF) samples processed using our laboratory's standard DNA extraction, and processing the same samples after being dried and stored on FTATM card, and DNA extracted following the manufacturer's instructions. The sensitivities for detection of Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae from CSF dried and stored on FTATM cards were 98%, 92%, and 100%, respectively, and with serum samples were 73%, 88%, and 100%, respectively. When compared to our laboratory's standard methodology, results showed high concordance, with Kappa index ranges of 0.9877-1.00 for CSF, and 0.8004-1.00 for serum samples. The use of FTATM cards for CSF and serum conservation and transport represents a rapid, reliable, and cost-effective alternative that will allow obtaining valuable epidemiological information that would otherwise be lost.

Comparison of Performance Characteristics of Aspergillus PCR in Testing a Range of Blood-Based Samples in Accordance with International Methodological Recommendations.

PubMed

Springer, Jan; White, P Lewis; Hamilton, Shanna; Michel, Denise; Barnes, Rosemary A; Einsele, Hermann; Löffler, Juergen

2016-03-01

Standardized methodologies for the molecular detection of invasive aspergillosis (IA) have been established by the European Aspergillus PCR Initiative for the testing of whole blood, serum, and plasma. While some comparison of the performance of Aspergillus PCR when testing these different sample types has been performed, no single study has evaluated all three using the recommended protocols. Standardized Aspergillus PCR was performed on 423 whole-blood pellets (WBP), 583 plasma samples, and 419 serum samples obtained from hematology patients according to the recommendations. This analysis formed a bicenter retrospective anonymous case-control study, with diagnosis according to the revised European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) consensus definitions (11 probable cases and 36 controls). Values for clinical performance using individual and combined samples were calculated. For all samples, PCR positivity was significantly associated with cases of IA (for plasma, P = 0.0019; for serum, P = 0.0049; and for WBP, P = 0.0089). Plasma PCR generated the highest sensitivity (91%); the sensitivities for serum and WBP PCR were 80% and 55%, respectively. The highest specificity was achieved when testing WBP (96%), which was significantly superior to the specificities achieved when testing serum (69%, P = 0.0238) and plasma (53%, P = 0.0002). No cases were PCR negative in all specimen types, and no controls were PCR positive in all specimens. This study confirms that Aspergillus PCR testing of plasma provides robust performance while utilizing commercial automated DNA extraction processes. Combining PCR testing of different blood fractions allows IA to be both confidently diagnosed and excluded. A requirement for multiple PCR-positive plasma samples provides similar diagnostic utility and is technically less demanding. Time to diagnosis may be enhanced by testing multiple contemporaneously obtained sample types. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Glycomics meets artificial intelligence - Potential of glycan analysis for identification of seropositive and seronegative rheumatoid arthritis patients revealed.

PubMed

Chocholova, Erika; Bertok, Tomas; Jane, Eduard; Lorencova, Lenka; Holazova, Alena; Belicka, Ludmila; Belicky, Stefan; Mislovicova, Danica; Vikartovska, Alica; Imrich, Richard; Kasak, Peter; Tkac, Jan

2018-06-01

In this study, one hundred serum samples from healthy people and patients with rheumatoid arthritis (RA) were analyzed. Standard immunoassays for detection of 10 different RA markers and analysis of glycan markers on antibodies in 10 different assay formats with several lectins were applied for each serum sample. A dataset containing 2000 data points was data mined using artificial neural networks (ANN). We identified key RA markers, which can discriminate between healthy people and seropositive RA patients (serum containing autoantibodies) with accuracy of 83.3%. Combination of RA markers with glycan analysis provided much better discrimination accuracy of 92.5%. Immunoassays completely failed to identify seronegative RA patients (serum not containing autoantibodies), while glycan analysis correctly identified 43.8% of these patients. Further, we revealed other critical parameters for successful glycan analysis such as type of a sample, format of analysis and orientation of captured antibodies for glycan analysis. Copyright © 2018 Elsevier B.V. All rights reserved.

Simultaneous detection of antibodies to five Actinobacillus pleuropneumoniae serovars using bead-based multiplex analysis.

PubMed

Berger, Sanne Schou; Lauritsen, Klara Tølbøll; Boas, Ulrik; Lind, Peter; Andresen, Lars Ole

2017-11-01

We developed and made a preliminary validation of a bead-based multiplexed immunoassay for simultaneous detection of porcine serum antibodies to Actinobacillus pleuropneumoniae serovars 1, 2, 6, 7, and 12. Magnetic fluorescent beads were coupled with A. pleuropneumoniae antigens and tested with a panel of serum samples from experimentally infected pigs and with serum samples from uninfected and naturally infected pigs. The multiplex assay was compared to in-house ELISAs and complement fixation (CF) tests, which have been used for decades as tools for herd classification in the Danish Specific Pathogen Free system. Assay specificities and sensitivities as well as the corresponding cutoff values were determined using receiver operating characteristic (ROC) curve analysis, and the A. pleuropneumoniae multiplex assay showed good correlation with the in-house ELISAs and CF tests with areas under ROC curves ≥ 0.988. Benefits of multiplexed assays compared to ELISAs and CF tests include reduced serum sample volumes needed for analysis, less labor, and shorter assay time.

Higher serum 25-hydroxyvitamin D concentrations are related to a reduced risk of depression.

PubMed

Jääskeläinen, Tuija; Knekt, Paul; Suvisaari, Jaana; Männistö, Satu; Partonen, Timo; Sääksjärvi, Katri; Kaartinen, Niina E; Kanerva, Noora; Lindfors, Olavi

2015-05-14

Vitamin D has been suggested to protect against depression, but epidemiological evidence is scarce. The present study investigated the relationship of serum 25-hydroxyvitamin D (25(OH)D) with the prevalence of depressive and anxiety disorders. The study population consisted of a representative sample of Finnish men and women aged 30-79 years from the Health 2000 Survey. The sample included 5371 individuals, of which 354 were diagnosed with depressive disorder and 222 with anxiety disorder. Serum 25(OH)D concentration was determined from frozen samples. In a cross-sectional study, a total of four indicators of depression and one indicator of anxiety were used as dependent variables. Serum 25(OH)D was the risk factor of interest, and logistic models used further included sociodemographic and lifestyle variables as well as indicators of metabolic health as confounding and/or effect-modifying factors. The population attributable fraction (PAF) was estimated. Individuals with higher serum 25(OH)D concentrations showed a reduced risk of depression. The relative odds between the highest and lowest quartiles was 0.65 (95% CI 0.46, 0.93; P for trend = 0.006) after adjustment for sociodemographic, lifestyle and metabolic factors. Higher serum 25(OH)D concentrations were associated with a lower prevalence of depressive disorder especially among men, younger, divorced and those who had an unhealthy lifestyle or suffered from the metabolic syndrome. The PAF was estimated to be 19% for depression when serum 25(OH)D concentration was at least 50 nmol/l. These results support the hypothesis that higher serum 25(OH)D concentrations protect against depression even after adjustment for a large number of sociodemographic, lifestyle and metabolic factors. Large-scale prospective studies are needed to confirm this finding.

Clinical utility of serum samples for human parechovirus type 3 infection in neonates and young infants: The 2014 epidemic in Japan.

PubMed

Aizawa, Yuta; Suzuki, Yuko; Watanabe, Kanako; Oishi, Tomohiro; Saitoh, Akihiko

2016-02-01

During the 2014 human parechovirus type 3 (HPeV3) epidemic in Niigata, Japan, this prospective observational study identified HPeV3 from 43/85 (51%) febrile young infants <4 months using PCR analysis of serum (n = 42) and/or cerebrospinal fluid (CSF) (n = 32) and genetic sequencing of the VP1 region of HPeV3. HPeV3-infected patients (median age, 32 days; range, 4-113 days) were diagnosed as having sepsis (79%), sepsis-like syndrome (19%), or encephalitis with septic shock (2%). Other than fever, mottled skin (67%) was significantly more frequent in HPeV3-infected patients than other virus-infected patients (P = 0.005). The rate of HPeV3 RNA detection in CSF without pleocytosis was high (88%; 28/32). Among the 32 patients whose serum and CSF samples were available, all patients were positive for serum PCR; however, 4 (12%) patients were negative for CSF PCR. Serum HPeV3 RNA level on admission was associated with younger age (P = 0.002), bad temper (P = 0.041), and grunting (P = 0.008). Among 6 patients with sequential data on serum HPeV3 RNA level, levels decreased rapidly without specific therapy. In conclusion, serum samples at disease onset are the most useful compared to CSF in detection of HPeV RNA and serum HPeV3 RNA level on admission was associated with important clinical manifestations in HPeV3-infected patients. Copyright © 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Brix refractometry in serum as a measure of failure of passive transfer compared to measured immunoglobulin G and total protein by refractometry in serum from dairy calves.

PubMed

Hernandez, D; Nydam, D V; Godden, S M; Bristol, L S; Kryzer, A; Ranum, J; Schaefer, D

2016-05-01

A series of trials were conducted to evaluate Brix refractometry (Brix %) for the assessment of failure of passive transfer (FPT) in dairy calves compared to: (1) serum IgG (reference standard) when measured by radial immunodiffusion (RID) or a turbidometric immunoassay (TIA), and (2) serum total protein refractometry (STP). For the serum samples tested with TIA, STP, and Brix % (n = 310; Holstein calves), the median concentrations were 21.3 g/L IgG, 58 g/L STP, and 9.2%, respectively. For the serum samples tested with RID, STP and Brix % (n = 112; Jersey calves), the mean concentrations were 38 g/L IgG, 68 g/L STP, and 10.2%, respectively. For samples tested with only Brix % and STP (n = 265; Holstein calves), median STP and Brix % were 50 g/L STP and 8.5%, respectively. Correlations between Brix % and RID, and between Brix % and TIA were equal (r = 0.79, respectively). Brix % and STP were positively correlated (r = 0.99). Brix % estimated serum IgG concentrations determined by TIA and RID (r(2) = 0.63, 0.62, respectively). When FPT was defined as serum IgG < 10 g/L, Brix % ≤ 8.5% showed optimal sensitivity (100%) and specificity (89.2%) to predict FPT. At the same IgG cut-point, an STP value of ≤ 52 g/L showed a similar sensitivity (100%) and specificity (80.4%) to predict FPT. Brix refractometry predicted successful transfer of passive immunity in dairy calves, but further evaluation as a diagnostic tool for the diagnosis of FPT is warranted. Copyright © 2016. Published by Elsevier Ltd.

Concentrations of Phthalate Metabolites in Milk, Urine, Saliva, and Serum of Lactating North Carolina Women

PubMed Central

Hines, Erin P.; Calafat, Antonia M.; Silva, Manori J.; Mendola, Pauline; Fenton, Suzanne E.

2009-01-01

Background Phthalates are ubiquitous in the environment, but concentrations in multiple media from breast-feeding U.S. women have not been evaluated. Objectives The objective of this study was to accurately measure and compare the concentrations of oxidative monoester phthalate metabolites in milk and surrogate fluids (serum, saliva, and urine) of 33 lactating North Carolina women. Methods We analyzed serum, saliva, urine, and milk for the oxidative phthalate metabolites mono(3-carboxypropyl) phthalate, mono(2-ethyl-5-carboxypentyl) phthalate (MECPP), mono(2-ethyl-5-hydroxyhexyl) phthalate, and mono(2-ethyl-5-oxohexyl) phthalate using isotope-dilution high-performance liquid chromatography tandem mass spectroscopy. Because only urine lacks esterases, we analyzed it for the hydrolytic phthalate monoesters. Results We detected phthalate metabolites in few milk (< 10%) and saliva samples. MECPP was detected in > 80% of serum samples, but other metabolites were less common (3–22%). Seven of the 10 urinary metabolites were detectable in ≥ 85% of samples. Monoethyl phthalate had the highest mean concentration in urine. Metabolite concentrations differed by body fluid (urine > serum > milk and saliva). Questionnaire data suggest that frequent nail polish use, immunoglobulin A, and fasting serum glucose and triglyceride levels were increased among women with higher concentrations of urinary and/or serum phthalate metabolites; motor vehicle age was inversely correlated with certain urinary phthalate concentrations. Conclusions Our data suggest that phthalate metabolites are most frequently detected in urine of lactating women and are less often detected in serum, milk, or saliva. Urinary phthalate concentrations reflect maternal exposure and do not represent the concentrations of oxidative metabolites in other body fluids, especially milk. PMID:19165392

Sustained high serum caspase-3 concentrations and mortality in septic patients.

PubMed

Lorente, L; Martín, M M; Pérez-Cejas, A; González-Rivero, A F; López, R O; Ferreres, J; Solé-Violán, J; Labarta, L; Díaz, C; Palmero, S; Jiménez, A

2018-02-01

Caspase-3 is the main executor of the apoptotic process. Higher serum caspase-3 concentrations in non-survivor compared to survivor septic patients have been found. The objectives of this work (with the increase of sample size to 308 patients, and the determination of serum caspase-3 concentrations also on days 4 and 8 of diagnosis of severe sepsis) were to know whether an association between serum caspase-3 concentrationss during the first week, degree of apoptosis, sepsis severity, and sepsis mortality exists. We collected serum samples of 308 patients with severe sepsis from eight intensive care units on days 1, 4 and 8 to measure concentrations of caspase-3 and caspase-cleaved cytokeratin (CCCK)-18 (to assess degree of apoptosis). End point was 30-day mortality. We found higher serum concentrations of caspase-3 and CCCK-18 in non-survivors compared to survivors on days 1 (p < 0.001), 4 (p < 0.001), and 8 (p < 0.001). We found an association between serum caspase-3 concentrations on days 1, 4 and 8 of severe sepsis diagnosis and serum CCCK-18 concentrations (p < 0.001), SOFA (p < 0.001), serum acid lactic concentrations (p < 0.001), and 30-day sepsis mortality (p < 0.001). The new findings of this work were that an association between serum caspase-3 concentrations during the first week, apoptosis degree, sepsis severity, and sepsis mortality exists.

Screening for chromosomal anomalies in the first trimester: does repeat maternal serum screening improve detection rates?

PubMed

Spencer, Kevin; Cuckle, Howard S

2002-10-01

To assess the within person biological variability of first trimester maternal serum biochemical markers of trisomy 21 across the 10-14 week gestational period. To evaluate whether repeat sampling and testing of free beta-hCG and PAPP-A during this period would result in an improved detection rate. Women presenting at the first trimester OSCAR clinic have blood collected prior to ultrasound dating and nuchal translucency measurement. All samples are analysed for free beta-hCG and PAPP-A before an accurate estimate of gestation is available. In 10% of cases the gestation is prior to the minimum time for NT measurement (11 weeks) and these women are rebooked for a repeat visit to the clinic at the appropriate time. Our fetal database was interrogated to obtain cases in which two maternal blood samples had been collected and analysed in the 10-14 week period. Using data from the marker correlations and statistical modelling, the impact of repeat testing on detection rate for trisomy 21 at a fixed 5% false positive rate, was assessed. 261 pairs of data were available for analysis collected over a 3 year period. The correlation between free beta-hCG in sample 1 and sample 2 was 0.890 and that for PAPP-A was 0.827. The average within person biological variation for free beta-hCG was 21% and 32% for PAPP-A. The increase in detection rate when using both sets of marker data was 3.5% when using serum biochemistry and maternal age, and 1.3% when using nuchal translucency, serum biochemistry and maternal age. Repeat sampling and testing of maternal serum biochemical markers is unlikely to substantially improve first trimester screening performance. Copyright 2002 John Wiley & Sons, Ltd.

An improved reference measurement procedure for triglycerides and total glycerides in human serum by isotope dilution gas chromatography-mass spectrometry.

PubMed

Chen, Yizhao; Liu, Qinde; Yong, Sharon; Teo, Hui Ling; Lee, Tong Kooi

2014-01-20

Triglycerides are widely tested in clinical laboratories using enzymatic methods for lipid profiling. As enzymatic methods can be affected by interferences from biological samples, this together with the non-specific nature of triglycerides measurement makes it necessary to verify the accuracy of the test results with a reference measurement procedure. Several such measurement procedures had been published. These procedures generally involved lengthy and laborious sample preparation steps. In this paper, an improved reference measurement procedure for triglycerides and total glycerides was reported which simplifies the sample preparation steps and greatly shortens the time taken. The procedure was based on isotope dilution gas chromatography-mass spectrometry (IDGC-MS)with tripalmitin as the calibration standard. Serum samples were first spiked with isotope-labeled tripalmitin. For the measurement of triglycerides, the serum samples were subjected to lipid extraction followed by separation of triglycerides from diglycerides and monoglycerides. Triglycerides were then hydrolyzed to glycerol, derivatized and injected into the GC–MS for quantification. For the measurement of total glycerides, the serum samples were hydrolyzed directly and derivatized before injection into the GC-MS for quantification. All measurement results showed good precision with CV <1%. A certified reference material (CRM) of lipids in frozen human serum was used to verify the accuracy of the measurement. The obtained values for both triglycerides and total glycerides were well within the certified ranges of the CRM, with deviation <0.4% from the certified values. The relative expanded uncertainties were also comparable with the uncertainties associated with the certified values of the CRM. The validated procedure was used in an External Quality Assessment (EQA) Program organized by our laboratory to establish the assigned values for triglycerides and total glycerides.

Development and validation of a liquid chromatography tandem mass spectrometry assay for the quantitation of a protein therapeutic in cynomolgus monkey serum.

PubMed

Zhao, Yue; Liu, Guowen; Angeles, Aida; Hamuro, Lora L; Trouba, Kevin J; Wang, Bonnie; Pillutla, Renuka C; DeSilva, Binodh S; Arnold, Mark E; Shen, Jim X

2015-04-15

We have developed and fully validated a fast and simple LC-MS/MS assay to quantitate a therapeutic protein BMS-A in cynomolgus monkey serum. Prior to trypsin digestion, a recently reported sample pretreatment method was applied to remove more than 95% of the total serum albumin and denature the proteins in the serum sample. The pretreatment procedure simplified the biological sample prior to digestion, improved digestion efficiency and reproducibility, and did not require reduction and alkylation. The denatured proteins were then digested with trypsin at 60 °C for 30 min and the tryptic peptides were chromatographically separated on an Acquity CSH column (2.1 mm × 50 mm, 1.7 μm) using gradient elution. One surrogate peptide was used for quantitation and another surrogate peptide was selected for confirmation. Two corresponding stable isotope labeled peptides were used to compensate variations during LC-MS detection. The linear analytical range of the assay was 0.50-500 μg/mL. The accuracy (%Dev) was within ± 5.4% and the total assay variation (%CV) was less than 12.0% for sample analysis. The validated method demonstrated good accuracy and precision and the application of the innovative albumin removal sample pretreatment method improved both assay sensitivity and robustness. The assay has been applied to a cynomolgus monkey toxicology study and the serum sample concentration data were in good agreement with data generated using a quantitative ligand-binding assay (LBA). The use of a confirmatory peptide, in addition to the quantitation peptide, ensured the integrity of the drug concentrations measured by the method. Copyright © 2015 Elsevier B.V. All rights reserved.

Clinical evaluation of a Mucorales-specific real-time PCR assay in tissue and serum samples.

PubMed

Springer, Jan; Lackner, Michaela; Ensinger, Christian; Risslegger, Brigitte; Morton, Charles Oliver; Nachbaur, David; Lass-Flörl, Cornelia; Einsele, Hermann; Heinz, Werner J; Loeffler, Juergen

2016-12-01

Molecular diagnostic assays can accelerate the diagnosis of fungal infections and subsequently improve patient outcomes. In particular, the detection of infections due to Mucorales is still challenging for laboratories and physicians. The aim of this study was to evaluate a probe-based Mucorales-specific real-time PCR assay (Muc18S) using tissue and serum samples from patients suffering from invasive mucormycosis (IMM). This assay can detect a broad range of clinically relevant Mucorales species and can be used to complement existing diagnostic tests or to screen high-risk patients. An advantage of the Muc18S assay is that it exclusively detects Mucorales species allowing the diagnosis of Mucorales DNA without sequencing within a few hours. In paraffin-embedded tissue samples this PCR-based method allowed rapid identification of Mucorales in comparison with standard methods and showed 91 % sensitivity in the IMM tissue samples. We also evaluated serum samples, an easily accessible material, from patients at risk from IMM. Mucorales DNA was detected in all patients with probable/proven IMM (100 %) and in 29 % of the possible cases. Detection of IMM in serum could enable an earlier diagnosis (up to 21 days) than current methods including tissue samples, which were gained mainly post-mortem. A screening strategy for high-risk patients, which would enable targeted treatment to improve patient outcomes, is therefore possible.

Acute phase protein haptoglobin as inflammatory marker in serum and synovial fluid in an equine model of arthritis.

PubMed

Barrachina, Laura; Remacha, Ana Rosa; Soler, Lourdes; García, Natalia; Romero, Antonio; Vázquez, Francisco José; Vitoria, Arantza; Álava, María Ángeles; Lamprave, Fermín; Rodellar, Clementina

2016-12-01

Acute phase proteins are useful inflammatory markers in horses. Haptoglobin (Hp) serum level is increased in horses undergoing different inflammatory processes, including arthritis. However, Hp concentration has not been assessed in inflammatory synovial fluid (SF). The aim of the present study was to investigate the Hp response in serum and SF in horses undergoing experimentally induced arthritis. For this purpose, serum and SF samples were collected from 12 animals before amphotericin B-induced arthritis was created (T0, healthy) and 15days after the lesion induction (T1, joint inflammation) and Hp was determined by single radial immunodiffusion. The Hp increase between T0 and T1 was significant in both serum and SF, and serum Hp concentration at T0 was significantly higher than in SF, but significant differences were not found at T1, indicating a higher Hp increase in SF. A significant positive correlation for Hp concentration between serum and SF samples was found. These results highlight the potential usefulness of Hp as inflammatory marker in horses, showing for the first time the increase of Hp in SF from joint inflammation in the horse. Copyright © 2016 Elsevier B.V. All rights reserved.

Correction of vitamin D deficiency in a cohort of newborn infants using daily 200 IU vitamin D supplementation.

PubMed

Onwuneme, C; Diya, B; Uduma, O; McCarthy, R A; Murphy, N; Kilbane, M T; McKenna, M J; Molloy, E J

2016-08-01

Although the role of vitamin D in the prevention of rickets has long been well established, controversies still exist on the ideal dose of vitamin D supplementation in infants. We assessed serum 25-hydroxyvitamin D (25OHD) status simultaneously in maternal and cord samples and the response to vitamin D3 supplementation in neonates. Serum 25OHD levels were evaluated from maternal, and umbilical cord samples from term normal pregnancies. Repeat 25OHD levels were assessed in neonates with 25OHD below 30 nmol/L following vitamin D3 200 IU daily after 6 weeks. Blood samples were taken including 57 cord samples and 16 follow-up neonatal samples. Maternal and cord serum 25OHD were 43 ± 21 and 29 ± 15 nmol/L, respectively. Infants with 25OHD < 30 nmol/L (19.8 ± 4.7 nmol/L) had a significant increase in serum 25OHD (63.3 ± 14.5 nmol/L) following vitamin D3 200 IU daily after 6 weeks. Healthy Irish infants born at term are at high risk of vitamin D deficiency, but vitamin D3 200 IU daily, rapidly corrects poor vitamin D status.

Exposure of Piglets to Enteroviruses Investigated by an Immunoassay Based on the EV-G1 VP4 Peptide.

PubMed

Benkahla, Mehdi A; Sane, Famara; Desailloud, Rachel; Hober, Didier

2016-01-01

The aim of this study was to investigate the exposure of piglets to enteroviruses-G (EV-G) through the presence of antibodies in their serum. Serum samples were obtained from the vena cava of 10 piglets at 9 weeks of age and again 39 days later (day 39). They were tested using an immunoassay based on the EV-G1 VP4 peptide, since VP4 is highly conserved among the four Enterovirus capsid proteins, and by using a seroneutralization assay. For each serum collected on day 39 the optical density was high compared to the value obtained in serum collected earlier (p = 0.002). However, the titers of anti-EV-G1 serum neutralizing activity were not different in paired samples (p > 0.999). The sequence alignment of the EV-G1 VP4 peptide, encompassing 50 amino acids, used in the immunoassay showed 88% homology with EV-G, suggesting that antibodies directed toward other EV-G than EV-G1 may be detected. An immunoassay based on EV-G1 VP4 can detect an increased level of EV-G antibodies in piglet serum samples. Further studies are needed to determine whether this immunoassay may be useful for diagnosis and/or epidemiological studies and to monitor EV-G infection in pigs to evaluate strategies aimed to prevent enterovirus infections. © 2016 S. Karger AG, Basel.

Polyfluoroalkyl Chemicals in the Serum and Milk of ...

EPA Pesticide Factsheets

Polyfluoroalkyl chemicals (PFCs) comprise a group of man-made organic compounds, some of which are persistent contaminants with developmental toxicity shown in laboratory animals. There is a paucity of human perinatal exposure data. The US EPA conducted a pilot study (Methods Advancement in Milk Analysis) including 34 breastfeeding women in North Carolina. Milk and serum samples were collected at 2-7 weeks and 3-4 months postpartum; 9 PFCs were assessed in milk and 7 in serum. Perfluorooctane sulfonic acid (PFOS), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), and perfluorohexane sulfonic acid (PFHxS) were found in nearly 100% of the serum samples. PFOS and PFOA were found at the highest concentrations. PFCs were below the limit of detection in most milk samples. Serum concentrations of PFOS, PFOA and PFHxS were lower (p<0.01) at the second visit compared to the first visit, and living in North Carolina 10 years or longer was related to elevated PFOS, PFOA and PFNA (p<0.03). These pilot data support the need to further explore perinatal PFC exposures and potentially related health effects, as planned in the upcoming National Children’s Study which provided the framework for this investigation. These data demonstrate very low transfer of perfluorinated chemicals (PFCs) to breast milk, even though PFCs are readily detected in the sera of lactating NC women. Our data correlation statistics suggest that there is an elevation of average serum PFCs i

Distribution of serum total homocysteine and its association with diabetes and cardiovascular risk factors of the insulin resistance syndrome in Mexican American men: the Third National Health and Nutrition Examination Survey.

PubMed

Gillum, Richard

2003-08-05

Few data have been published on the association of variables of the insulin resistance syndrome and serum total homocysteine (tHcy), a putative risk factor for cardiovascular morbidity, in representative samples of total populations or in Hispanic Americans. To describe the distributions of serum tHcy concentration and variables associated with insulin resistance in Mexican American men and to assess their association, data from a cross-sectional survey of a large national sample, the Third National Health and Nutrition Examination Survey were analyzed. Analyses were restricted to Mexican American men aged 40-74 years with data on glycated hemoglobin (%), body mass index (BMI), body fat distribution, HDL cholesterol, fasting serum insulin, serum triglycerides and serum tHcy concentrations. Cumulative distributions of serum tHcy shifted to the right with increasing age. Log serum tHcy was not associated with prevalence of diagnosed diabetes mellitus or glycated hemoglobin percent or other risk factors other than age. Log serum tHcy concentration showed borderline significant (p = 0.049) positive association with fasting serum insulin concentration independent of age and BMI, only in men aged 60-74. No consistent association of tHcy with diabetes prevalence or variables of the insulin resistance syndrome were found in Mexican American men aged 40-74 years. Further research is needed on the associations of serum tHcy concentration with insulin resistance and other components of the insulin resistance syndrome in persons of varying ethnicity.

Sensitive detection of C-reactive protein in serum by immunoprecipitation-microchip capillary gel electrophoresis.

PubMed

Herwig, Ela; Marchetti-Deschmann, Martina; Wenz, Christian; Rüfer, Andreas; Redl, Heinz; Bahrami, Soheyl; Allmaier, Günter

2015-06-01

Sepsis represents a significant cause of mortality in intensive care units. Early diagnosis of sepsis is essential to increase the survival rate of patients. Among others, C-reactive protein (CRP) is commonly used as a sepsis marker. In this work we introduce immune precipitation combined with microchip capillary gel electrophoresis (IP-MCGE) for the detection and quantification of CRP in serum samples. First high-abundance proteins (HSA, IgG) are removed from serum samples using affinity spin cartridges, and then the remaining proteins are labeled with a fluorescence dye and incubated with an anti-CRP antibody, and the antigen/antibody complex is precipitated with protein G-coated magnetic beads. After precipitation the complex is eluted from the beads and loaded onto the MCGE system. CRP could be reliably detected and quantified, with a detection limit of 25 ng/μl in serum samples and 126 pg/μl in matrix-free samples. The overall sensitivity (LOQ = 75 ng/μl, R(2) = 0.9668) of the method is lower than that of some specially developed methods (e.g., immune radiometric assay) but is comparable to those of clinically accepted ELISA methods. The straightforward sample preparation (not prone to mistakes), reduced sample and reagent volumes (including the antibodies), and high throughput (10 samples/3 h) are advantages and therefore IP-MCGE bears potential for point-of-care diagnosis. Copyright © 2015 Elsevier Inc. All rights reserved.

Serological evidence of hepatitis E virus infection in dromedary camels in Ethiopia.

PubMed

Li, Tian-Cheng; Yoshizaki, Sayaka; Zhou, Xianfeng; Sentsui, Hiroshi; Shirato, Kazuya; Matsuyama, Shutoku; Melaku, Simenew Keskes; Bazartseren, Boldbaatar; Takeda, Naokazu; Wakita, Takaji

2017-08-01

The genome of dromedary camel hepatitis E virus (DcHEV) has been detected in stool and serum samples from dromedary camels, but the sero-epidemiological information of DcHEV infection remains unclear. A total of 246 serum samples collected from dromedary camels (Camelus dromedarius) in Ethiopia, and 40 serum samples from Bactrian camels (Camelus ferus) in Mongolia were examined for the detection of anti-DcHEV IgG antibody by a newly developed enzyme-linked immunosorbent assay (ELISA) by using DcHEV-like particles (DcHEV-LPs) as the antigen. The results revealed that 55 of the 246 (22.4%) dromedary camels were positive for anti-DcHEV IgG, whereas all 40 samples from the Bactrian camels were negative for DcHEV IgG antibody. A total of 98 serum samples from dromedary camels, including 25 anti-DcHEV-IgG positive samples, were used for the detection of DcHEV RNA by reverse transcription-polymerase chain reaction (RT-PCR), however, no positive samples were identified. These results suggested that the DcHEV infection occurred in the dromedary camels in Ethiopia. Further studies are required to determine whether Bactrian camels are susceptible to DcHEV infection. In addition, not only DcHEV-LPs, but also virus-like particles (VLPs) delivered from G1, G3 and G5 HEV are likely applicable for the detection of the anti-DcHEV IgG antibody. Copyright © 2017 Elsevier B.V. All rights reserved.

Oral fluid samples used for PRRSV acclimatization program and sow performance monitoring in endemic PRRS-positive farms.

PubMed

Woonwong, Yonlayong; Kedkovid, Roongtham; Arunorat, Jirapat; Sirisereewan, Chaitawat; Nedumpun, Teerawut; Poonsuk, Korakrit; Panyasing, Yaowalak; Poolperm, Pariwat; Boonsoongnern, Alongkot; Thanawongnuwech, Roongroje

2018-02-01

An effective gilt acclimatization program is one of the most important management strategies for controlling porcine reproductive and respiratory syndrome virus (PRRSV) infection. Recently, oral fluid samples have been used as alternative diagnostic samples for various swine diseases. This study utilized oral fluids for PRRSV monitoring during the gilt acclimatization period in PRRSV endemic farms. The study was performed in two selected commercial breeding herds (farm A and farm B). PRRSV RNA and PRRSV-specific antibodies were monitored using oral fluid and serum samples. Sow performance parameters related to PRRSV infection were recorded and assessed. After PRRSV exposure during acclimatization, viral RNA was demonstrated in oral fluids from 1 to 10 weeks post-exposure (WPE). PRRSV RNA was detected in serum at 1 and 4 WPE in farm A and at 1, 4, 8, and 12 WPE in farm B. Prolonged viremia of gilts from farm B was possibly due to re-infection (within the herd) and later, reproductive problems were found in the breeding herd. The correlation of PRRSV RNA concentration in oral fluids and serum was evident. The S/P ratio values of PRRSV antibodies in oral fluid samples were higher and had similar patterns of antibody responses to the serum samples. The results suggest that the use of oral fluid samples for PRRSV monitoring during gilt acclimatization in endemic farms is effective, convenient, practical, and economical and would be most beneficial when used with other parameters.

Nipah Virus in Lyle's Flying Foxes, Cambodia

PubMed Central

Counor, Dorian; Ong, Sivuth; Faure, Caroline; Seng, Vansay; Molia, Sophie; Walston, Joe; Georges-Courbot, Marie Claude; Deubel, Vincent; Sarthou, Jean-Louis

2005-01-01

We conducted a survey in Cambodia in 2000 on henipavirus infection among several bat species, including flying foxes, and persons exposed to these animals. Among 1,072 bat serum samples tested by enzyme-linked immunosorbent assay, antibodies reactive to Nipah virus (NiV) antigen were detected only in Pteropus lylei species; Cynopterus sphinx, Hipposideros larvatus, Scotophilus kuhlii, Chaerephon plicata, Taphozous melanopogon, and T. theobaldi species were negative. Seroneutralization applied on a subset of 156 serum samples confirmed these results. None of the 8 human serum samples was NiV seropositive with the seroneutralization test. One virus isolate exhibiting cytopathic effect with syncytia was obtained from 769 urine samples collected at roosts of P. lylei specimens. Partial molecular characterization of this isolate demonstrated that it was closely related to NiV. These results strengthen the hypothesis that flying foxes could be the natural host of NiV. Surveillance of human cases should be implemented. PMID:16022778

Reproducibility of positive results for the detection of serum galactomannan by Platelia™ aspergillus EIA.

PubMed

Pedroza, Kelly C M C; de Matos, Sócrates B; de Moura, Daniel L; Oliveira, Mônica B B; Araújo, Marco A S; Nascimento, Roberto J M; Lima, Fernanda W M

2013-10-01

Galactomannan (GM) was recently included in consensus guidelines as an indirect mycological criterion for the diagnosis of invasive aspergillosis. Currently, there is an enzyme immunoassay available to detect GM in biological samples, the Platelia™ Aspergillus EIA. In this study, the reproducibility of positive results obtained using this assay was evaluated using serum samples from neutropenic patients. A trend toward lower values was observed, and 55 %(27/49) of positive results were negative after retesting. A low reproducibility of positive results for the detection of GM in serum was observed.

Prevalence of antibodies to Trypanosoma cruzi, Leishmania infantum, Encephalitozoon cuniculi, Sarcocystis neurona, and Neospora caninum in Capybara, Hydrochoerus hydrochaeris, from São Paulo State, Brazil.

PubMed

Valadas, Samantha; Gennari, Solange Maria; Yai, Lucia Eiko Oishi; Rosypal, Alexa C; Lindsay, David S

2010-06-01

Little is known about the importance of capybara, Hydrochoerus hydrochaeris, as reservoirs for parasites of zoonotic or veterinary importance. Sera from 63 capybaras, from 6 counties in the state of São Paulo, Brazil, were examined for antibodies to Trypanosoma cruzi, Leishmania infantum, Encephalitozoon cuniculi, Sarcocystis neurona, and Neospora caninum using an indirect immunofluorescent antibody test. Five (8%) of the 63 capybaras had antibodies to T. cruzi epimastigotes. None of the samples from capybara reacted positively with L. infantum promastigotes or with spores of E. cuniculi . Two (3%) of the serum samples were positive for antibodies to S. neurona merozoites, and 2 (3%) of the serum samples were positive for antibodies to N. caninum tachyzoites. A serum sample from 1 capybara was positive for antibodies to both T. cruzi and N. caninum. None of the remaining 62 samples reacted with more than 1 parasite.

Evaluation of three high abundance protein depletion kits for umbilical cord serum proteomics

PubMed Central

2011-01-01

Background High abundance protein depletion is a major challenge in the study of serum/plasma proteomics. Prior to this study, most commercially available kits for depletion of highly abundant proteins had only been tested and evaluated in adult serum/plasma, while the depletion efficiency on umbilical cord serum/plasma had not been clarified. Structural differences between some adult and fetal proteins (such as albumin) make it likely that depletion approaches for adult and umbilical cord serum/plasma will be variable. Therefore, the primary purposes of the present study are to investigate the efficiencies of several commonly-used commercial kits during high abundance protein depletion from umbilical cord serum and to determine which kit yields the most effective and reproducible results for further proteomics research on umbilical cord serum. Results The immunoaffinity based kits (PROTIA-Sigma and 5185-Agilent) displayed higher depletion efficiency than the immobilized dye based kit (PROTBA-Sigma) in umbilical cord serum samples. Both the PROTIA-Sigma and 5185-Agilent kit maintained high depletion efficiency when used three consecutive times. Depletion by the PROTIA-Sigma Kit improved 2DE gel quality by reducing smeared bands produced by the presence of high abundance proteins and increasing the intensity of other protein spots. During image analysis using the identical detection parameters, 411 ± 18 spots were detected in crude serum gels, while 757 ± 43 spots were detected in depleted serum gels. Eight spots unique to depleted serum gels were identified by MALDI- TOF/TOF MS, seven of which were low abundance proteins. Conclusions The immunoaffinity based kits exceeded the immobilized dye based kit in high abundance protein depletion of umbilical cord serum samples and dramatically improved 2DE gel quality for detection of trace biomarkers. PMID:21554704

Refractometer assessment of colostral and serum IgG and milk total solids concentrations in dairy cattle

PubMed Central

2014-01-01

Background Estimation of the quantity of colostral IgG or serum IgG absorbed following ingestion of colostrum by calves is essential for monitoring the effectiveness of colostrum feeding practices on dairy farms. Milk total solids concentrations determination is a critical part of quality assessment of nonsaleable whole milk prior to feeding to calves. To date, on-farm methods to assess colostral IgG, serum IgG or milk total solids concentrations have been performed separately with various instruments. The objective of this study was to evaluate the diagnostic performance of a single electronic, hand-held refractometer for assessing colostral and serum IgG concentrations and milk total solids in dairy cattle. Colostral IgG, serum IgG and milk total solids concentrations were determined by the refractometer. Corresponding analysis of colostral and serum IgG concentrations were determined by radial immunodiffusion (RID) while milk total solids were determined by spectrophotometry. Sensitivity and specificity of the refractometer for colostrum and serum samples were calculated as determined by RID. Sensitivity and specificity of the refractometer for milk samples was calculated as determined by spectrophotometry. Results The sensitivity of the refractometer was 1 for colostral IgG, serum IgG and milk total solids determinations. Specificity of the refractometer was 0.66, 0.24 and 0 for colostral IgG, serum IgG and milk total solids determinations, respectively. The refractometer underestimated colostral IgG, serum IgG and milk total solids concentrations compared to the concentrations determined by RID or spectrophotometry. Conclusions The refractometer was an acceptable, rapid, convenient on-farm method for determining colostral IgG and milk total solids. The refractometer was not an acceptable method for determination of serum IgG concentrations as it severely underestimated the serum IgG concentrations. PMID:25125217

[Sterility in drained serums; before their use in emergency wards and in non-hospital emergency use].

PubMed

Carrasco, Juan Antonio Péculo; Escorza, Juana Gómez; Sánchez, María Del Mar Casal; Bouza, Mónica Rodríguez; Sanchez, José Luis Avecilla; Ruiz, Hugo José Rodríguez

2008-01-01

The authors detail the characteristics for the sterility of drained serums placed in a mobile emergency service unit and they determine how much time they can maintain optimum conditions for the use of these serums by means of a controlled, random, triple blind clinical test in which the units studied were serums prepared by an infusion system and a three-way valve. Data were gathered during a 12 month study period. The authors designed six operating groups based on their exposure time and where they were kept. These samples totaled 672 units. The authors carried out 12 microbiological cultural analyses for each unit; these analyses were tested for bacteria and fungi, both aerobic and anaerobic. The authors contrasted their hypothesis with a multivariable logistic regression model used as the variable for sterility A value of p < 0.05 was considered to be significant. Results were obtained on 669 serums, 98.82% of samples. Colonization was found in 1.7% of the 8028 cultures analyzed; in only 2 cases was there a register greater than 10 UFC/ml. This colonization corresponded to 103 serums which were then classified as non-sterile. For sterility the logistic regression model did not show any significant association with the various operating groups, which were controlled by means of two possible variable methods. The authors did not find evidence that recently drained serums were different, from the point of view of sterility, than the serums exposed during 24, 48 or 72 hours but, as well as for the groups, the incidences of these serums tripled the risk of colonization. The authors concluded they could use serums prepared previously and these serums sterility were guaranteed to the same degree as recently drained serums.

Detection of autoantibodies against thyroid peroxidase in serum samples of hypothyroid dogs.

PubMed

Skopek, Evelyn; Patzl, Martina; Nachreiner, Raymond F

2006-05-01

To establish a sensitive test for the detection of autoantibodies against thyroid peroxidase (TPO) in canine serum samples. 365 serum samples from dogs with hypothyroidism as determined on the basis of serum concentrations of total and free triiodothyronine (T3), total and free thyroxine (T4), and thyroid-stimulating hormone, of which 195 (53%) had positive results for at least 1 of 3 thyroid autoantibodies (against thyroglobulin [Tg], T4, or T3) and serum samples from 28 healthy dogs (control samples). TPO was purified from canine thyroid glands by extraction with detergents, ultracentrifugation, and precipitation with ammonium sulfate. Screening for anti-TPO autoantibodies in canine sera was performed by use of an immunoblot assay. Thyroid extract containing TPO was separated electrophoretically, blotted, and probed with canine sera. Alkaline phosphatase-conjugated rabbit anti-dog IgG was used for detection of bound antibodies. TPO bands were observed at 110, 100, and 40 kd. Anti-TPO autoantibodies against the 40-kd fragment were detected in 33 (17%) sera of dogs with positive results for anti-Tg, anti-T4, or anti-T3 autoantibodies but not in sera of hypothyroid dogs without these autoantibodies or in sera of healthy dogs. The immunoblot assay was a sensitive and specific method for the detection of autoantibodies because it also provided information about the antigen. Anti-TPO autoantibodies were clearly detected in a fraction of hypothyroid dogs. The value of anti-TPO autoantibodies for use in early diagnosis of animals with thyroid gland diseases should be evaluated in additional studies.

Long-term in vitro reactivity for HLA antibodies and comparison of detection using serum vs. plasma

PubMed Central

Norris, Philip J.; Lee, Jar-How; Carrick, Danielle M.; Gottschall, Jerome L.; Lebedeva, Mila; de Castro, B.R.; Kleinman, Steven H.; Busch, Michael P.

2010-01-01

BACKGROUND HLA antibodies are a possible cause of transfusion-related acute lung injury (TRALI), and fluorescent bead assays are often used for antibody detection. Serum is the manufacturer’s recommended sample, but plasma may be easier to obtain for studies of HLA antibody prevalence and TRALI case investigations. STUDY DESIGN AND METHODS Specimens were obtained from 44 multiparous females positive for HLA antibodies by lymphocytotoxicity testing at least 13 years prior, and from 1,000 contemporary blood donors. Screening tests were performed using a Luminex-based assay. In addition to comparing results obtained with paired plasma and serum samples, the effects of storage at 4 °C for one week and of multiple freeze-thaw cycles were evaluated. RESULTS Of 42 evaluable subjects with HLA antibodies documented >13 years earlier, only 1 showed loss of detectable antibodies, with 39 (93%) positive in the screening assay for class I and 24 (57%) positive in the screening assay for HLA class II antibodies. In 968 evaluable contemporary donors, 291 screened positive for HLA class I and 206 for HLA class II antibodies using a low assay cut-off. Screening test concordance using paired plasma and serum samples was high, particularly for subjects with higher level antibodies. Refrigeration of samples for one week did not significantly affect assay results, while repeated freeze-thaw cycles caused a decrement in signal level. CONCLUSION Serum and plasma samples gave concordant results in the majority of cases, particularly for specimens with higher-level antibodies. High-level HLA antibodies were present in most individuals for over 13 years. PMID:18980615

SIMPLIFIED METHOD FOR EXTRACTING BOUND PESTICIDES FROM AVIAN SERUM

EPA Science Inventory

A simple solid-phase extraction (SPE) method was developed to extract organochlorine pesticides (OCs) and persistent organic pollutants (POPs) from avian serum. In this method, a 1-mL serum sample fortified with two levels of OCs or POPs was treated with 8M urea or 4M urea and 4...

Inositol and hepatic lipidosis. I. Effect of inositol supplementation and time from parturition on liver and serum lipids in dairy cattle.

PubMed

Gerloff, B J; Herdt, T H; Wells, W W; Liesman, J S; Emery, R S

1986-06-01

Percutaneous liver biopsies and blood samples were obtained from 80 multiparous dairy cows in nine Michigan herds. Biopsies and samples were obtained serially over the peripartum period. Thirty-nine cows received 17 g of supplemental myoinositol in the diet to test its use as a possible lipotropic substance and 41 received a placebo. Liver biopsies were assayed for triglyceride (TG) and total myoinositol content. Serum was assayed for dextran precipitable cholesterol and non-esterified fatty acids (NEFA). Inositol supplementation had no effect on any of the lipid variables. There was a significant herd effect on liver inositol, serum dextran precipitable cholesterol and NEFA concentrations. Serum NEFA and liver TG concentrations increased in the immediate postpartum period, while dextran precipitable cholesterol decreased. A significant herd X period interaction existed for liver TG and serum dextran precipitable cholesterol concentrations. Liver TG and serum NEFA concentrations were positively correlated. Excessive infiltration of bovine liver with lipid at calving appears to be an exaggerated manifestation of normal metabolic changes.

Comparison of high-pressure liquid chromatography (HPLC) and Griess reagent-spectroscopic methods for the measurement of nitrate in serum from healthy individuals in the Nordic countries.

PubMed

Larsen, Tine Lise; Nilsen, Valentina; Andersen, Dag Olav; Francis, George; Rustad, Pål; Mansoor, Mohammad Azam

2008-12-01

Bioavailability of NO can be estimated by measuring the concentration of nitrate (NO(3)) in serum. However, the methods used for the measurement NO(3) in plasma or serum show a great degree of variation. Therefore, we compared two analytical methods for the measurement of NO(3) in serum. The concentration of NO(3) in 600 serum samples collected from healthy individuals was determined by the HPLC and by the Griess reagent-spectroscopic method. The concentration of NO(3) in the samples was 29.4+/-16.1 micromol/L and 26.2+/-14.0 micromol/L (mean+/-SD) measured by HPLC and Griess reagent-spectroscopic method respectively (p<0.0001). We detected a significant correlation between the two methods (R=0.81, p<0.0001). A significant correlation between the two methods may suggest that either method can be used for the measurement of NO(3) in serum, however the Griess reagent-spectroscopic method measures lower concentrations of NO(3) than the HPLC method.

Utility of the Department of Defense Serum Repository in Assessing Deployment Exposure.

PubMed

Lushniak, Boris; Mallon, Col Timothy M; Gaydos, Joel C; Smith, David J

2016-08-01

This paper describes why the research project was conducted in terms of demonstrating the utility of the Department of Defense Serum Repository in addressing deployment environmental exposures. The history deployment exposure surveillance was reviewed and the rationale for developing validated biomarkers that were detected in sera in postdeployment samples and compared with nondeployed controls was described. The goal was to find validated biomarkers that are associated with both exposures and health outcomes. The articles in this supplement described novel serum biomarkers that were found to be associated with deployment exposures and weakly associated with some health outcomes. Future research must continue to validate the use of serum biomarkers when operational contingencies prevent the gold standard collection of real-time breathing zone samples in deployed service members.

A SIMPLE AND RAPID METHOD FOR THE SIMULTANEOUS DETERMINATION OF CALCIUM AND MAGNESIUM FROM THE SAME SAMPLE OF BLOOD SERUM

PubMed Central

Kovács, G. S.; Tárnoky, K. E.

1960-01-01

A simple and rapid procedure has been developed for the complexometric titration of serum calcium and magnesium using “plasmocorinth B” as indicator. Both determinations can be carried out from the same 0.5 ml. sample. The method is in good agreement with the established calcium and magnesium methods. PMID:14411396

Protein electrophoresis in cranes with presumed insect bite.

PubMed

Hartup, Barry K; Schroeder, Carrie A

2006-06-01

Serum protein electrophoresis (SPE) has emerged as a potentially valuable diagnostic tool in avian medicine; yet, there is limited information regarding SPE in cranes. Since 2000, 20 cases of unilateral periocular or facial soft tissue swelling, blepharitis, feather loss, and ocular or nasal discharge attributed to insect bite hypersensitivity were observed in cranes from a captive breeding center. SPE may be useful for evaluating these lesions. The aim of this study was to characterize the inflammatory response in cranes with hypersensitivity reactions using SPE. Serum samples from 7 cranes diagnosed with hypersensitivity reactions were submitted to a diagnostic laboratory for agarose gel electrophoresis. Results were compared to those in control serum samples obtained from the same cranes during routine physical examination, when they were clinically healthy. Total protein and a- and g-globulin concentrations were significantly increased and albumin/globulin ratios were significantly decreased in serum samples from cranes with hypersensitivity lesions compared with control samples. Using SPE, we documented changes in protein fraction concentrations in cranes with clinical signs of hypersensitivity. The increase in alpha- and gamma-globulin concentrations suggested inflammation and antigenic stimulation, consistent with a Type I hypersensitivity reaction.

Bayesian estimation of the sensitivity and specificity of individual fecal culture and Paralisa to detect Mycobacterium avium subspecies paratuberculosis infection in young farmed deer.

PubMed

Stringer, Lesley A; Jones, Geoff; Jewell, Chris P; Noble, Alasdair D; Heuer, Cord; Wilson, Peter R; Johnson, Wesley O

2013-11-01

A Bayesian latent class model was used to estimate the sensitivity and specificity of an immunoglobulin G1 serum enzyme-linked immunosorbent assay (Paralisa) and individual fecal culture to detect young deer infected with Mycobacterium avium subsp. paratuberculosis. Paired fecal and serum samples were collected, between July 2009 and April 2010, from 20 individual yearling (12-24-month-old) deer in each of 20 South Island and 18 North Island herds in New Zealand and subjected to culture and Paralisa, respectively. Two fecal samples and 16 serum samples from 356 North Island deer, and 55 fecal and 37 serum samples from 401 South Island deer, were positive. The estimate of individual fecal culture sensitivity was 77% (95% credible interval [CI] = 61-92%) with specificity of 99% (95% CI = 98-99.7%). The Paralisa sensitivity estimate was 19% (95% CI = 10-30%), with specificity of 94% (95% CI = 93-96%). All estimates were robust to variation of priors and assumptions tested in a sensitivity analysis. The data informs the use of the tests in determining infection status at the individual and herd level.

Improvement for identification of heterophile antibody interference and AFP hook effect in immunoassays with multiplex suspension bead array system.

PubMed

Wang, Yajie; Yu, Jinsheng; Ren, Yuan; Liu, Li; Li, Haowen; Guo, Anchen; Shi, Congning; Fang, Fang; Juehne, Twyla; Yao, Jianer; Yang, Enhuan; Zhou, Xuelei; Kang, Xixiong

2013-11-15

A variety of immunoassays including multiplex suspension bead array have been developed for tumor marker detections; however, these assays could be compromised in their sensitivity and specificity by well-known heterophile antibody interference and hook effect. Using Luminex® multiplex suspension bead arrays, we modified protocols with two newly-developed solutions that can identify heterophile antibody interference and AFP hook effect. Effectiveness of the two solutions was assessed in serum samples from patients. Concentrations of 9 tumor markers in heterophile antibody positive samples assayed with Solution A, containing murine monoclonal antibodies and mouse serum, were significantly reduced when compared with those false high signals assayed without Solution A (all p<0.01). With incorporation of Solution H (fluorescent beads linked with AFP antigen), a new strategy for identification of AFP hook effect was established, and with this strategy AFP hook effect was identified effectively in serum samples with very high levels of AFP. Two proprietary solutions improve the identification of heterophile antibody interference and AFP hook effect. With these solutions, multiplex suspension bead arrays provide more reliable testing results in tumor marker detection where complex clinical serum samples are used. © 2013.

Bayesian validation of a serum and milk ELISA for antibodies against Mycobacterium avium subspecies paratuberculosis in Greek dairy goats across lactation.

PubMed

Angelidou, E; Kostoulas, P; Leontides, L

2014-02-01

We validated a commercial (Idexx Pourquier, Montpellier, France) serum and milk indirect ELISA that detects antibodies against Mycobacterium avium ssp. paratuberculosis (MAP) in Greek dairy goats. Each goat was sampled 4 times, starting from kidding and covering early, mid, and late lactation. A total of 1,268 paired milk (or colostrum) and serum samples were collected during the 7-mo lactation period. Bayesian latent class models, which allow for the continuous interpretation of test results, were used to derive the distribution of the serum and milk ELISA response for healthy and MAP-infected individuals at each lactation stage. Both serum and milk ELISA, in all lactation stages, had average and similar overall discriminatory ability as measured by the area under the curve (AUC). For each test, the smallest overlap between the distribution of the healthy and MAP-infected does was in late lactation. At this stage, the AUC was 0.89 (95% credible interval: 0.70; 0.98) and 0.92 (0.74; 0.99) for the milk and serum ELISA, respectively. Both tests had comparable sensitivities and specificities at the recommended cutoffs across lactation. Lowering the cutoffs led to an increase in sensitivity without serious loss in specificity. In conclusion, the milk ELISA was as accurate as the serum ELISA. Therefore, it could serve as the diagnostic tool of choice, especially during the implementation of MAP control programs that require frequent testing, because milk sampling is a noninvasive, rapid, and easy process. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Blood serum concentrations of selenium in female llamas (Lama glama) in relationship to feeding practices, region of United States, reproductive stage, and health of offspring.

PubMed

Herdt, T H

1995-02-01

Serum Se and vitamin E concentrations were determined twice in each of 35 female llamas from a high-Se region and 96 female llamas from a low-Se region of the United States. The first determination was taken at a random time during gestation and the second shortly after parturition. At the time of postpartum sampling, a sample was also taken from the cria. Dietary information was collected from each llama owner and a subjective estimate of strength and vigor was assigned by the owner to each cria studied. The mean blood serum concentrations of Se in llamas during gestation and at parturition were 213 and 203 ng/mL, respectively. The mean serum Se concentration was 113 ng/mL in neonatal cria. Among herds, serum Se means varied for the cria (P < .05) and at both sampling times in the females (P < .01). In adult females, mean serum Se was actually higher during gestation (P = .06) and at parturition (P < .01) for the low-Se region than for the high-Se region, but this effect was removed by covariate adjustment for intake of supplemental Se. Serum Se in females was correlated (P < .05) to supplemental Se consumption. Vigor and (or) viability of the cria were not affected by the Se status of the dams (P > .3) or cria (P > .2). However, there were very few weak or stillborn cria in this study.(ABSTRACT TRUNCATED AT 250 WORDS)

Season and location effects on serum and liver mineral concentrations of Senepol cattle on St Croix, Virgin Islands.

PubMed

Wildeus, S; McDowell, L R; Fugle, J R

1992-11-01

Serum and liver concentrations of selected macro- and trace minerals were determined in Senepol cattle at 8 sites (4 each in a high and low rainfall region) during the dry and wet season on St Croix. At each site an average of 15 mature, lactating cows, grazing native grass/legume pastures without supplementation were blood sampled each season. Liver samples were collected (n = 51) at slaughter from mature animals originating from the same sites. A preliminary analysis indicated no differences in serum mineral concentrations between mature lactating cows and growing heifers. There were differences between sites for serum magnesium (Mg) (P < 0.001), copper (Cu) (P < 0.05) selenium (Se) (P < 0.001) and zinc (Zn) (P < 0.01) in the dry season, and for Cu (P < 0.01), iron (Fe) (P < 0.001) and Zn (P < 0.01) in the wet season. Higher (P < 0.001) serum concentrations of Mg, Cu, Fe and Zn were observed in the dry season, while Se was higher (P < 0.01) in the wet season. Liver concentrations of Cu and Fe were lower (P < 0.01) and liver molybdenum (Mo) (P < 0.001) and Se (P < 0.05) higher during the dry season. The seasonal differences in serum Cu, Se and Zn concentrations have not been observed in other studies in the Central American region. More than 50% of serum samples were deficient in phosphorus (P) regardless of season, and in Cu and Zn during the wet season. Mineral supplementation should be considered.

Multicenter Comparison of Serum and Whole-Blood Specimens for Detection of Aspergillus DNA in High-Risk Hematological Patients

PubMed Central

Springer, Jan; Morton, C. O.; Perry, Michael; Heinz, Werner J.; Paholcsek, Melinda; Alzheimer, Mona; Rogers, T. R.; Barnes, Rosemary A.; Einsele, Hermann; White, P. Lewis

2013-01-01

Samples from patients at high risk for invasive aspergillosis (IA) were prospectively collected and analyzed for the presence of molecular markers of fungal infection. Serum specimens were screened for galactomannan and Aspergillus DNA, and whole-blood specimens were screened only for Aspergillus DNA. Fungal infections were categorized according to the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group, National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) criteria. Forty-seven cases (proven and probable IA) and 31 controls (no evidence of IA) were selected retrospectively for this case-control study, comprising 803 samples, in order to determine the performance of whole-blood PCR, serum PCR, and serum galactomannan testing. Although no single assay was able to detect every case of IA, a combination of different assays provided the best performance. There was no significant difference between the use of whole-blood and serum specimens for PCR-based diagnosis of IA, but there was a trend for whole blood to be more sensitive (85% versus 79%) and to yield an earlier positive result (36 days versus 15 days) than for serum. However, DNA extraction from serum specimens is easier and faster than that from whole-blood specimens, and it allows the same specimen to be used for both galactomannan and PCR assays. In conclusion, the appropriate sample type for DNA extraction should be determined by the local requirements and the technical platforms available at each individual center. A combination of biomarker tests offered the best diagnostic utility for detecting IA. PMID:23426930

First Detection of Antibodies Against African Swine Fever Virus in Faeces Samples.

PubMed

Nieto-Pelegrín, E; Rivera-Arroyo, B; Sánchez-Vizcaíno, J M

2015-12-01

African swine fever (ASF) is a viral, highly lethal haemorrhagic disease of swine with no available vaccine or effective treatment. Introduction of ASF into a country triggers immediate restriction measures that cause significant economic losses and threatens spread to neighbouring countries. Wild boar populations have been recently assigned an essential role in the spread of African swine fever virus (ASFV) to European countries. Therefore, effective surveillance and monitoring of wild boar populations is required, but sampling wild boar is logistically challenging and expensive. This study assessed the feasibility of detecting antibodies against ASFV in faeces for later implementation in surveillance and control programmes. Two groups of pigs were experimentally infected with an attenuated ASFV isolate Ken05, and blood, oral fluid and faecal samples were tested for the presence of viral DNA using quantitative real-time polymerase chain reaction (qPCR) to monitor infection progress. Faecal samples were analysed using two indirect enzyme-linked immunosorbent assays (ELISAs) based on semipurified viral protein (vp) 72 or purified recombinant vp30 expressed in mammalian cells. Faecal samples from 9 of 10 pigs with non-haemorrhagic diarrhoea tested positive for antibodies against ASFV using the two ELISA tests that showed a positive correlation. The serum sample results from the two indirect ELISAs were compared against results from the reference ELISA technique and the immunoperoxidase test. Our findings indicate the feasibility of faecal sampling for detecting anti-ASFV antibodies, which may provide a practical non-invasive alternative for sampling wild boar populations. In conclusion, the application of these ELISA tests to faecal field samples could be particularly useful to screen for the presence of ASF in field conditions. © 2015 Blackwell Verlag GmbH.

Countercurrent distribution of biological cells

NASA Technical Reports Server (NTRS)

Brooks, D. E.

1979-01-01

A neutral polymer phase system consisting of 7.5 percent dextran 40/4.5 percent PEG 6, 0.11 M Na phosphate, 5 percent fetal bovine serum (FBS), pH 7.5, was developed which has a high phase droplet electrophoretic mobility and retains cell viability over many hours. In this and related systems, the drop mobility was a linear function of drop size, at least in the range 4-30 micron diameter. Applications of and electric field of 4.5 v/cm to a system containing 10 percent v/v bottom phase cleared the system more than two orders of magnitude faster than in the absence of the field. At higher bottom phase concentrations a secondary phenomenon intervened in the field driven separations which resulted in an increase in turbidity after clearing had commenced. The increase was associated with a dilution of the phase system in the chamber. The effect depended on the presence of the electric field. It may be due to electroosmotic flow of buffer through the Amicon membranes into the sample chamber and flow of phase system out into the rinse stream. Strategies to eliminate this problem are proposed.

Ultrasensitive Hybridization-Based ELISA Method for the Determination of Phosphorodiamidate Morpholino Oligonucleotides in Biological samples.

PubMed

Burki, Umar; Straub, Volker

2017-01-01

Determining the concentration of oligonucleotide in biological samples such as tissue lysate and serum is essential for determining the biodistribution and pharmacokinetic profile, respectively. ELISA-based assays have shown far greater sensitivities compared to other methods such as HPLC and LC/MS. Here, we describe a novel ultrasensitive hybridization-based ELISA method for quantitating morpholino oligonucleotides in mouse tissue lysate and serum samples. The assay has a linear detection range of 5-250 pM (R2 > 0.99).

Micro-bead injection spectroscopy for label-free automated determination of immunoglobulin G in human serum.

PubMed

Ramos, Inês I; Magalhães, Luís M; Barreiros, Luisa; Reis, Salette; Lima, José L F C; Segundo, Marcela A

2018-01-01

Immunoglobulin G (IgG) represents the major fraction of antibodies in healthy adult human serum, and deviations from physiological levels are a generic marker of disease corresponding to different pathologies. Therefore, screening methods for IgG evaluation are a valuable aid to diagnostics. The present work proposes a rapid, automatic, and miniaturized method based on UV-vis micro-bead injection spectroscopy (μ-BIS) for the real-time determination of human serum IgG with label-free detection. Relying on attachment of IgG in rec-protein G immobilized in Sepharose 4B, a bioaffinity column is automatically assembled, where IgG is selectively retained and determined by on-column optical density measurement. A "dilution-and-shoot" approach (50 to 200 times) was implemented without further sample treatment because interferences were flushed out of the column upon sample loading, with minimization of carryover and cross-contamination by automatically discarding the sorbent (0.2 mg) after each determination. No interference from human serum albumin at 60 mg mL -1 in undiluted sample was found. The method allowed IgG determination in the range 100-300 μg mL -1 (corresponding to 5.0-60 mg mL -1 in undiluted samples), with a detection limit of 33 μg mL -1 (1.7 mg mL -1 for samples, dilution factor of 50). RSD values were < 9.4 and < 11.7%, for intra and inter-assay precision, respectively, while recovery values for human serum spiked with IgG at high pathological levels were 97.8-101.4%. Comparison to commercial ELISA kit showed no significant difference for tested samples (n = 8). Moreover, time-to-result decreased from several hours to < 5 min and analysis cost decreased 10 times, showing the potential of the proposed approach as a point-of-care method. Graphical abstract Micro-Bead Injection Spectroscopy method for real time, automated and label-free determination of total serum human Immunoglobulin G (IgG). The method was designed for Lab-on-Valve (LOV) platforms using a miniaturised protein G bioaffinity separative approach. IgG are separated from serum matrix components upon quantification with low non-specific binding in less than 5 min.

Assessment of serum antibody titers against canine distemper virus, canine adenovirus type II, and canine parvovirus in Alaskan sled dogs before and after a long-distance race.

PubMed

Banse, Heidi E; McKenzie, Erica C; Nelson, Stuart; Hinchcliff, Kenneth W

2008-06-01

To determine serum antibody titers against canine distemper virus (CDV), canine adenovirus type II (CAV-2), and canine parvovirus (CPV) in trained sled dogs prior to and after completion of a long-distance race. Prospective cohort study. 195 Alaskan sled dogs (from 18 kennels) that participated in the 2006 Iditarod Trail Race. All 1,323 dogs participating in the race had been vaccinated against the 3 viruses at 19 to 286 days prior to initial blood sample collection (obtained within the month preceding the race). Within 12 hours of race completion, blood samples were collected from 195 dogs (convenience sample) and matched with each dog's prerace sample. Serum antibody titers (90% confidence intervals [CIs]) were determined via serum neutralization assays. After racing, geometric mean titers against CDV and CPV were significantly higher (2,495 [90% CI, 321 to 16,384] and 6,323 [90% CI, 512 to 32,768], respectively) than prerace values (82 [90% CI, 11 to 362] and 166 [90% CI, 32 to 1,024], respectively). Sixty-one of 194 (31.4%) dogs had > or = 4-fold increases in anti-CPV antibody titers after racing. Prerace serum antibody titers against CDV, CPV, and CAV-2 varied significantly by sled team but were not associated with time since vaccination. Postrace increases in serum anti-CDV and anti-CPV antibody titer might reflect exposure of dogs to these agents immediately before or during racing. Dogs had no clinical signs of CDV-, CAV-2-, or CPV-associated disease; therefore, the clinical importance of these titer changes is uncertain.

High Concentrations of Angiopoietin-Like Protein 4 Detected in Serum from Patients with Rheumatoid Arthritis Can Be Explained by Non-Specific Antibody Reactivity.

PubMed

Makoveichuk, Elena; Ruge, Toralph; Nilsson, Solveig; Södergren, Anna; Olivecrona, Gunilla

2017-01-01

Angiopoietin-like protein 4 (ANGPTL4) is suggested to be a master regulator of plasma triglyceride metabolism. Our aim was to study whether the previously reported high levels of ANGPTL4 detected in serum from patients with rheumatoid arthritis (RA) by ELISA was due to any specific molecular form of this protein (oligomers, monomers or fragments). ANGPTL4 levels were first determined in serum from 68 RA patients and 43 age and sex matched control subjects and the mean values differed by a factor of 5.0. Then, ANGPTL4 was analyzed after size exclusion chromatography (SEC) of serum samples. With serum from one of the RA patients with high levels of ANGPTL4, the dominant reactivity was found in fractions corresponding to high-molecular weight proteins. In addition, a minor peak of reactivity eluting late from the column was found both in the patient and in controls. By the use of HeteroBlock®, and by careful selection of antibodies, we documented non-specific reactions for ANGPTL4 in 39% of samples from the RA patients, most likely due to cross-reactivity of the antibodies with rheumatoid factor (RF). The corresponding figure for control subjects was 6.3%. After corrections for non-specific reactions, the mean level of ANGPTL4 in serum from RA patients was still significantly higher than in control individuals (mean levels were 101±62 and 67±39 ng/ml respectively, P = 0.02). We re-analyzed samples from our previously published studies on ANGPL4 levels in patients on hemodialysis and patients with diabetes type 2. These samples did not show false positive reactions. The levels of ANGPTL4 were comparable to those detected previously.

Cure of tuberculosis despite serum concentrations of antituberculosis drugs below published reference ranges.

PubMed

Meloni, Monica; Corti, Natascia; Müller, Daniel; Henning, Lars; Gutteck, Ursula; von Braun, Amrei; Weber, Rainer; Fehr, Jan

2015-01-01

Therapeutic target serum concentrations of first-line antituberculosis drugs have not been well defined in clinical studies in tuberculosis (TB) patients. We retrospectively investigated the estimated maximum serum concentrations (eC max) of antituberculosis drugs and clinical outcome of TB patients with therapeutic drug monitoring performed between 2010-2012 at our institution, and follow-up until March 2014. The eC max was defined as the highest serum concentration during a sampling period (2, 4 and 6 hours after drug ingestion). We compared the results with published eC max values, and categorised them as either "within reference range", "low eC max", or "very low eC max".Low/very low eC max-levels were defined as follows: isoniazid 2-3/<2 mg/l, rifampicin 4-8/<4 mg/l, rifabutin 0.2-0.3/<0.2 mg/l, ethambutol 1-2/<0.1 mg/l and pyrazinamide <20 mg/l. Concentrations of antituberculosis drugs in 175 serum samples of 17 patients with TB were analysed. In 12 (71%) patients, multiple therapeutic drug monitoring samples were collected over time, in 5 (29%) patients only one sample was available for therapeutic drug monitoring. Overall, 94% of all patients had at least one low antituberculosis drug concentration. Overall, 64% of all eC max levels were classified as "low" or "very low". The eC max was below the relevant reference range in 80% of isoniazid, 95% of rifampicin, 30% of pyrazinamide, and 30% of ethambutol measurements. All but one patient were cured of tuberculosis. Although many antituberculosis drug serum concentrations were below the widely used reference ranges, 16 of 17 patients were cured of tuberculosis. These results challenge the use of the published reference ranges for therapeutic drug monitoring.

Analysis of free drug fractions in serum by ultrafast affinity extraction and two-dimensional affinity chromatography using α1-acid glycoprotein microcolumns.

PubMed

Bi, Cong; Zheng, Xiwei; Hage, David S

2016-02-05

In the circulatory system, many drugs are reversibly bound to serum proteins such as human serum albumin (HSA) and alpha1-acid glycoprotein (AGP), resulting in both free and protein-bound fractions for these drugs. This report examined the use of microcolumns containing immobilized AGP for the measurement of free drug fractions by ultrafast affinity extraction and a two-dimensional affinity system. Several drugs known to bind AGP were used as models to develop and evaluate this approach. Factors considered during the creation of this method included the retention of the drugs on the microcolumns, the injection flow rate, the microcolumn size, and the times at which a second AGP column was placed on-line with the microcolumn. The final system had residence times of only 110-830ms during sample passage through the AGP microcolumns and allowed free drug fractions to be determined within 10-20min when using only 3-10μL of sample per injection. This method was used to measure the free fractions of the model drugs at typical therapeutic levels in serum, giving good agreement with the results obtained by ultrafiltration. This approach was also used to estimate the binding constants for each drug with AGP in serum, even for drugs that had significant interactions with both AGP and HSA in such samples. These results indicated that AGP microcolumns could be used with ultrafast affinity extraction to measure free drug fractions in a label-free manner and to study the binding of drugs with AGP in complex samples such as serum. Copyright © 2015 Elsevier B.V. All rights reserved.

Serum neuron-specific enolase levels from the same patients differ between laboratories: assessment of a prospective post-cardiac arrest cohort.

PubMed

Mlynash, Michael; Buckwalter, Marion S; Okada, Ami; Caulfield, Anna Finley; Venkatasubramanian, Chitra; Eyngorn, Irina; Verbeek, Marcel M; Wijman, Christine A C

2013-10-01

In comatose post-cardiac arrest patients, a serum neuron-specific enolase (NSE) level of >33 μg/L within 72 h was identified as a reliable marker for poor outcome in a large Dutch study (PROPAC), and this level was subsequently adopted in an American Academy of Neurology practice parameter. Later studies reported that NSE >33 μg/L is not a reliable predictor of poor prognosis. To test whether different clinical laboratories contribute to this variability, we compared NSE levels from the laboratory used in the PROPAC study (DLM-Nijmegen) with those of our hospital's laboratory (ARUP) using paired blood samples. We prospectively enrolled cardiac arrest patients who remained comatose after resuscitation. During the first 3 days, paired blood samples for serum NSE were drawn at a median of 10 min apart. After standard preparation for each lab, one sample was sent to ARUP laboratories and the other to DLM-Nijmegen. Fifty-four paired serum samples from 33 patients were included. Although the serum NSE measurements correlated well between laboratories (R = 0.91), the results from ARUP were approximately 30% lower than those from DLM-Nijmegen. Therapeutic hypothermia did not affect this relationship. Two patients had favorable outcomes after hypothermia despite NSE levels measured by DLM-Nijmegen as >33 μg/L. Absolute serum NSE levels of comatose cardiac arrest patients differ between laboratories. Any specific absolute cut-off levels proposed to prognosticate poor outcome should not be used without detailed data on how neurologic outcomes correspond to a particular laboratory's method, and even then only in conjunction with other prognostic variables.

Serum biochemical, blood gas and antioxidant status in search and rescue dogs before and after simulated fieldwork.

PubMed

Spoo, J W; Zoran, D L; Downey, R L; Bischoff, K; Wakshlag, J J

2015-10-01

The aim of the study was to assess the physiological and antioxidant status before and after a 4 h search and rescue field exercise, with handlers, under warm-weather conditions performing activities compared to a control group of similarly trained dogs at rest. Serum chemistry demonstrated a decrease in serum sodium (Na) and potassium (K) in both exercising and control groups, however only Na was decreased significantly (P < 0.05) after exercise and hematocrits (HCTs) remained unchanged. The exercise group demonstrated significantly decreased serum phosphorus (P) and magnesium (Mg) compared to pre-exercise values, as did the control group (P < 0.025). There was also a significant increase in creatinine kinase concentrations in the exercise groups (P < 0.025). Serum non-esterified fatty acids were increased only in the exercise group after exercise, suggesting fat mobilization to produce energy. The mean total serum antioxidant potential in searching dogs was no different from the pre- and post-exercise values in the control dogs. Serum vitamin E concentrations did not differ between the two groups, with a decreasing trend in both groups. There was a modest decrease in serum uric acid in the control group, while there was a significant rise after exercise in the searching group (P < 0.01). Multiple changes in serum chemistry, HCTs and blood gases were documented in this study, and were similar to those observed after other endurance activities. The lack of hemoconcentration in this field search exercise suggested that even in extreme environmental conditions (heat and humidity), dogs with access to water do not experience significant dehydration or diminished antioxidant status. Copyright © 2015. Published by Elsevier Ltd.

Comparative analysis of EV isolation procedures for miRNAs detection in serum samples.

PubMed

Andreu, Zoraida; Rivas, Eva; Sanguino-Pascual, Aitana; Lamana, Amalia; Marazuela, Mónica; González-Alvaro, Isidoro; Sánchez-Madrid, Francisco; de la Fuente, Hortensia; Yáñez-Mó, María

2016-01-01

Extracellular vesicles (EVs) are emerging as potent non-invasive biomarkers. However, current methodologies are time consuming and difficult to translate to clinical practice. To analyse EV-encapsulated circulating miRNA, we searched for a quick, easy and economic method to enrich frozen human serum samples for EV. We compared the efficiency of several protocols and commercial kits to isolate EVs. Different methods based on precipitation, columns or filter systems were tested and compared with ultracentrifugation, which is the most classical protocol to isolate EVs. EV samples were assessed for purity and quantity by nanoparticle tracking analysis and western blot or cytometry against major EV protein markers. For biomarker validation, levels of a set of miRNAs were determined in EV fractions and compared with their levels in total serum. EVs isolated with precipitation-based methods were enriched for a subgroup of miRNAs that corresponded to miRNAs described to be encapsulated into EVs (miR-126, miR-30c and miR-143), while the detection of miR-21, miR-16-5p and miR-19a was very low compared with total serum. Our results point to precipitation using polyethylene glycol (PEG) as a suitable method for an easy and cheap enrichment of serum EVs for miRNA analyses. The overall performance of PEG was very similar, or better than other commercial precipitating reagents, in both protein and miRNA yield, but in comparison to them PEG is much cheaper. Other methods presented poorer results, mostly when assessing miRNA by qPCR analyses. Using PEG precipitation in a longitudinal study with human samples, we demonstrated that miRNA could be assessed in frozen samples up to 8 years of storage. We report a method based on a cut-off value of mean of fold EV detection versus serum that provides an estimate of the degree of encapsulation of a given miRNA.

Antibody responses to botulinum neurotoxin type A of toxin-treated spastic equinus children with cerebral palsy: A randomized clinical trial comparing two injection schedules.

PubMed

Oshima, Minako; Deitiker, Philip; Hastings-Ison, Tandy; Aoki, K Roger; Graham, H Kerr; Atassi, M Zouhair

2017-05-15

We have conducted a 26-month-long comparative study involving young patients (2-6years old) with a clinical diagnosis of spastic equinus secondary to cerebral palsy who have been treated with BoNT/A (BOTOX®, Allergan) tri-annually or annually. Serum samples were obtained to determine the presence or absence of blocking antibodies (Abs) by a mouse protection assay (MPA) and levels of anti-BoNT/A Abs by radioimmune assay (RIA). HLA DQ alleles were typed using blood samples to determine the possible association of certain HLA type(s) with the disease or with the Ab status. Blocking Abs were detected in only two out of 18 serum samples of the tri-annual group, but none were found in 20 samples of the annual group. The MPA-positive serum samples gave in RIA significantly higher anti-BoNT/A Ab-binding levels than the MPA-negative samples. On the other hand, when two MPA-positive sample data were excluded, serum samples from tri-annual and annual groups showed similar anti-BoNT/A Ab levels. Linkage of the disorder with a particular HLA DQA1 and DQB1 allele types was not observed due to the small sample size. However, by combining results with other studies on BoNT/A-treated Caucasian patients with cervical dystonia (CD), we found that, among Caucasian patients treated with BoNT/A, DQA1*01:02 and DQB1*06:04 were higher in Ab-positive than in Ab-negative patients. The genetic linkage was on the threshold of corrected significance. Copyright © 2017. Published by Elsevier B.V.

Quantification of a biomarker of acetaminophen protein adducts in human serum by high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry: clinical and animal model applications.

PubMed

Cook, Sarah F; King, Amber D; Chang, Yan; Murray, Gordon J; Norris, Hye-Ryun K; Dart, Richard C; Green, Jody L; Curry, Steven C; Rollins, Douglas E; Wilkins, Diana G

2015-03-15

The aims of this study were to develop, validate, and apply a high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) method for quantification of protein-derived 3-(cystein-S-yl)-acetaminophen (APAP-Cys) in human serum. Formation of acetaminophen (APAP) protein adducts is thought to be a critical, early event in the development of APAP-induced hepatotoxicity, and quantification of these protein adducts in human serum represents a valuable tool for assessment of APAP exposure, metabolism, and toxicity. In the reported procedure, serum samples were first dialyzed or passed through gel filtration columns to remove APAP-Cys not covalently bound to proteins. Serum eluates were then subjected to enzymatic protease digestion to liberate protein-bound APAP-Cys. Norbuprenorphine-D3 was utilized as an internal standard (IS). APAP-Cys and IS were recovered from digested serum by protein precipitation with acetonitrile, and sample extracts were analyzed by HPLC-ESI-MS/MS. The method was validated by assessment of intra- and inter-assay accuracy and imprecision on two different analytical instrument platforms. APAP-Cys could be accurately quantified from 0.010 to 10μM, and intra- and inter-assay imprecision were <15% on both analytical instruments. APAP-Cys was stable in human serum for three freeze-thaw cycles and for 24h at ambient temperature. Extracted samples were stable when stored in refrigerated autosamplers for the typical duration of analysis or when stored at -20°C for six days. Results from process efficiency and matrix effect experiments indicated adequate recovery from human serum and insignificant ion suppression or enhancement. The utility and sensitivity of the reported procedure were illustrated by analysis of clinical samples collected from subjects taking chronic, therapeutic doses of APAP. Applicability to other biological matrices was also demonstrated by measurement of protein-derived APAP-Cys in plasma collected from APAP-treated mice, a common animal model of APAP-induced hepatotoxicity. Copyright © 2015 Elsevier B.V. All rights reserved.

Circadian and longitudinal variation of serum C-telopeptide, osteocalcin, and skeletal alkaline phosphatase in C3H/HeJ mice.

PubMed

Srivastava, A K; Bhattacharyya, S; Li, X; Mohan, S; Baylink, D J

2001-10-01

Inbred strains of mice are increasingly being used as an animal model to investigate skeletal disorders relevant to humans. In the bone field, one of the most convenient endpoints for evaluating genetic, physiological, or pharmaceutical perturbations is the use of biochemical markers. To apply biochemical markers in an effective manner, it is of key importance to establish the biological variation and appropriate sampling time. In this study, we evaluate two components: (i) circadian changes, and (ii) longitudinal variation for three serum markers, osteocalcin, C-telopeptide, and skeletal alkaline phosphatase (sALP), using 6-week-old C3H/HeJ (C3H) mice. To study circadian rhythms, the mice were randomly divided into eight groups of 15 mice each. Blood was collected at 3 h intervals, starting at 9:00 A.M. and continuing until 6:00 A.M. the next day. To determine whether circadian rhythm is intrinsically regulated or influenced by restricted food intake, it was also studied after a 12 h fasting period. Serum osteocalcin and C-telopeptide levels were measured by enzyme-linked immunoassay (ELISA) and skeletal alkaline phosphatase by a kinetic assay. The results demonstrated significant circadian variations in osteocalcin and C-telopeptide levels with a peak value between 0900 and 1200 h during daytime and a nadir between 15:00 and 18:00 h. The peak levels of C-telopeptide and osteocalcin were 26%-66% higher as compared with 24 h mean values. The pattern of the circadian variation of C-telopeptide and osteocalcin was similar in female and male animals and was not significantly affected by restricted food intake. The sALP levels were only marginally affected by the circadian rhythm. Longitudinal variations, expressed as coefficient of variation (CV), for osteocalcin, C-telopeptide, and sALP concentrations were 17%, 14%, and 16%, respectively. In addition, the longitudinal variations were not significantly influenced by the time of blood collection in sALP and osteocalcin levels, whereas C-telopeptide levels showed significantly higher within-subject day-to-day variation in morning samples, as compared with blood samples collected in the afternoon. The results highlight the importance of: (i) the timing of sample collection for appropriate interpretation of the bone marker data; and (ii) using the appropriate number of samples based on the variance obtained herein.

Rapid and simple determination of selenium in blood serum by inductively coupled plasma-mass spectrometry (ICP-MS).

PubMed

Labat, L; Dehon, B; Lhermitte, M

2003-05-01

An inductively coupled plasma mass spectrometer (ICP-MS) with a rapid sample-preparative procedure was used for the determination of selenium in blood serum. Blood serum was prepared by dilution in an acidic solution consisting of nitric acid (1%), X-triton (0.1%) and 1-butanol (0.8%). A calibration curve was established for 1-40 microg mL(-1) (r(2)>0.99). The limit of detection was 0.5 microg mL(-1). Repeatability and intermediate precision were satisfactory with relative standard deviations (RSD) of 2.0% and 3.2%, respectively. This method was easily applied to reference materials with satisfactory accuracy. Good correlation (r(2)=0.96) was observed between ICP-MS and atomic absorption spectrometry (AAS) for the determination of (82)Se in blood serum from 23 patients. These results suggest that the sample preparative procedure coupled with ICP-MS can be used for the routine determination of (82)Se in human blood serum.

Non-invasive optical detection of HBV based on serum surface-enhanced Raman spectroscopy

NASA Astrophysics Data System (ADS)

Zheng, Zuci; Wang, Qiwen; Weng, Cuncheng; Lin, Xueliang; Lin, Yao; Feng, Shangyuan

2016-10-01

An optical method of surface-enhanced Raman spectroscopy (SERS) was developed for non-invasive detection of hepatitis B surface virus (HBV). Hepatitis B virus surface antigen (HBsAg) is an established serological marker that is routinely used for the diagnosis of acute or chronic hepatitis B virus(HBV) infection. Utilizing SERS to analyze blood serum for detecting HBV has not been reported in previous literature. SERS measurements were performed on two groups of serum samples: one group for 50 HBV patients and the other group for 50 healthy volunteers. Blood serum samples are collected from healthy control subjects and patients diagnosed with HBV. Furthermore, principal components analysis (PCA) combined with linear discriminant analysis (LDA) were employed to differentiate HBV patients from healthy volunteer and achieved sensitivity of 80.0% and specificity of 74.0%. This exploratory work demonstrates that SERS serum analysis combined with PCA-LDA has tremendous potential for the non-invasive detection of HBV.

Association between Nitrogen Stable Isotope Ratios in Human Hair and Serum Levels of Leptin.

PubMed

Ahn, Song Vogue; Koh, Sang-Baek; Lee, Kwang-Sik; Bong, Yeon-Sik; Park, Jong-Ku

2017-10-01

Stable isotope ratios have been reported to be potential biomarkers of dietary intake and nutritional status. High serum levels of leptin, a hormone which regulates energy metabolism and food intake, are associated with insulin resistance and metabolic syndrome. However, little is known about the association between stable isotope ratios and the metabolic risk in humans. We investigated whether the carbon and nitrogen stable isotope ratios in hair are associated with serum leptin levels. Hair samples were collected from 399 healthy adults (233 men and 166 women) aged 40 to 70 years of a community-based cohort in Korea and the bulk stable isotope ratios of carbon (δ 13 C) and nitrogen (δ 15 N) were measured for all hair samples. Serum leptin levels were analyzed by radioimmunoassay. δ 15 N showed positive correlations with serum leptin levels. In multivariate models, increasing δ 15 N were associated with elevated serum leptin levels (defined as ≥ the median values), whereas δ 13 C were not significantly associated with serum leptin levels. The odds ratio (95% confidence interval) per 1‰ increase in δ 15 N for an elevated serum leptin level was 1.58 (1.11-2.26). In participants with high body mass index, δ 15 N showed positive associations with serum leptin levels, whereas these associations were not seen in participants with low body mass index. The nitrogen stable isotopic ratio in hair is positively associated with serum leptin levels. The hair δ 15 N could be used as a clinical marker to estimate metabolic risk.

Reciprocal contribution of clinical studies and the HP10 antigen ELISA for the diagnosis of extraparenchymal neurocysticercosis.

PubMed

Parkhouse, R Michael E; Carpio, Arturo; Campoverde, Alfredo; Sastre, Patricia; Rojas, Glenda; Cortez, María Milagros

2018-02-01

To evaluate diagnosis of active neurocysticercosis, paired cerebral spinal fluid (CSF) and serum samples from 24 neurocysticercosis (NCC) patients and 17 control neurological patients were assayed in the HP10 Taenia antigen (Ag) ELISA. The CSF samples were also tested with an HP10 Lateral Flow Assay (LFA). The HP10 Ag was detected by ELISA in the CSF of 5/5 patients with Definitive extraparenchymal NCC, and in 4/5 of the corresponding sera. In the Definitive parenchymal group, on the other hand, the HP10 Ag was absent in 2/3 CSF (with a very low value in the one positive sample) and all the corresponding serum samples. Samples of CSF from 4/7 patients in the Probable parenchymal group, were also significantly HP10 Ag positive, suggesting the presence of extraparenchymal cysts not identified by the imaging studies. With the possible exception of one patient, the corresponding serum samples of the Probable parenchymal NCC group, were all HP10 Ag negative. Samples of CSF from 9 NCC patients diagnosed with Mixed parenchymal and extraparenchymal NCC were all significantly HP10 Ag positive, confirming the presence of extraparenchymal cysts, with only 7/9 of the corresponding serum samples being HP10 positive. Thus detection of the HP10 Ag indicates extraparenchymal and not parenchymal cyst localization and is more sensitive with CSF than serum. Three neurological patients clinically diagnosed as subarachnoid cyst, hydrocephalus and tuberculoma, respectively, were clearly positive for HP10 Ag. Of these, two were confirmed as NCC by subsequent imaging; the third died prior to further examination. Thus, a total of 8 patients had their clinical diagnosis questioned. Finally, there was good agreement between the HP10 Ag ELISA and LFA with CSF samples giving an optical density ≥0.4 in the ELISA assay. In conclusion, the HP10 Ag assay should provide a valuable and reciprocal tool in the clinical diagnosis and follow up of extraparenchymal NCC. Copyright © 2017 Elsevier B.V. All rights reserved.

Inositol and hepatic lipidosis. II. Effect of inositol supplementation and time from parturition on serum insulin, thyroxine and triiodothyronine and their relationship to serum and liver lipids in dairy cows.

PubMed

Gerloff, B J; Herdt, T H; Wells, W W; Nachreiner, R F; Emery, R S

1986-06-01

Percutaneous liver biopsies and blood samples were obtained from 80 dairy cows in nine Michigan herds over the peripartum period. Thirty-nine cows were fed 17 g of supplemental inositol and 41 were fed a placebo. Liver biopsies were assayed for total myoinositol and triglyceride (TG) concentrations. Blood samples were assayed for serum dextran precipitable cholesterol, nonesterified fatty acids (NEFA), insulin, thyroxine (T4), free (FT4), triiodothyronine (T3) and free T3 (FT3) concentrations. Serum concentrations of insulin and the thyroid hormones decreased near parturition, with lowest concentrations occurring in the immediate postpartum period. Concentrations of T3 correlated well with T4, and the concentrations of free thyroid hormones reflected concentrations of total thyroid hormones. The percentage of hormone in the free fraction remained constant over time. Serum insulin, T3 and T4 were negatively correlated with serum NEFA and liver TG concentrations. Thyroid hormone concentrations were positively correlated with serum dextran precipitable cholesterol concentrations. Inositol supplementation was associated with reduced circulating T3 and FT3 concentrations, but not T4 and FT4 concentrations. Changes in hormone concentrations at parturition and their relationship to liver TG and serum NEFA concentrations were consistent with a metabolic adaptation by the dairy cow to the negative energy balance of early lactation.

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2010 CFR

2010-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2012 CFR

2012-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2011 CFR

2011-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2013 CFR

2013-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2014 CFR

2014-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

Brominated flame retardant (BFRs) and Dechlorane Plus (DP) in paired human serum and segmented hair.

PubMed

Qiao, Lin; Zheng, Xiao-Bo; Yan, Xiao; Wang, Mei-Huang; Zheng, Jing; Chen, She-Jun; Yang, Zhong-Yi; Mai, Bi-Xian

2018-01-01

Brominated flame retardants (BFRs) and Dechlorane Plus (DP) were measured in both human hair and paired serum samples from a cohort of university students in South China. Segmental analysis was conducted to explore gender difference and the relationships between the hair and serum. The concentrations of total PBDEs in the hair and serum samples were in a range of 0.28-34.1ng/g dry weight (dw) and 0.16-156ng/g lipid weight (lw), respectively. Concentrations of ∑DPs (sum of the syn-DP and anti-DP isomers) in all hair samples ranged from nd-5.45ng/g dry weight. Concentrations of most PBDEs and decabromodiphenylethane (DBDPE) in distal segments (5-10cm from the scalp) were higher than those in the proximal segments (0-5cm from the scalp) (t-test, p < 0.05), which could be due to the longer exposure time of distal segments. The proximal segments exhibited a unique congener profile, more close to that in the serum rather than the distal segments of hair. An obvious gender difference was found in the levels of ∑PBDEs using integrated hair samples, while the difference disappeared when considering alone the proximal segments of hair (0-5cm from scalp) for both genders. This paper provides supplement to the current knowledge on sources of BFRs and DPs in hair and declares the importance of segmental analysis. Copyright © 2017 Elsevier Inc. All rights reserved.

Identification of periparturient mare and foal associated predictors of post parturient immunoglobulin A concentrations in Thoroughbred foals.

PubMed

Jenvey, C; Caraguel, C; Howarth, G B; Riley, C B

2012-12-01

Prior to the start of endogenous production of immunoglobulins (Igs), absorption of maternal Igs is important to protect against pathogens in the early neonatal period. It is possible that mare- or foal-associated factors may influence neonatal IgA concentrations. The temporal relationships among serum and milk IgA concentrations in Thoroughbred mare-foal pairs were explored to determine if periparturient mare- and foal-associated factors contribute to the prediction of foal serum IgA concentrations. Blood and milk samples as well as complete veterinary records, were collected for 84 Thoroughbred mare-foal pairs from one month before to 2 months after parturition. Samples were tested using enzyme-linked immunosorbent assay (ELISA) for concentrations of IgA. Pairwise correlation coefficients were estimated (P < 0.01) and simple linear regression used to investigate unconditional associations between mare IgA levels, mare and foal risk factors and foal serum IgA concentration at 12 h. Backwards, stepwise elimination of nonsignificant factors was used to create a final model. There were significant temporal relationships among mare serum IgA and among colostrum and milk IgA concentrations within mares (P < 0.01). Mare serum IgA concentrations up to one month before parturition were associated with foal serum IgA concentrations at all time points and with colostrum and milk IgA concentrations. Mare serum IgA at -28 days and parity were associated with foal serum IgA concentration at 12 h (P < 0.001). Mare serum IgA concentrations up to 28 days before parturition, together with mare parity, are indicative of neonatal foal serum IgA concentrations. Mare serum and colostrum IgA concentrations may be useful peripartum predictors of neonatal mucosal immune status, enabling earlier intervention to prevent the consequences of mucosal infections.

Glycoblotting method allows for rapid and efficient glycome profiling of human Alzheimer's disease brain, serum and cerebrospinal fluid towards potential biomarker discovery.

PubMed

Gizaw, Solomon T; Ohashi, Tetsu; Tanaka, Masakazu; Hinou, Hiroshi; Nishimura, Shin-Ichiro

2016-08-01

Understanding of the significance of posttranslational glycosylation in Alzheimer's disease (AD) is of growing importance for the investigation of the pathogenesis of AD as well as discovery research of the disease-specific serum biomarkers. We designed a standard protocol for the glycoblotting combined with MALDI-TOFMS to perform rapid and quantitative profiling of the glycan parts of glycoproteins (N-glycans) and glycosphingolipids (GSLs) using human AD's post-mortem samples such as brain tissues (dissected cerebral cortices such as frontal, parietal, occipital, and temporal domains), serum and cerebrospinal fluid (CSF). The structural profiles of the major N-glycans released from glycoproteins and the total expression levels of the glycans were found to be mostly similar between the brain tissues of the AD patients and those of the normal control group. In contrast, the expression levels of the serum and CSF protein N-glycans such as bisect-type and multiply branched glycoforms were increased significantly in AD patient group. In addition, the levels of some gangliosides such as GM1, GM2 and GM3 appeared to alter in the AD patient brain and serum samples when compared with the normal control groups. Alteration of the expression levels of major N- and GSL-glycans in human brain tissues, serum and CSF of AD patients can be monitored quantitatively by means of the glycoblotting-based standard protocols. The changes in the expression levels of the glycans derived from the human post-mortem samples uncovered by the standardized glycoblotting method provides potential serum biomarkers in central nervous system disorders and can contribute to the insight into the molecular mechanisms in the pathogenesis of neurodegenerative diseases and future drug discovery. Most importantly, the present preliminary trials using human post-mortem samples of AD patients suggest that large-scale serum glycomics cohort by means of various-types of human AD patients as well as the normal control sera can facilitate the discovery research of highly sensitive and reliable serum biomarkers for an early diagnosis of AD. This article is part of a Special Issue entitled "Glycans in personalised medicine" Guest Editor: Professor Gordan Lauc. Copyright © 2016 Elsevier B.V. All rights reserved.

Calcium isotope signature: new proxy for net change in bone volume for chronic kidney disease and diabetic rats.

PubMed

Tanaka, Yu-Ki; Yajima, Nobuyuki; Higuchi, Yusuke; Yamato, Hideyuki; Hirata, Takafumi

2017-12-01

Herein, we measure the Ca isotope ratios ( 44 Ca/ 42 Ca and 43 Ca/ 42 Ca) in serum and bone samples collected from rats with chronic kidney disease (CKD) or diabetes mellitus (DM). For the serum samples, the isotope ratios are lower for the CKD (δ 44 Ca/ 42 Ca serum = 0.16 ± 0.11‰; 2SD, n = 6) and the DM (δ 44 Ca/ 42 Ca serum = -0.11 ± 0.25‰; 2SD, n = 7) rats than that for the control rats (δ 44 Ca/ 42 Ca serum = 0.25 ± 0.04‰; 2SD, n = 7). Bone samples from two distinct positions of 20 rats in total, namely, the center and proximal parts of the tibial diaphysis, are subject to Ca isotope analysis. The resulting δ 44 Ca/ 42 Ca values for the bone of the proximal part are about 0.3‰ lower than that for the serum samples from the same rats. The larger isotope fractionations between the serum and bone are consistent with previously reported data for vertebrate animals (e.g., Skulan and DePaolo, 1999), which suggests the preferential incorporation of lighter Ca isotopes through bone formation. For the bones from the control and CKD rats, there were no differences in the δ 44 Ca/ 42 Ca values between the positions of the bone. In contrast, the δ 44 Ca/ 42 Ca values of the bone for the DM rats were different between the positions of the bone. Due to the lower bone turnover rate for the DM rats, the δ 44 Ca/ 42 Ca for the middle of the diaphysis can reflect the Ca isotopes in the bone formed prior to the progression of DM states. Thus, the resulting δ 44 Ca/ 42 Ca values show a clear correlation with bone mineral density (BMD). This can be due to the release of isotopically lighter Ca from the bone to the serum. In the present study, our data demonstrate that the δ 44 Ca/ 42 Ca value for serum can be used as a new biomarker for evaluating changes in bone turnover rate, followed by changes in bone volume.

Variations in the detection of anti-PEDV antibodies in serum samples using three diagnostic tests - short communication.

PubMed

Plut, Jan; Toplak, Ivan; Štukelj, Marina

2018-06-01

Over the last few years several porcine epidemic diarrhoea (PED) outbreaks have been discovered in Europe including the first PED case in Slovenia in January 2015. The aim of this study was to determine when PED virus (PEDV) infection started in Slovenia. Serum samples collected between 2012 and 2016 were tested. Three hundred and seventy-five serum samples were collected from 132 Slovenian small, one-site pig farms. Samples were tested for PEDV antibodies utilising three different serological methods: commercially-available indirect ELISA, in-house blocking ELISA test and Immunoperoxidase Monolayer Assay (IPMA) test. One hundred and seventy (45.33%) tested samples were found positive by the commercially-available ELISA test kit, and 10 (5.68%) of these 170 samples found positive were positive by the in-house blocking ELISA. Only these 10 samples were collected from a farm where clinical signs of PED infection had been observed and PEDV was confirmed by RT-PCR methodology; the other 160 samples were collected randomly. Thirty-two samples with the highest S/P value obtained with the commercial ELISA were all negative with IPMA. Reasons for the high variance in the results obtained remain unclear; more research is required to ensure higher sensitivity and specificity in terms of PEDV antibody tests and other PED diagnostic methods.

NMR Metabolomics Investigates the Influence of Flavonoid-Enriched Rations on Chicken Plasma.

PubMed

Fotakis, Charalambos; Lantzouraki, Dimitra Z; Goliomytis, Michael; Simitzis, Panagiotis E; Charismiadou, Maria; Deligeorgis, Stelios G; Zoumpoulakis, Panagiotis

2017-03-01

The use of flavonoids as dietary supplements is well established, mainly due to their intense antioxidant and anti-inflammatory properties. In the present study, hesperidin, naringin, and vitamin E were used as additives at different concentrations in poultry rations in order to achieve meat of improved quality. NMR metabolomics was applied to chicken blood serum samples to discern whether and how the enriched rations affected the animals' metabolic profile. Variations in the metabolic patterns according to sustenance consumption were traced by orthogonal projections to latent structures discriminant analysis (OPLS-DA) models and were attributed to specific metabolites by using S-line plots. In particular, serum samples from chickens fed with vitamin E displayed higher concentrations of glycine and succinic acid compared to control samples, which were mainly characterized by betaine, formic acid, and lipoproteins. Samples from chickens fed with hesperidin were characterized by increased levels of lactic acid, citric acid, creatine, carnosine, creatinine, phosphocreatine, anserine, glucose, and alanine compared to control samples. Lastly, naringin samples exhibited increased levels of citric and acetic acids. Results verify the scalability of NMR metabolomics to highlight metabolite variations among chicken serum samples in relation to food rations.

Long-term stability of glucose: glycolysis inhibitor vs. gel barrier tubes.

PubMed

Winter, Theresa; Hannemann, Anke; Suchsland, Juliane; Nauck, Matthias; Petersmann, Astrid

2018-03-12

Measuring the glucose concentration in whole blood samples is critical due to unsatisfactory glycolysis inhibition. Previous studies showed that Terumo tubes were superior, but they were taken off the European market in 2016 and alternatives were required. This initiated the present evaluation of glucose stability in five available tube types. Venous blood samples were collected from 61 healthy volunteers to test tubes supplied by Terumo (two sets), Greiner FC-Mix, BD FX-Mixture and BD serum. After sampling, the contents were thoroughly mixed and centrifuged within an hour. The glucose concentrations were determined and the samples resuspended except for BD serum tubes (gel barrier). The first 30 samples were stored at room temperature and the remaining 31 at 4°C. After 24, 48, 72 and 96 h, all tubes were (re)centrifuged, and glucose concentration measurements were repeated. Changes in glucose concentrations over time differed significantly between the investigated tube types and to a certain extent between the two storing conditions. Glycolysis was most evident in the BD FX-mixture tubes. Good glucose stability was observed in samples retrieved form BD serum and Greiner tubes. The stability in both Terumo tubes was comparable to that in other studies. Although Greiner and both Terumo tubes are supposed to contain the same glycolysis inhibitor, glucose stability differed between these tubes. We showed that Greiner is an acceptable alternative to Terumo and that glucose in serum that was rapidly separated from corpuscles by a gel barrier is stable for an extended time.

Serum proteomics reveals systemic dysregulation of innate immunity in type 1 diabetes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Qibin; Fillmore, Thomas L.; Schepmoes, Athena A.

Using global liquid chromatography-mass spectrometry (LC-MS)-based proteomics analyses, we identified 24 serum proteins significantly variant between those with type 1 diabetes and healthy controls. Functionally, these proteins represent innate immune responses, the activation cascade of complement, inflammatory responses and blood coagulation. Targeted verification analyses were performed on 52 surrogate peptides representing these proteins with serum samples from an antibody standardization program cohort of 100 healthy control and 50 type 1 diabetic subjects, and 16 peptides were verified having very good discriminating power, with areas under the receiver operator characteristic curve ≥ 0.8. Further validation with blinded serum samples from anmore » independent cohort (10 healthy control and 10 type 1 diabetic) demonstrated that peptides from platelet basic protein and C1 inhibitor achieved both 100% sensitivity and 100% specificity for classification of samples. The disease specificity of these proteins was assessed using serum from 50 age matched type 2 diabetic individuals, and a subset of proteins, particularly C1 inhibitor were exceptionally good discriminators between these two forms of diabetes. The panel of biomarkers distinguishing those with type 1 diabetes from healthy control and type 2 diabetes suggests dysregulated innate immune responses may be associated with the development of this disorder.« less

Assessment of cytologic evaluation of preputial epithelial cells as a diagnostic test for detection of adrenocortical disease in castrated ferrets.

PubMed

Protain, Holly J; Kutzler, Michelle A; Valentine, Beth A

2009-05-01

To determine whether results of cytologic evaluation of preputial epithelial cells correspond to results of a serum endocrine hormone assay and clinical signs associated with adrenocortical disease in castrated ferrets. 13 clinically normal ferrets and 8 ferrets with signs of adrenocortical disease. Blood and preputial lavage samples were collected from each ferret. Serum samples were submitted to the University of Tennessee Veterinary Diagnostic Laboratory for performance of an endocrine hormone assay. Differential epithelial cell counts were performed on preputial lavage samples to determine the percentage of cornified cells. Results of cytologic evaluation were compared with results of the endocrine hormone assay and clinical status of ferrets. The percentage of cornified preputial epithelial cells was not significantly correlated with serum 17B-estradiol or androstenedione concentration but was significantly correlated with serum 17-hydroxyprogesterone concentration (r = 0.60). The percentage of cornified preputial epithelial cells was higher in ferrets with clinical signs of adrenocortical disease (mean +/- SD, 71.3 +/- 16.9%) than in clinically normal ferrets (55.5 +/- 19.0%). Cornification of preputial epithelial cells was correlated with an increase in serum 17-hydroxyprogesterone concentration as well as clinical signs of adrenocortical disease in castrated ferrets. Additional investigation is needed to elucidate the mechanism of preputial epithelial cell cornification in castrated ferrets.

Simplifying sampling for African swine fever surveillance: Assessment of antibody and pathogen detection from blood swabs.

PubMed

Carlson, J; Zani, L; Schwaiger, T; Nurmoja, I; Viltrop, A; Vilem, A; Beer, M; Blome, S

2018-02-01

African swine fever (ASF) is a notifiable disease with serious socio-economic consequences that has been present in wild boar in the Baltic States and Poland since 2014. An introduction of ASF is usually accompanied by increased mortality, making fallen wild boar and hunted animals with signs of disease the main target for early warning and passive surveillance. It is difficult, however, to encourage hunters and foresters to report and take samples from these cases. A pragmatic and easy sampling approach with quick-drying swabs could facilitate this. In this study, we further evaluated the use of dry blood swabs for the detection of ASFV antibody and genome with samples from animal trials and diagnostic submissions (blood, bone and organs) from Estonia. Compared to serum samples, dried blood swabs yielded 93.1% (95% confidence interval: [83.3, 98.1]) sensitivity and 100% [95.9, 100.0] specificity in a commercial ASFV antibody ELISA. Similarly, the swabs gave a sensitivity of 98.9% [93.4, 100.0] and a specificity of 98.1% [90.1, 100.0] for genome detection by a standard ASFV p72 qPCR when compared to EDTA blood. The same swabs were tested in a VP72-antibody lateral flow device, with a sensitivity of 94.7% [85.4, 98.9] and specificity of 96.1% [89.0, 99.2] compared to the serum ELISA. When GenoTube samples tested in ELISA and LFD were compared, the sensitivity was 96.3% [87.3, 99.5] and the specificity was 93.8% [86.0, 97.9]. This study demonstrates reliable detection of ASFV antibody and genome from swabs. A field test of the swabs with decomposed wild boar carcasses in an endemic area in Estonia also gave promising results. Thus, this technique is a practical approach for surveillance of ASF in both free and endemic areas. © 2017 Blackwell Verlag GmbH.

Porcine reproductive and respiratory syndrome virus (PRRSV) surveillance using pre-weaning oral fluid samples detects circulation of wild-type PRRSV.

PubMed

Kittawornrat, Apisit; Panyasing, Yaowalak; Goodell, Christa; Wang, Chong; Gauger, Phillip; Harmon, Karen; Rauh, Rolf; Desfresne, Luc; Levis, Ian; Zimmerman, Jeffrey

2014-01-31

Oral fluid samples collected from litters of piglets (n=600) one day prior to weaning were evaluated as a method to surveil for porcine reproductive and respiratory syndrome virus (PRRSV) infections in four sow herds of approximately 12,500 sow each. Serum samples from the litters' dam (n=600) were included for comparison. All four herds were endemically infected with PRRSV and all sows had been vaccinated ≥ 2 times with PRRSV modified-live virus vaccines. After all specimens had been collected, samples were randomized and assayed by PRRSV real-time reverse transcription polymerase chain reaction (RT-qPCR) and four PRRSV antibody ELISA assays (IgM, IgA, IgG, and Commercial Kit). All sow serum samples were negative by PRRSV RT-qPCR, but 9 of 600 oral fluid samples tested positive at two laboratories. Open reading frame 5 (ORF5) sequencing of 2 of the 9 positive oral fluid samples identified wild-type viruses as the source of the infection. A comparison of antibody responses in RT-qPCR positive vs. negative oral fluid samples showed significantly higher IgG S/P ratios in RT-qPCR-positive oral fluid samples (mean S/P 3.46 vs. 2.36; p=0.02). Likewise, sow serum samples from RT-qPCR-positive litter oral fluid samples showed significantly higher serum IgG (mean S/P 1.73 vs. 0.98; p<0.001) and Commercial Kit (mean S/P 1.97 vs. 0.98; p<0.001) S/P ratios. Overall, the study showed that pre-weaning litter oral fluid samples could provide an efficient and sensitive approach to surveil for PRRSV in infected, vaccinated, or presumed-negative pig breeding herds. Copyright © 2014 Elsevier B.V. All rights reserved.

Human serum miR-34a as an indicator of exposure to ionizing radiation.

PubMed

Halimi, Mohammad; Shahabi, Ahmad; Moslemi, Dariush; Parsian, Hadi; Asghari, S Mohsen; Sariri, Reyhaneh; Yeganeh, Farshid; Zabihi, Ebrahim

2016-11-01

Radiation exposure in industrial accidents or nuclear device attacks is a major public health concern. There is an urgent need for markers that rapidly identify people exposed to ionizing radiation (IR). Finding a blood-based marker is advantageous because of the ease of sample collection. This study was designed to test the hypothesis that serum miR-34a could serve as an indicator of exposure to IR. Therefore, 44 women with breast cancer, where radiotherapy was part of their therapeutic protocol, were investigated in this study. After demonstrating the appropriateness of our microRNA (miRNA) extraction efficiency and miRNA assay in human serum, we analyzed the miR-34a level in paired serum samples before and after radiotherapy. Fifty Gy X-ray irradiation in daily dose fractions of 2 Gy, 5 days per week, was used in this study. We demonstrated that IR significantly increased serum level of miR-34a. By measuring miR-34a in serum, we could distinguish irradiated patients with sensitivity of 65 % and specificity of 75 %. According to this study, serum miR-34a has the potential to be used as an indicator of radiation exposure.

A Serum Protein Profile Predictive of the Resistance to Neoadjuvant Chemotherapy in Advanced Breast Cancers*

PubMed Central

Hyung, Seok-Won; Lee, Min Young; Yu, Jong-Han; Shin, Byunghee; Jung, Hee-Jung; Park, Jong-Moon; Han, Wonshik; Lee, Kyung-Min; Moon, Hyeong-Gon; Zhang, Hui; Aebersold, Ruedi; Hwang, Daehee; Lee, Sang-Won; Yu, Myeong-Hee; Noh, Dong-Young

2011-01-01

Prediction of the responses to neoadjuvant chemotherapy (NACT) can improve the treatment of patients with advanced breast cancer. Genes and proteins predictive of chemoresistance have been extensively studied in breast cancer tissues. However, noninvasive serum biomarkers capable of such prediction have been rarely exploited. Here, we performed profiling of N-glycosylated proteins in serum from fifteen advanced breast cancer patients (ten patients sensitive to and five patients resistant to NACT) to discover serum biomarkers of chemoresistance using a label-free liquid chromatography-tandem MS method. By performing a series of statistical analyses of the proteomic data, we selected thirteen biomarker candidates and tested their differential serum levels by Western blotting in 13 independent samples (eight patients sensitive to and five patients resistant to NACT). Among the candidates, we then selected the final set of six potential serum biomarkers (AHSG, APOB, C3, C9, CP, and ORM1) whose differential expression was confirmed in the independent samples. Finally, we demonstrated that a multivariate classification model using the six proteins could predict responses to NACT and further predict relapse-free survival of patients. In summary, global N-glycoproteome profile in serum revealed a protein pattern predictive of the responses to NACT, which can be further validated in large clinical studies. PMID:21799047

Polyfluorinated Compounds in Serum Linked to Indoor Air in Office Environments

PubMed Central

Fraser, Alicia J; Webster, Thomas F; Watkins, Deborah J; Nelson, Jessica W; Stapleton, Heather M; Calafat, Antonia M; Kato, Kayoko; Shoeib, Mahiba; Vieira, Verónica M; McClean, Michael D

2011-01-01

We aimed to investigate the role of indoor office air on exposure to polyfluorinated compounds (PFCs) among office workers. Week-long, active air sampling was conducted during the winter of 2009 in 31 offices in Boston, MA. Air samples were analyzed for fluorotelomer alcohols (FTOHs), sulfonamides (FOSAs), and sulfonamidoethanols (FOSEs). Serum was collected from each participant (n=31) and analyzed for twelve PFCs including PFOA and PFOS. In air, FTOHs were present in the highest concentrations, particularly 8:2-FTOH (GM=9,920 pg/m3). FTOHs varied significantly by building with the highest levels observed in a newly constructed building. PFOA in serum was significantly correlated with air levels of 6:2-FTOH (r=0.43), 8:2-FTOH (r=0.60), and 10:2-FTOH (r=0.62). Collectively, FTOHs in air significantly predicted PFOA in serum (p < 0.001) and explained approximately 36% of the variation in serum PFOA concentrations. PFOS in serum was not associated with air levels of FOSAs/FOSEs. In conclusion, FTOH concentrations in office air significantly predict serum PFOA concentrations in office workers. Variation in PFC air concentrations by building is likely due to differences in the number, type, and age of potential sources such as carpeting, furniture and/or paint. PMID:22148395

Polyfluorinated compounds in serum linked to indoor air in office environments.

PubMed

Fraser, Alicia J; Webster, Thomas F; Watkins, Deborah J; Nelson, Jessica W; Stapleton, Heather M; Calafat, Antonia M; Kato, Kayoko; Shoeib, Mahiba; Vieira, Verónica M; McClean, Michael D

2012-01-17

We aimed to investigate the role of indoor office air on exposure to polyfluorinated compounds (PFCs) among office workers. Week-long, active air sampling was conducted during the winter of 2009 in 31 offices in Boston, MA. Air samples were analyzed for fluorotelomer alcohols (FTOHs), sulfonamides (FOSAs), and sulfonamidoethanols (FOSEs). Serum was collected from each participant (n = 31) and analyzed for 12 PFCs including PFOA and PFOS. In air, FTOHs were present in the highest concentrations, particularly 8:2-FTOH (GM = 9920 pg/m(3)). FTOHs varied significantly by building with the highest levels observed in a newly constructed building. PFOA in serum was significantly correlated with air levels of 6:2-FTOH (r = 0.43), 8:2-FTOH (r = 0.60), and 10:2-FTOH (r = 0.62). Collectively, FTOHs in air significantly predicted PFOA in serum (p < 0.001) and explained approximately 36% of the variation in serum PFOA concentrations. PFOS in serum was not associated with air levels of FOSAs/FOSEs. In conclusion, FTOH concentrations in office air significantly predict serum PFOA concentrations in office workers. Variation in PFC air concentrations by building is likely due to differences in the number, type, and age of potential sources such as carpeting, furniture, and/or paint.

Improved micromethod for mezlocillin quantitation in serum and urine by high-pressure liquid chromatography.

PubMed Central

Fiore, D; Auger, F A; Drusano, G L; Dandu, V R; Lesko, L J

1984-01-01

A rapid, sensitive, and specific method of analysis for mezlocillin in serum and urine by high-pressure liquid chromatography is described. A solid-phase extraction column was used to remove interfering substances from samples before chromatography. Quantitation included the use of an internal standard, nafcillin. Mezlocillin was chromatographed with a phosphate buffer-acetonitrile (73:27) mobile phase and a C-18 reverse-phase column and detected at a wavelength of 220 nm. The assay had a sensitivity of 1.6 micrograms/ml and a linearity of up to 600 micrograms/ml and 16 mg/ml in serum and urine, respectively, with only 0.1 ml of sample. The interday and intraday coefficients of variation for replicate analyses of spiked serum and urine specimens were less than 6.5%. PMID:6517560

Evidence to Support a Contribution of Polyreactive Antibodies to HLA Serum Reactivity.

PubMed

Gao, Baoshan; Rong, Chunshu; Porcheray, Fabrice; Moore, Carolina; Girouard, Timothy C; Saidman, Susan L; Wong, Waichi; Fu, Yaowen; Zorn, Emmanuel

2016-01-01

Assessing the serum reactivity to HLA is essential for the evaluation of transplant candidates and the follow-up of allograft recipients. In this study, we look for evidence at the clonal level that polyreactive antibodies cross-reactive to apoptotic cells and multiple autoantigens can also react to HLA and contribute to the overall serum reactivity. We immortalized B cell clones from the blood of 2 kidney transplant recipients and characterized their reactivity to self-antigens, apoptotic cells as well as native, denatured, and cryptic HLA determinants using enzyme-linked immunosorbent assay (ELISA), immunofluorescence, flow cytometry and Luminex assays. We also assessed the reactivity of 300 pretransplant serum specimens to HLA and apoptotic cells. We report here 4 distinct B cell clones cross-reactive to self and HLA class I. All 4 clones reacted to numerous HLA class I alleles but did not appear to target canonical "shared" epitopes. In parallel experiments, we observed a strong correlation between IgG reactivity to HLA and apoptotic cells in pretransplant serum samples collected from 300 kidney transplant recipients. Further analysis revealed that samples with higher reactivity to apoptotic cells displayed significantly higher class I percent panel-reactive antibodies compared to samples with low reactivity to apoptotic cells. We provide here (1) proof of principle at the clonal level that human polyreactive antibodies can cross-react to HLA, multiple self-antigens and apoptotic cells and (2) supportive evidence that polyreactive antibodies contribute to overall HLA reactivity in the serum of patients awaiting kidney transplant.

Massive glutamine cyclization to pyroglutamic acid in human serum discovered using NMR spectroscopy.

PubMed

Nagana Gowda, G A; Gowda, Yashas N; Raftery, Daniel

2015-04-07

Glutamine is one of the most abundant metabolites in blood and is a precursor as well as end product central to numerous important metabolic pathways. A number of surprising and unexpected roles for glutamine, including cancer cell glutamine addiction discovered recently, stress the importance of accurate analysis of glutamine concentrations for understanding its role in health and numerous diseases. Utilizing a recently developed NMR approach that offers access to an unprecedented number of quantifiable blood metabolites, we have identified a surprising glutamine cyclization to pyroglutamic acid that occurs during protein removal. Intact, ultrafiltered and protein precipitated samples from the same pool of human serum were comprehensively investigated using (1)H NMR spectroscopy at 800 MHz to detect and quantitatively evaluate the phenomenon. Interestingly, although glutamine cyclization occurs in both ultrafiltered and protein precipitated serum, the cyclization was not detected in intact serum. Strikingly, due to cyclization, the apparent serum glutamine level drops by up to 75% and, concomitantly, the pyroglutamic acid level increases proportionately. Further, virtually under identical conditions, the magnitude of cyclization is vastly different for different portions of samples from the same pool of human serum. However, the sum of glutamine and pyroglutamic acid concentrations in each sample remains the same for all portions. These unexpected findings indicate the importance of considering the sum of apparent glutamine and pyroglutamic acid levels, obtained from the contemporary analytical methods, as the actual blood glutamine level for biomarker discovery and biological interpretations.

Simultaneous determination of neonicotinoid insecticides in human serum and urine using diatomaceous earth-assisted extraction and liquid chromatography-tandem mass spectrometry.

PubMed

Yamamuro, Tadashi; Ohta, Hikoto; Aoyama, Mika; Watanabe, Daisuke

2014-10-15

A rapid and sensitive analytical method was developed for simultaneous determination of eight neonicotinoid insecticides (acetamiprid, clothianidin, dinotefuran, flonicamid, imidacloprid, nitenpyram, thiacloprid and thiamethoxam) and three specific metabolites of acetamiprid (N-desmethylacetamiprid, 5-(N-acetyl-N-methylaminomethyl)-2-chloropyridine and 5-(N-acetylaminomethyl)-2-chloropyridine) in human serum and urine. A diatomaceous earth-assisted extraction using Extrelut NT3 column with chloroform/2-propanol (3:1, v/v) as eluent was selected for the single step cleanup procedure for all the target compounds. Qualitative and quantitative analyses were conducted by liquid chromatography-tandem mass spectrometry with multiple reaction monitoring mode. The limits of detection and the limits of quantification of eleven compounds were in the ranges of 0.1-0.2ng/mL and 0.5-10ng/mL for serum, 0.1-1ng/mL and 1-10ng/mL for urine, respectively. The extraction recoveries were between 80.9% and 101.8% for serum samples, 91.9% and 106% for urine samples. The intra-day RSDs and the inter-day RSDs were less than 11.5% and 13.2% for serum, less than 8.3% and 8.8% for urine. The proposed procedure will be suitable for forensic investigations of human poisoning cases with neonicotinoid insecticides. This is the first report of simultaneous determination of eight neonicotinoids in serum and urine samples. Copyright © 2014. Published by Elsevier B.V.

Exploration of attenuated total reflectance mid-infrared spectroscopy and multivariate calibration to measure immunoglobulin G in human sera.

PubMed

Hou, Siyuan; Riley, Christopher B; Mitchell, Cynthia A; Shaw, R Anthony; Bryanton, Janet; Bigsby, Kathryn; McClure, J Trenton

2015-09-01

Immunoglobulin G (IgG) is crucial for the protection of the host from invasive pathogens. Due to its importance for human health, tools that enable the monitoring of IgG levels are highly desired. Consequently there is a need for methods to determine the IgG concentration that are simple, rapid, and inexpensive. This work explored the potential of attenuated total reflectance (ATR) infrared spectroscopy as a method to determine IgG concentrations in human serum samples. Venous blood samples were collected from adults and children, and from the umbilical cord of newborns. The serum was harvested and tested using ATR infrared spectroscopy. Partial least squares (PLS) regression provided the basis to develop the new analytical methods. Three PLS calibrations were determined: one for the combined set of the venous and umbilical cord serum samples, the second for only the umbilical cord samples, and the third for only the venous samples. The number of PLS factors was chosen by critical evaluation of Monte Carlo-based cross validation results. The predictive performance for each PLS calibration was evaluated using the Pearson correlation coefficient, scatter plot and Bland-Altman plot, and percent deviations for independent prediction sets. The repeatability was evaluated by standard deviation and relative standard deviation. The results showed that ATR infrared spectroscopy is potentially a simple, quick, and inexpensive method to measure IgG concentrations in human serum samples. The results also showed that it is possible to build a united calibration curve for the umbilical cord and the venous samples. Copyright © 2015 Elsevier B.V. All rights reserved.

Aspergillus colonization in patients with bronchogenic carcinoma.

PubMed

Ali, Sana; Malik, Abida; Bhargava, Rakesh; Shahid, Mohammad; Fatima, Nazish

2014-05-01

Aspergillus antigens such as galactomannan antigen, a cell wall polysaccharide, can be detected in patient's serum or bronchoalveolar lavage. To study the prevalence of Aspergillus infection in patients with bronchogenic carcinoma, we measured galactomannan antigen in serum and bronchoalveolar lavage samples of patients with bronchogenic carcinoma. The study was conducted on 45 bronchogenic carcinoma patients. The diagnosis of lung cancer was confirmed by bronchoscopy, histopathological and radiological examinations. Bronchoalveolar lavage fluid collected from each patient by fiberoptic bronchoscopy was subjected to direct microscopy and culture on Sabouraud's dextrose agar and Czapek-Dox agar, and Aspergillus galactomannan antigen was measured in serum and bronchoalveolar lavage samples. The majority of patients were male (93.3%) in the age group 51-60 years, 88.9% were addicted to gutka chewing, and 82.1% were addicted to smoking. Most patients complained of cough (73%) and shortness of breath (51.1%). Squamous cell carcinoma (64.4%) was the most common malignancy, followed by adenocarcinoma (13.3%). On culture of bronchoalveolar lavage samples, 35.5% showed growth of Aspergillus spp. (Aspergillus fumigatus in 17.8%, Aspergillus flavus in 13.3%, and Aspergillus niger in 4.4%). Galactomannan antigen was detected in 58.3% of bronchoalveolar lavage samples and 47.2% of serum samples. There is a high prevalence of aspergillosis in patients with lung carcinoma, especially among smokers and gutka chewers.

An occupational exposure assessment of a perfluorooctanesulfonyl fluoride production site: biomonitoring.

PubMed

Olsen, Geary W; Logan, Perry W; Hansen, Kristen J; Simpson, Cathy A; Burris, Jean M; Burlew, Michele M; Vorarath, Phanasouk P; Venkateswarlu, Pothapragada; Schumpert, John C; Mandel, Jeffrey H

2003-01-01

This investigation randomly sampled a fluorochemical manufacturing employee population to determine the distribution of serum fluorochemical levels according to employees' jobs and work areas. Previous analyses of medical surveillance data have not shown significant associations between fluorochemical production employees' clinical chemistry and hematology tests and their serum PFOS and perfluorooctanoate (PFOA, C(7)F(15)COO(-)) concentrations, but may have been subject to nonparticipation bias. A random sample of the on-site film plant employee population, where fluorochemicals are not produced, determined their serum concentrations also. Of the 232 employees randomly selected for serum sampling, 186 (80%) employees participated (n=126 chemical plant; n=60 film plant). Sera samples were extracted using an ion-pairing extraction procedure and were quantitatively analyzed for seven fluorochemicals using high-pressure liquid chromatography electrospray tandem mass spectrometry methods. Geometric means (in parts per million) and 95% confidence intervals (in parentheses) of the random sample of 126 chemical plant employees were: PFOS 0.941 (0.787-1.126); PFOA 0.899 (0.722-1.120); perfluorohexanesulfonate 0.180 (0.145-0.223); N-ethyl perfluorooctanesulfonamidoacetate 0.008 (0.006-0.011); N-methyl perfluorooctanesulfonamidoacetate 0.081 (0.067-0.098); perfluorooctanesulfonamide 0.013 (0.009-0.018); and perfluorooctanesulfonamidoacetate 0.022 (0.018-0.029). These geometric means were approximately one order of magnitude higher than those observed for the film plant employees.

The prevalence of low serum zinc and copper levels and dietary habits associated with serum zinc and copper in 12- to 36-month-old children from low-income families at risk for iron deficiency.

PubMed

Schneider, Julie M; Fujii, Mary L; Lamp, Catherine L; Lönnerdal, Bo; Zidenberg-Cherr, Sheri

2007-11-01

Iron and zinc share common food sources, and children at risk of iron deficiency may also develop zinc deficiency. We determined the prevalence of zinc and copper deficiency and examined factors associated with serum zinc and copper in young children from low-income families at risk of iron deficiency. A cross-sectional study design was used to assess serum zinc and copper, along with an interview-assisted survey to assess factors associated with serum zinc and copper in a convenience sample. Participants were 435 children aged 12 to 36 months recruited from select clinics of the Special Supplemental Nutrition Program for Women, Infants, and Children in Contra Costa and Tulare Counties, California. Frequencies were used to report prevalence. Multiple linear regressions were conducted to examine factors associated with serum zinc and copper, controlling for age, sex, and ethnicity. The prevalence of low serum zinc level (<70 microg/dL [<10.7 micromol/L]) was 42.8%, and low serum copper level (<90 microg/dL [<14.2 micromol/L]) was <1%. Mean+/-standard deviation of serum copper was 150+/-22 microg/dL (23.6+/-3.5 micromol/L) and 140+/-24 microg/dL (22.1+/-3.8 micromol/L) for anemic and non-anemic children, respectively (t test, P=0.026). In multiple linear regression consumption of sweetened beverages was negatively associated with serum zinc level, and consumption of >15 g/day meat was positively associated with serum zinc level, whereas current consumption of breast milk and >15 g/day beans were positively associated with serum copper level. The prevalence of low serum zinc concentration in the sample was high, and warrants further investigation amongst vulnerable populations.

Longitudinal Study of Hepatitis A Infection by Saliva Sampling: The Kinetics of HAV Markers in Saliva Revealed the Application of Saliva Tests for Hepatitis A Study.

PubMed

Amado Leon, Luciane Almeida; de Almeida, Adilson José; de Paula, Vanessa Salete; Tourinho, Renata Santos; Villela, Daniel Antunes Maciel; Gaspar, Ana Maria Coimbra; Lewis-Ximenez, Lia Laura; Pinto, Marcelo Alves

2015-01-01

Despite the increasing numbers of studies investigating hepatitis A diagnostic through saliva, the frequency and the pattern of hepatitis A virus (HAV) markers in this fluid still remains unknown. To address this issue, we carried on a longitudinal study to examine the kinetics of HAV markers in saliva, in comparison with serum samples. The present study followed-up ten patients with acute hepatitis A infection during 180 days post diagnosis (dpd). Total anti-HAV was detected in paired serum and saliva samples until the end of the follow-up, showing a peak titer at 90th. However, total anti-HAV level was higher in serum than in saliva samples. This HAV marker showed a probability of 100% to be detected in both serum and saliva during 180 dpd. The IgM anti-HAV could be detected in saliva up to 150 dpd, showing the highest frequency at 30th, when it was detected in all individuals. During the first month of HAV infection, this acute HAV marker showed a detection probability of 100% in paired samples. The detection of IgM anti-HAV in saliva was not dependent on its level in serum, HAV-RNA detection and/or viral load, since no association was found between IgM anti-HAV positivity in saliva and any of these parameter (p>0.05). Most of the patients (80%) were found to contain HAV-RNA in saliva, mainly at early acute phase (30th day). However, it was possible to demonstrate the HAV RNA presence in paired samples for more than 90 days, even after seroconversion. No significant relationship was observed between salivary HAV-RNA positivity and serum viral load, demonstrating that serum viral load is not predictive of HAV-RNA detection in saliva. Similar viral load was seen in paired samples (on average 104 copies/mL). These data demonstrate that the best diagnostic coverage can be achieved by salivary anti-HAV antibodies and HAV-RNA tests during 30-90 dpd. The long detection and high probability of specific-HAV antibodies positivity in saliva samples make the assessment of salivary antibodies a useful tool for diagnosis and epidemiological studies. The high frequency of HAV-RNA in saliva and the probability of detection of about 50%, during the first 30 dpd, demonstrate that saliva is also useful for molecular investigation of hepatitis A cases, mainly during the early course of infection. Therefore, the collection of saliva may provide a simple, cheap and non-invasive means of diagnosis, epidemiological surveys and monitoring of hepatitis A infection purposes.

Longitudinal Study of Hepatitis A Infection by Saliva Sampling: The Kinetics of HAV Markers in Saliva Revealed the Application of Saliva Tests for Hepatitis A Study

PubMed Central

Amado Leon, Luciane Almeida; de Almeida, Adilson José; de Paula, Vanessa Salete; Tourinho, Renata Santos; Villela, Daniel Antunes Maciel; Gaspar, Ana Maria Coimbra; Lewis-Ximenez, Lia Laura; Pinto, Marcelo Alves

2015-01-01

Despite the increasing numbers of studies investigating hepatitis A diagnostic through saliva, the frequency and the pattern of hepatitis A virus (HAV) markers in this fluid still remains unknown. To address this issue, we carried on a longitudinal study to examine the kinetics of HAV markers in saliva, in comparison with serum samples. The present study followed-up ten patients with acute hepatitis A infection during 180 days post diagnosis (dpd). Total anti-HAV was detected in paired serum and saliva samples until the end of the follow-up, showing a peak titer at 90th. However, total anti-HAV level was higher in serum than in saliva samples. This HAV marker showed a probability of 100% to be detected in both serum and saliva during 180 dpd. The IgM anti-HAV could be detected in saliva up to 150 dpd, showing the highest frequency at 30th, when it was detected in all individuals. During the first month of HAV infection, this acute HAV marker showed a detection probability of 100% in paired samples. The detection of IgM anti-HAV in saliva was not dependent on its level in serum, HAV-RNA detection and/or viral load, since no association was found between IgM anti-HAV positivity in saliva and any of these parameter (p>0.05). Most of the patients (80%) were found to contain HAV-RNA in saliva, mainly at early acute phase (30th day). However, it was possible to demonstrate the HAV RNA presence in paired samples for more than 90 days, even after seroconversion. No significant relationship was observed between salivary HAV-RNA positivity and serum viral load, demonstrating that serum viral load is not predictive of HAV-RNA detection in saliva. Similar viral load was seen in paired samples (on average 104 copies/mL). These data demonstrate that the best diagnostic coverage can be achieved by salivary anti-HAV antibodies and HAV-RNA tests during 30–90 dpd. The long detection and high probability of specific-HAV antibodies positivity in saliva samples make the assessment of salivary antibodies a useful tool for diagnosis and epidemiological studies. The high frequency of HAV-RNA in saliva and the probability of detection of about 50%, during the first 30 dpd, demonstrate that saliva is also useful for molecular investigation of hepatitis A cases, mainly during the early course of infection. Therefore, the collection of saliva may provide a simple, cheap and non-invasive means of diagnosis, epidemiological surveys and monitoring of hepatitis A infection purposes. PMID:26690904

Severe reactions from roe without concomitant fish allergy.

PubMed

Mäkinen-Kiljunen, Soili; Kiistala, Raija; Varjonen, Elina

2003-10-01

Although fish allergy is common, no studies have been published on allergy to fish roe. To describe 2 cases of IgE-mediated allergy to 2 roe species. Two patients, one with local symptoms and the other with anaphylaxis following ingestion of roe, underwent skin prick testing (SPT) with 2 roe species, whitefish roe (WFR) and rainbow trout roe (RTR). Serum samples were taken for IgE measurement and immunoblotting to identify roe allergens. Inhibition studies were performed to scrutinize the cross-reactivity between the roes and to fish. The results of the SPTs with the roes were clearly positive in both patients but negative in control persons. The results of SPTs to all other foods were negative. Roe-specific IgE levels were elevated in the serum samples of both patients. Immunoblotting revealed different IgE-binding patterns of the extracts and different inhibition profiles with the serum samples. In WFR blotting, both serum samples detected a heavy IgE-binding band at approximately 20 kDa, which was not inhibited with fish. Cross-reactivity between the roes was demonstrated in the patient with local symptoms from RTR but not in the patient with anaphylaxis from WFR. The first serum sample also detected several IgE-binding bands in the RTR blot, the most intensive at 21 to 23 kDa and 30 kDa, which were partially inhibited by WFR and more completely with fish. The anaphylaxis patient did not detect allergens in the RTR blot. After the investigation, the patients have remained symptom free and able to consume all kinds of fish without problems. IgE-mediated allergy to roe is possible without concomitant fish allergy. Roe allergy should be explored in patients who test negative to fish but are suspected of having seafood-related allergy.

Erythrocyte migration and gap formation in rabbit blood clots in vitro.

PubMed

Ueki, T; Yazama, F; Horiuchi, T; Yamada, M

2008-04-01

Thrombolytic agents must be carried by the blood circulation to thrombi to exert their functions. Structural gaps exist between blood vessels and thrombi or in the area surrounding thrombi. Therefore, information about fundamental gap formation at thrombotic areas is critically important for thrombolytic therapy. We previously reported that t-PA accelerates the activities of bovine erythrocytes and hemoglobin (Hb) towards bovine plasminogen activation. Here, we examined gap generation by observing morphological changes during thrombolytic processes in rabbit blood clots deformation of erythrocytes from blood clots and Hb transfer from erythrocytes to serum in vitro. Rabbit venous blood samples (1 ml) were stored under sterile conditions in glass tubes at 37 degrees C for 2, 24, 48 h, 1, and 2 weeks. We examined clot diameter, erythrocyte diameter and number as well as Hb volume in the serum, as well as histological changes in the clots. The diameter of blood clots did not change until 2 weeks after sampling. Erythrocyte diameter decreased within 48 h and at 2 weeks after sampling at the clot surface (p < 0.001) and interior (p < 0.001). The number of erythrocytes in the serum started to increase starting from 24 h after sampling (p < 0.01). Serum Hb volume also gradually increased from 24 h until 2 weeks after sampling (p < 0.01). The erythrocyte envelope became disrupted and cytoplasm started to flow through pores into the serum at 24 h. The results indicated that blood clots are reduced due to clot retraction, erythrocyte dissociation and cytoplasm leakage without a distinct fibrinolytic reaction. These results indicated that gaps start to form between 2 and 24 h after blood clotting.

Analysis of Serum Total and Free PSA Using Immunoaffinity Depletion Coupled to SRM: Correlation with Clinical Immunoassay Tests

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Tao; Hossain, Mahmud; Schepmoes, Athena A.

2012-08-03

Sandwich immunoassay is the standard technique used in clinical labs for quantifying protein biomarkers for disease detection, monitoring and therapeutic intervention. Albeit highly sensitive, the development of a specific immunoassay is rather time-consuming and associated with extremely high cost due to the requirement for paired immunoaffinity reagents of high specificity. Recently, mass spectrometry-based methods, specifically selected reaction monitoring mass spectrometry (SRM-MS), have been increasingly applied to measure low abundance biomarker candidates in tissue and biofluids, owing to high sensitivity and specificity, simplicity of assay configuration, and great multiplexing capability. In this study, we report for the first time the developmentmore » of immunoaffinity depletion-based workflows and SRM-MS assays that enable sensitive and accurate quantification of total and free prostate-specific antigen (PSA) in serum without the requirement for specific PSA antibodies. With stable isotope dilution and external calibration, low ng/mL level detection of both total and free PSA was consistently achieved in both PSA-spiked female serum samples and actual patient serum samples. Moreover, comparison of the results obtained when SRM PSA assays and conventional immunoassays were applied to the same samples showed very good correlation (R2 values ranging from 0.90 to 0.99) in several independent clinical serum sample sets, including a set of 33 samples assayed in a blinded test. These results demonstrate that the workflows and SRM assays developed here provide an attractive alternative for reliably measuring total and free PSA in human blood. Furthermore, simultaneous measurement of free and total PSA and many other biomarkers can be performed in a single analysis using high-resolution liquid chromatographic separation coupled with SRM-MS.« less

Prevalence of Parvovirus B19 and Parvovirus V9 DNA and Antibodies in Paired Bone Marrow and Serum Samples from Healthy Individuals

PubMed Central

Heegaard, Erik D.; Petersen, Bodil Laub; Heilmann, Carsten J.; Hornsleth, Allan

2002-01-01

Parvovirus B19 (hereafter referred to as B19) exhibits a marked tropism to human bone marrow (BM), and infection may lead to erythema infectiosum, arthropathy, hydrops fetalis, and various hematologic disorders. Recently, a distinct parvovirus isolate termed V9 with an unknown clinical spectrum was discovered. In contrast to the many studies of B19 serology and viremia, valid information on the frequency of B19 or V9 DNA in the BM of healthy individuals is limited. To develop a reference value, paired BM and serum samples from healthy subjects were tested for the presence of B19 and V9 DNA and specific antibodies. Immunoglobulin M (IgM) was not found in any of the serum samples. The prevalence of IgG showed a gradual and steady increase from 37% in children aged 1 to 5 years to 87% in people aged >50 years. When 190 well-characterized subjects were examined, B19 DNA was detected in the BM of 4 individuals (2.1%; 95% confidence interval, 0.58 to 5.3%) while none of the paired serum samples showed evidence of circulating viral DNA. V9 DNA was not found in any of the BM or serum samples. The finding of B19 DNA probably indicated a primary infection in one 7-year-old individual and reinfection or reactivation of persistent infection in the remaining three persons, aged 47 to 58 years. Serving as a benchmark for future studies, these findings are useful when interpreting epidemiologic data, performing BM transplantation, or considering clinical implications of parvovirus infection. PMID:11880419

Prevalence of parvovirus B19 and parvovirus V9 DNA and antibodies in paired bone marrow and serum samples from healthy individuals.

PubMed

Heegaard, Erik D; Petersen, Bodil Laub; Heilmann, Carsten J; Hornsleth, Allan

2002-03-01

Parvovirus B19 (hereafter referred to as B19) exhibits a marked tropism to human bone marrow (BM), and infection may lead to erythema infectiosum, arthropathy, hydrops fetalis, and various hematologic disorders. Recently, a distinct parvovirus isolate termed V9 with an unknown clinical spectrum was discovered. In contrast to the many studies of B19 serology and viremia, valid information on the frequency of B19 or V9 DNA in the BM of healthy individuals is limited. To develop a reference value, paired BM and serum samples from healthy subjects were tested for the presence of B19 and V9 DNA and specific antibodies. Immunoglobulin M (IgM) was not found in any of the serum samples. The prevalence of IgG showed a gradual and steady increase from 37% in children aged 1 to 5 years to 87% in people aged >50 years. When 190 well-characterized subjects were examined, B19 DNA was detected in the BM of 4 individuals (2.1%; 95% confidence interval, 0.58 to 5.3%) while none of the paired serum samples showed evidence of circulating viral DNA. V9 DNA was not found in any of the BM or serum samples. The finding of B19 DNA probably indicated a primary infection in one 7-year-old individual and reinfection or reactivation of persistent infection in the remaining three persons, aged 47 to 58 years. Serving as a benchmark for future studies, these findings are useful when interpreting epidemiologic data, performing BM transplantation, or considering clinical implications of parvovirus infection.

Simple and rapid quantification of vancomycin in serum, urine and peritoneal/pleural effusion via UHPLC-MS/MS applicable to personalized antibiotic dosing research.

PubMed

Javorska, Lenka; Krcmova, Lenka Kujovska; Solich, Petr; Kaska, Milan

2017-08-05

Management of the therapy of life-threatening bacterial infection is extremely based on an optimal antibiotic treatment. Achieving the correct vancomycin dosage in blood and target tissues can be complicated in special situations, e.g., where large fluid sequestration and/or acute renal failure occur. A UHPLC-MS/MS method operating in electrospray (ESI) positive ion mode was applied for the determination of vancomycin in serum, urine and peritoneal/pleural effusion. Sample pretreatment was composed of dilution and simple protein precipitation where only a small volume (50μL) of serum, urine or peritoneal/pleural effusion was required. The separation of vancomycin was performed on a Meteoric Core C18 BIO column (100×4.6mm, 2.7μm) by gradient elution with 0.1% formic acid in water and acetonitrile. The total time of analysis was 4.5min. The method was found to be linear in the range of 2-60μM (or 0.5-10μM) for serum, 0.27-10μM (or 2-60μM) for peritoneal/pleural effusion and 25-300μM for urine, which was adequate for the determination of vancomycin in patient samples. The intra- and inter-day precision was below 8% RSD, and accuracy was from 89 to 104%. The UHPLC/MS-MS method offers a fast and reliable approach to determine vancomycin concentrations in three different human body fluid samples (serum, urine and peritoneal/pleural effusion) with a simple sample pretreatment that was the same for all selected specimens. This method should be applicable to large sample series in clinical (pharmacokinetic/pharmacodynamic) studies. Copyright © 2017 Elsevier B.V. All rights reserved.

Anti-Saccharomyces cerevisiae and antineutrophil cytoplasmic antibodies as predictors of inflammatory bowel disease.

PubMed

Israeli, E; Grotto, I; Gilburd, B; Balicer, R D; Goldin, E; Wiik, A; Shoenfeld, Y

2005-09-01

Several antibodies have been reported in the sera of patients with Crohn's disease (CD) and ulcerative colitis (UC). The most commonly described are anti-Saccharomyces cerevisiae mannan antibodies (ASCA) in CD and perinuclear antineutrophil cytoplasm antibodies (pANCA) in UC. Familial clustering of these antibodies has been described, suggesting they might be genetic markers. Our aim was to investigate the presence of these antibodies before the emergence of overt clinical manifestations. Since 1980, the Israeli Defense Force (IDF) Medical Corps Serum Repository has stored serum samples obtained systematically from 5% of all recruits on enlistment, and from the same population on discharge from compulsory military service. We evaluated serum samples obtained from 32 subjects with CD and eight with UC before they were clinically diagnosed, along with samples from matched controls. ASCA were present in 10/32 (31.3%) CD patients before clinical diagnosis compared with 0/95 (0%) controls (p<0.001). None of the eight patients with serum samples available before diagnosis of UC were ASCA positive. ASCA was positive in 54.5% of patients after diagnosis of CD. The mean interval between ASCA detection and diagnosis was 38 months. In 90% of patients, antibodies were detected in the first available serum sample; therefore, measurements of the average time from the presence of ASCA to diagnosis may be even longer. pANCA were present in 2/8 (25%) patients with available sera before the diagnosis of UC. None of their 24 matched controls were positive (p = 0.014). ASCA and pANCA may predict development of inflammatory bowel disease years before the disease is clinically diagnosed.

Schoolchildren with Learning Difficulties Have Low Iron Status and High Anemia Prevalence

PubMed Central

Arcanjo, C. P. C.; Santos, P. R.

2016-01-01

Background. In developing countries there is high prevalence of iron deficiency anemia, which reduces cognitive performance, work performance, and endurance; it also causes learning difficulties and negative impact on development for infant population. Methods. The study concerns a case-control study; data was collected from an appropriate sample consisting of schoolchildren aged 8 years. The sample was divided into two subgroups: those with deficient initial reading skills (DIRS) (case) and those without (control). Blood samples were taken to analyze hemoglobin and serum ferritin levels. These results were then used to compare the two groups with Student's t-test. Association between DIRS and anemia was analyzed using odds ratio (OR). Results. Hemoglobin and serum ferritin levels of schoolchildren with DIRS were statistically lower when compared to those without, hemoglobin p = 0.02 and serum ferritin p = 0.04. DIRS was statistically associated with a risk of anemia with a weighted OR of 1.62. Conclusions. In this study, schoolchildren with DIRS had lower hemoglobin and serum ferritin levels when compared to those without. PMID:27703806

Schoolchildren with Learning Difficulties Have Low Iron Status and High Anemia Prevalence.

PubMed

Arcanjo, F P N; Arcanjo, C P C; Santos, P R

2016-01-01

Background . In developing countries there is high prevalence of iron deficiency anemia, which reduces cognitive performance, work performance, and endurance; it also causes learning difficulties and negative impact on development for infant population. Methods . The study concerns a case-control study; data was collected from an appropriate sample consisting of schoolchildren aged 8 years. The sample was divided into two subgroups: those with deficient initial reading skills (DIRS) (case) and those without (control). Blood samples were taken to analyze hemoglobin and serum ferritin levels. These results were then used to compare the two groups with Student's t -test. Association between DIRS and anemia was analyzed using odds ratio (OR). Results . Hemoglobin and serum ferritin levels of schoolchildren with DIRS were statistically lower when compared to those without, hemoglobin p = 0.02 and serum ferritin p = 0.04. DIRS was statistically associated with a risk of anemia with a weighted OR of 1.62. Conclusions . In this study, schoolchildren with DIRS had lower hemoglobin and serum ferritin levels when compared to those without.

Fecal corticosterone reflects serum corticosterone in Florida sandhill cranes.

PubMed

Ludders, J W; Langenberg, J A; Czekala, N M; Erb, H N

2001-07-01

Florida sandhill cranes (Grus canadensis pratensis) were conditioned to confinement 6 hr/day for 7 days. On day 8, each bird's jugular vein was catheterized, blood samples were drawn, and each crane was confined for 6 hr. Using a randomized, restricted cross-over design, cranes were injected intravenously with either 0.9% NaCl solution or ACTH (cosyntropin; Cortrosyn; 0.25 mg). During the 6 hr of confinement, fecal samples (feces and urine) were collected from each of five cranes immediately after defecation. Individual fecal samples were collected approximately at hourly intervals and assayed for corticosterone. We showed previously that serum corticosterone did not vary significantly following saline injection, but peaked significantly 60 min after ACTH injection. Maximal fecal corticosterone concentrations (ng/g) were greater (P < 0.10; median 1087 ng/g) following ACTH stimulation compared to maximal fecal corticosterone concentrations at the end of acclimation (day 7; median 176) and following saline treatment (median 541). In cranes under controlled conditions, fecal corticosterone concentration reflects serum corticosterone levels, fecal corticosterone, Grus canadensis pratensis, sandhill cranes, serum corticosterone levels.

[Metabolism of rat liver in the electrostatic field and in the faraday cage before and after hepatectomy (author's transl)].

PubMed

Klingenberg, H G; Möse, J R; Fischer, G; Porta, J; Sadjak, A

1975-10-01

Investigations were performed with the aim of establishing the influence of various environmental conditions (such as steady field conditions, climatized laboratories, Faraday's cage) on a number of enzymic activities in the rat (including glutamic oxaloacetic tic transaminase, glutamic pyruvic transaminase, lactic dehydrogenase, gamma-glutamyl transpeptidase, acid phosphatase), as well as the serum concentrations of triglycerides, the oxygen consumption of hepatic parenchyma cells, and the influence on the incorporation of 3H-thymidine (following partial hepatectomy). In the steady field, the activities of the cytoplasmic enzymes (GOT, GPT, LDH) were higher then under Faraday conditions. The same applies both to the hepatic oxygen consumption and to the neutral fat serum levels. The control values always remained within the range of the results obtained under steady field or Faraday conditions. In the structure-linked enzymes (gamma-glutamyl transpeptidase, acid phosphatase) the results were not uniform. Following partial hepatectomy, and under steady field conditions, the serum triglyceride concentrations showed a less pronounced drop than they did in the controls. Under selected environmental conditions, the results obtained lie within the physiological range. The present findings, therefore, do not permit definite conclusions to be drawn on favourable or unfavourable effects exerted by the different types of electroclimates.

Nucleic Acid-Based Cross-Linking Assay for Detection and Quantification of Hepatitis B Virus DNA

PubMed Central

Lai, Vicky C. H.; Guan, Richard; Wood, Michael L.; Lo, Su Kong; Yuen, Man-Fung; Lai, Ching-Lung

1999-01-01

A nucleic acid photo-cross-linking technology was used to develop a direct assay for the quantification of hepatitis B virus (HBV) DNA levels in serum. Cross-linker-modified DNA probes complementary to the viral genomes of the major HBV subtypes were synthesized and used in an assay that could be completed in less than 6 h. The quantification range of the assay, as determined by testing serial dilutions of Eurohep HBV reference standards and cloned HBV DNA, was 5 × 105 to 3 × 109 molecules of HBV DNA/ml of serum. Within-run and between-run coefficients of variation (CVs) for the assay were 4.3 and 4.0%, respectively. The assay was used to determine HBV DNA levels in 302 serum samples, and the results were compared to those obtained after testing the same samples with the Chiron branched-DNA (bDNA) assay for HBV DNA. Of the samples tested, 218 were positive for HBV DNA by both assays and 72 gave results below the cutoff for both assays. Of the remaining 12 samples, 10 were positive for HBV DNA by the cross-linking assay only; the 2 other samples were positive by the bDNA assay only. Twenty-eight samples had to be retested by the bDNA assay (CV, >20% between the results obtained from the testing of each sample in duplicate), whereas only three samples required retesting by the cross-linking assay. The correlation between the HBV DNA levels, as measured by the two tests, was very high (r = 0.902; P = 0.01). We conclude that the cross-linking assay is a sensitive and reproducible method for the detection and quantification of HBV DNA levels in serum. PMID:9854083

Serological evidence for hepatitis e virus infection in laboratory monkeys and pigs in animal facilities in Japan.

PubMed

Yamamoto, Hiroshi; Li, Tian-Cheng; Koshimoto, Chihiro; Ito, Kaori; Kita, Masakazu; Miyashita, Nobumoto; Arikawa, Jiro; Yagami, Kenichi; Asano, Masahide; Tezuka, Hideo; Suzuki, Noboru; Kurosawa, Tsutomu; Shibahara, Toshiyuki; Furuya, Masato; Mohri, Shirou; Sato, Hiroshi; Ohsawa, Kazutaka; Ibuki, Kentaro; Takeda, Naokazu

2008-07-01

In laboratory animal facilities, monkeys and pigs are used for animal experiments, but the details of hepatitis E virus (HEV) infection in these animals are unknown. The risk of infection from laboratory animals to humans has become a concern; therefore, much attention should be paid to the handling of these animals during their care and use, including surgical procedures performed on infected animals. In this connection, serum samples collected from 916 monkeys and 77 pigs kept in 23 animal facilities belonging to the Japanese Association of Laboratory Animal Facilities of National University Corporations (JALAN) and the Japanese Association of Laboratory Animal Facilities of Public and Private Universities (JALAP) in Japan were examined for the purpose of detecting antibodies to HEV and HEV RNA by using ELISA and RT-PCR, respectively. One hundred and seven serum samples of 916 (11.7%) monkeys were positive for anti-HEV IgG, and 7 and 17 serum samples of 916 (0.8% and 5.3%) monkeys were positive for anti-HEV IgM and IgA, respectively. Thirty-six samples from 62 (58.1%) farm pigs were positive for anti-HEV IgG, whereas all samples tested from miniature pigs were negative (0/15, 0%). Seven samples from 62 (9.1%) farm pigs and 7 samples from 916 (0.8%) monkeys were positive for IgM antibody, but these HEV-IgM antibody positive serum samples were HEV-RNA negative by RT-PCR. The IgM antibody positive rate (9.1%) of farm pigs was much higher than that of monkeys (0.8%). These results suggest the relative levels of risk of HEV infection from these animals to animal handlers and researchers who work with them in laboratory animal facilities.

Stability of selected serum hormones and lipids after long-term storage in the Janus Serum Bank.

PubMed

Gislefoss, Randi E; Grimsrud, Tom K; Mørkrid, Lars

2015-04-01

The potential value of a biobank depends on the quality of the samples, i.e. how well they reflect the biological or biochemical state of the donors at the time of sampling. Documentation of sample quality has become a particularly important issue for researchers and users of biobank studies. The aim of this study was to investigate the long-term stability of selected components: cholesterol, high density cholesterol (HDLC), low density cholesterol (LDLC), apolipoprotein A1 (apo-A1), apolipoprotein B (apo B), follicle stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL), thyroid stimulating hormone (TSH) and free thyroxin (FT4). Samples, stored at -25°C, from 520 men aged 40-49 years at blood sampling distributed in equally sized groups (n=130) according to length of storage, 0, 4, 17 and 29 years, respectively, were used in a cross sectional design. The freshly collected serum samples were used as a reference group to calculate storage related changes. The differences between fresh samples and samples stored for 29 years were substantial for apo-A1 (+12%), apo-B (+22.3%), HDLC (-69.2%), LDLC (+31.3%), and PRL (-33.5%), while total cholesterol, FSH, LH, TSH and FT4 did not show any significant difference. The study showed large differences in serum level of the selected components. The lipids and apolipoproteins were all changed except for total cholesterol. Most hormones investigated (FSH, LH, TSH and FT4) proved to be stable after 29 years of storage while PRL showed sign of degradation. The observed differences are probably due to long-term storage effects and/or external factors (i.e. diet and smoking). Copyright © 2014 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Identification of New Biomarkers of Low-Dose GH Replacement Therapy in GH-Deficient Patients

PubMed Central

Cruz-Topete, Diana; Jorgensen, Jens Otto L.; Christensen, Britt; Sackmann-Sala, Lucila; Krusenstjerna-Hafstrøm, Thomas; Jara, Adam; Okada, Shigeru

2011-01-01

Context: GH secretion peaks at puberty and continues to be secreted in adulthood, albeit at a declining rate. Profound GH deficiency (GHD) in adults with pituitary disease is associated with symptoms that improve with GH substitution, but it is important to tailor the GH dose to avoid overtreatment. Measurement of serum IGF-I levels is an important clinical tool in this regard, but it is well recognized that some patients receiving GH treatment do not show an increase in IGF-I. Objective: The objective of the study was to identify novel serum biomarkers of GH treatment in adults with GHD. Design and Patients: Eight patients with profound GHD as a consequence of a pituitary adenoma or its treatment were evaluated before and 3 months after GH replacement therapy (0.2–0.4 mg/d). Main Outcome Measures: Serum proteomic changes were studied using two-dimensional gel electrophoresis and mass spectrometry. Protein profiles were analyzed and compared in serum samples obtained before and after GH treatment. Results: The levels of six serum protein spots were significantly altered after GH substitution. These proteins were identified as five isoforms of haptoglobin (decreased in posttreatment samples) and one isoform of apolipoprotein A-I (increased in posttreatment samples). Importantly, changes in the levels of the identified proteins were associated with decreases in fat mass and increases in lean mass in all patients. These results were independent of serum IGF-I levels. Conclusions: Evaluation of the identified proteins provides a novel alternative to traditional markers of GH status, such as serum IGF-I levels, to assess GH therapy in GH deficient adults. PMID:21543428

Serum concentrations of pituitary and adrenal hormones in female pigs exposed to two photoperiods.

PubMed

Kraeling, R R; Rampacek, G B; Mabry, J W; Cunningham, F L; Pinkert, C A

1983-11-01

Serum concentrations of pituitary and adrenal hormones were determined in lactating sows and ovariectomized (OVX) gilts exposed to 8 h (8L:16D) or 16 h of light (16L:8D). In addition serum prolactin (PRL) concentrations were determined after a thyrotropin releasing hormone (TRH) challenge. At 103 +/- 2 d of gestation or 3 wk after ovariectomy of nulliparous gilts on d 7 to 9 of the estrous cycle (d - 10), blood samples were collected from jugular vein cannulae at 30-min intervals for 8 h beginning at 0800 h. Immediately after the last sample, 13 sows and five OVX gilts were assigned to 8L:16D and 14 sows and five OVX gilts were assigned to 16L:8D/d and placed in two identical chambers in the farrowing house. Blood sampling was repeated on d 7, 14 and 21 of lactation in the sows and on d 7, 14, 21 and 28 in the OVX gilts. In Exp. 1, serum cortisol (C) concentrations were similar for sows exposed to 8L:16D (n = 7) and 16L:8D (n = 6) treatments, whereas in Exp. 2, serum C concentrations for sows exposed to 8L:16D (n = 6) were lower than those exposed to 16L:8D (n = 6) on d 7, 14 and 21. Photoperiod failed to influence serum concentrations of PRL, luteinizing hormone (LH) and growth hormone in the lactating sows or PRL in the OVX gilts. Photoperiod also failed to affect mean basal serum concentrations, peak height and peak frequency for PRL and LH in the lactating sows or for PRL in the OVX gilts.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation and comparison of Abbott Jaffe and enzymatic creatinine methods: Could the old method meet the new requirements?

PubMed

Küme, Tuncay; Sağlam, Barıs; Ergon, Cem; Sisman, Ali Rıza

2018-01-01

The aim of this study is to evaluate and compare the analytical performance characteristics of the two creatinine methods based on the Jaffe and enzymatic methods. Two original creatinine methods, Jaffe and enzymatic, were evaluated on Architect c16000 automated analyzer via limit of detection (LOD) and limit of quantitation (LOQ), linearity, intra-assay and inter-assay precision, and comparability in serum and urine samples. The method comparison and bias estimation using patient samples according to CLSI guideline were performed on 230 serum and 141 urine samples by analyzing on the same auto-analyzer. The LODs were determined as 0.1 mg/dL for both serum methods and as 0.25 and 0.07 mg/dL for the Jaffe and the enzymatic urine method respectively. The LOQs were similar with 0.05 mg/dL value for both serum methods, and enzymatic urine method had a lower LOQ than Jaffe urine method, values at 0.5 and 2 mg/dL respectively. Both methods were linear up to 65 mg/dL for serum and 260 mg/dL for urine. The intra-assay and inter-assay precision data were under desirable levels in both methods. The higher correlations were determined between two methods in serum and urine (r=.9994, r=.9998 respectively). On the other hand, Jaffe method gave the higher creatinine results than enzymatic method, especially at the low concentrations in both serum and urine. Both Jaffe and enzymatic methods were found to meet the analytical performance requirements in routine use. However, enzymatic method was found to have better performance in low creatinine levels. © 2017 Wiley Periodicals, Inc.

A microbiological method for determining serum levels of broad spectrum β-lactam antibiotics in critically ill patients.

PubMed

Fridlund, Jimmy; Woksepp, Hanna; Schön, Thomas

2016-10-01

Recent studies show that suboptimal blood levels of β-lactam antibiotics are present in intensive care unit (ICU) patients. A common reference method for assessing drug concentrations is liquid chromatography coupled with mass-spectrometry (LC-MS) which is highly accurate but rarely available outside reference centres. Thus, our aim was to develop a microbiological method for monitoring β-lactam antibiotic serum levels which could be used at any hospital with a microbiological laboratory. The method was developed as a 96-well broth microdilution format to assess the concentrations of cefotaxime (CTX), meropenem (MER), and piperacillin (PIP). Patient serum containing antibiotics were diluted in suspensions of bacteria with known minimal inhibitory concentrations (MICs). Serum antibiotic concentrations were calculated by dividing the MIC with the dilution factor at which the serum inhibited growth of the bacterial suspension. Serum (n=88) from ICU patients at four hospitals in south-east Sweden were analysed and compared to LC-MS analysis. The overall accuracy and precision for spiked samples and patient samples was within the pre-set target of ±20.0% for all drugs. There was a significant correlation between the microbiological assay and LC-MS for the patient samples (CTX: r=0.86, n=31; MER: r=0.96, n=11; PIP: r=0.88, n=39) and the agreement around the clinical cut-off for CTX (4.0mg/l), MER (2.0mg/l) and PIP (16.0mg/l) was 90%, 100% and 87%, respectively. The microbiological method has a performance for determination of serum levels of meropenem, piperacillin and cefotaxime suitable for clinical use. It is an inexpensive method applicable in any microbiology laboratory. Copyright © 2016 Elsevier B.V. All rights reserved.

Supercritical fluid extraction of selected pharmaceuticals from water and serum.

PubMed

Simmons, B R; Stewart, J T

1997-01-24

Selected drugs from benzodiazepine, anabolic agent and non-steroidal anti-inflammatory drug (NSAID) therapeutic classes were extracted from water and serum using a supercritical CO2 mobile phase. The samples were extracted at a pump pressure of 329 MPa, an extraction chamber temperature of 45 degrees C, and a restrictor temperature of 60 degrees C. The static extraction time for all samples was 2.5 min and the dynamic extraction time ranged from 5 to 20 min. The analytes were collected in appropriate solvent traps and assayed by modified literature HPLC procedures. Analyte recoveries were calculated based on peak height measurements of extracted vs. unextracted analyte. The recovery of the benzodiazepines ranged from 80 to 98% in water and from 75 to 94% in serum. Anabolic drug recoveries from water and serum ranged from 67 to 100% and 70 to 100%, respectively. The NSAIDs were recovered from water in the 76 to 97% range and in the 76 to 100% range from serum. Accuracy, precision and endogenous peak interference, if any, were determined for blank and spiked serum extractions and compared with classical sample preparation techniques of liquid-liquid and solid-phase extraction reported in the literature. For the benzodiazepines, accuracy and precision for supercritical fluid extraction (SFE) ranged from 1.95 to 3.31 and 0.57 to 1.25%, respectively (n = 3). The SFE accuracy and precision data for the anabolic agents ranged from 4.03 to 7.84 and 0.66 to 2.78%, respectively (n = 3). The accuracy and precision data reported for the SFE of the NSAIDs ranged from 2.79 to 3.79 and 0.33 to 1.27%, respectively (n = 3). The precision of the SFE method from serum was shown to be comparable to the precision obtained with other classical preparation techniques.

Development and analytical validation of an enzyme linked immunosorbent assay for the measurement of canine gastric lipase immunoreactivity in serum

PubMed Central

2004-01-01

Abstract The objective of this study was to develop and analytically validate an enzyme linked immunosorbent assay (ELISA) for measurement of canine gastric lipase immunoreactivity (cGLI). A sandwich ELISA was developed using canine gastric lipase (cGL) purified from canine stomachs and polyclonal antibodies directed against cGL, raised in rabbits and purified by affinity chromatography. The assay was validated by determination of sensitivity, working range, linearity, accuracy, precision, reproducibility, and the upper limit of the control range by determining the 97.5th percentile of serum cGLI concentration in 74 healthy canines. Sensitivity and working range in serum were 200 ng/L and 200 to 39 160 ng/L, respectively. Observed to expected ratios for dilutional parallelism for 3 serum samples and 3 dilutions ranged from 86.1% to 244.2% (mean ± standard deviation [s]; 125.4% ± 48.2%). Observed to expected ratios for spiking recoveries for 3 serum samples and 6 spiking concentrations ranged from 66.4% to 152.5% (mean ± s; 104.5% ± 22.9%). Intra-assay and interassay variabilities for 3 different serum samples were 25.5%, 9.4%, and 13.4% and 26.0%, 17.2%, and 14.4%, respectively. The upper limit of the control range for serum cGLI was 662 ng/L. We concluded that the ELISA for cGLI described here is highly sensitive and shows a wide working range. However, the validation characteristics for this assay are suboptimal and below values of approximately 2.000 ng/L the assay is more semiquantitative in nature. Despite its limitations, whether this assay is useful for the diagnosis of canine gastric disorders remains to be determined. PMID:15352539

Association of serum ferritin with metabolic syndrome and diabetes mellitus in the South Korean general population according to the Korean National Health and Nutrition Examination Survey 2008.

PubMed

Lee, Byung-Kook; Kim, Yangho; Kim, Young-Il

2011-10-01

We examined the association of serum ferritin levels with metabolic syndrome (MS) and diabetes mellitus in a representative sample of the adult South Korean population using data from the 2008 Korean National Health and Nutrition Examination Survey. We conducted a cross-sectional study of 6311 adults older than 20 years who participated in the 2008 Korean National Health and Nutrition Examination Survey. Metabolic syndrome was defined as the presence of at least 3 of the following: elevated blood pressure, low high-density lipoprotein cholesterol, elevated serum triglycerides, elevated plasma glucose, and abdominal obesity. Diabetes mellitus was defined as fasting glucose of at least 126 mg/dL. Insulin resistance was determined using the homeostasis model assessment estimate of insulin resistance. In a representative sample of the adult Korean population, MS was more prevalent in the highest quartile compared with the lowest quartile of serum ferritin concentrations in women following adjustments for age, education, smoking, alcohol intake, body mass index, aspartate aminotransferase, and alanine aminotransferase. Diabetes mellitus was more prevalent in the highest quartile compared with the lowest quartile of serum ferritin concentrations in premenopausal women and men. The geometric means of fasting insulin and insulin resistance determined using the homeostasis model assessment of insulin resistance in the fourth serum ferritin quartiles of postmenopausal women and men were significantly higher compared with those in the first quartile of the respective groups. The present study demonstrates that elevated serum ferritin concentrations are associated with an increased risk of MS and diabetes mellitus in a representative sample of the adult South Korean population. Copyright © 2011. Published by Elsevier Inc.

Changes in Peripheral Serum Creatine Phosphokinase (CPK) and Lactic Dehydrogenase (LDH) in Acute Experimental Colonic Infarction

PubMed Central

Graeber, Geoffrey M.; Wukich, Dane K.; Cafferty, Patrick J.; O'Neill, John F.; Wolf, Robert E.; Ackerman, Norman B.; Harmon, John W.

1981-01-01

No satisfactory laboratory test for the early diagnosis of bowel infarction exists at this time. We have delineated changes in serum CPK levels after acute superior mesenteric artery infarction; whether or not comparable changes occur with inferior mesenteric artery infarction has not yet been determined. Furthermore, the changes in LDH associated with acute bowel infarction have not been documented. To determine the changes in serum CPK and LDH in acute colonic infarction, laparotomies were performed on dogs after peripheral baseline blood samples were drawn and each subject was randomly placed in one of three groups: laparotomy alone, acute colonic obstruction, and acute colonic infarction by ligation of the inferior mesenteric artery. The marginal artery of the colon was ligated at the peritoneal reflection and at the cecum to interrupt arterial collaterals. Blood samples were taken from each subject at intervals of three hours for 48 hours after injury. Serum from each sample was analyzed for total CPK and LDH by automated spectrophotometry. Isoenzymes were determined by agarose gel electrophoresis. Necropsies were conducted on all the dogs to confirm that the intended condition had been produced and that no intercurrent disease was present. The data support the conclusion that total CPK, total LDH and their isoenzymes become elevated in the peripheral serum after colonic infarction. The maximal elevations were all seen within the first 12 hours after acute colonic infarction. Total LDH and LDH3, the most prevalent isoenzyme of LDH in bowel, do not become elevated in the serum to as high a level as CPK, but the combination of serum elevations in both enzyme systems may prove to be of diagnostic significance. PMID:7305484

Introduction of a rapid, simple radioimmunoassay and quality control scheme for thyroxine.

PubMed Central

Nye, L; Hassan, M; Willmott, E; Landon, J

1976-01-01

A simple radioimmunoassay has been developed for service purposes to determine serum total thyroxine levels. Only three additions are required, of standard or sample, labelled thyroxine and antibody in polyethylene glycol. After 2 hours' incubation at room temperature the antibody-bound and free fractions are separated by centrifugation. Serum total thyroxine levels were measured in 195 euthyroid subjects and it was established that normal values lay within the range 57 to 155 nmol/1. Serial blood samples taken over a 24-hour period, from 11 subjects, indicated that there was no circadian rhythm so that samples for total thyroxine assay can be taken at any time of the day. Similar results were obtained using serum or plasma. Satisfactory results were obtained for three quality control sera when measured by seven different laboratories using this method. PMID:932232

Systems analysis of singly and multiply O-glycosylated peptides in the human serum glycoproteome via EThcD and HCD mass spectrometry.

PubMed

Zhang, Yong; Xie, Xinfang; Zhao, Xinyuan; Tian, Fang; Lv, Jicheng; Ying, Wantao; Qian, Xiaohong

2018-01-06

Human serum has been intensively studied to identify biomarkers via global proteomic analysis. The altered O-glycoproteome is associated with human pathological state including cancer, inflammatory and degenerative diseases and is an attractive source of disease biomarkers. Because of the microheterogeneity and macroheterogeneity of O-glycosylation, site-specific O-glycosylation analysis in human serum is still challenging. Here, we developed a systematic strategy that combined multiple enzyme digestion, multidimensional separation for sample preparation and high-resolution tandem MS with Byonic software for intact O-glycopeptide characterization. We demonstrated that multiple enzyme digestion or multidimensional separation can make sample preparation more efficient and that EThcD is not only suitable for the identification of singly O-glycosylated peptides (50.3%) but also doubly (21.2%) and triply (28.5%) O-glycosylated peptides. Totally, with the strict scoring criteria, 499 non-redundant intact O-glycopeptides, 173 O-glycosylation sites and 6 types of O-glycans originating from 49 O-glycoprotein groups were identified in human serum, including 121 novel O-glycosylation sites. Currently, this is the largest data set of site-specific native O-glycoproteome from human serum samples. We expect that the strategies developed by this study will facilitate in-depth analyses of native O-glycoproteomes in human serum and provide opportunities to understand the functional roles of protein O-glycosylation in human health and diseases. The altered O-glycoproteome is associated with human pathological state and is an attractive source of disease biomarkers. However, site-specific O-glycosylation analysis is challenging because of the microheterogeneity (different glycoforms attached to one glycosylation site) and macroheterogeneity (site occupancy) of O-glycosylation. In this work, we developed a systematic strategy for intact O-glycopeptide characterization. This study took advantage of the inherent properties of the new fragmentation method called EThcD, which provides more complete fragmentation information about O-glycosylated peptides and a more confident site localization of O-glycans than collision-induced dissociation (HCD). We demonstrated that multiple enzyme digestion or multidimensional separation can make sample preparation more efficient and that EThcD was not only suitable for the identification of singly O-glycosylated peptides (50.3%) but also doubly (21.2%) and triply (28.5%) O-glycosylated peptides. Finally, we got a largest data set of site-specific native O-glycoproteome from human serum samples. Furthermore, quantitative analysis of intact O-glycopeptides from the serum samples of IgA nephropathy (IgAN) patients and healthy donors was performed, and the results showed the potential of the strategy to discover O-glycosylation biomarkers. We expect that the strategies developed by this study will facilitate in-depth analyses of native O-glycoproteomes in human serum and lead to exciting opportunities to understand the functional roles of protein O-glycosylation in human health and diseases. Copyright © 2017. Published by Elsevier B.V.

Detection of cerebrospinal fluid leakage by specific measurement of transferrin glycoforms.

PubMed

Kwon, Seok-Joon; Zhang, Fuming; Dordick, Jonathan S; Sonstein, William J; Linhardt, Robert J

2015-10-01

A simple and rapid detection of cerebrospinal fluid (CSF) leakage would benefit spine surgeons making critical postoperative decisions on patient care. We have assessed novel approaches to selectively determine CSF β2-transferrin (β2TF), an asialo-transferrin (aTF) biomarker, without interference from serum sialo-transferrin (sTF) in test samples. First, we performed mild periodate oxidation to selectively generate aldehyde groups in sTF for capture with magnetic hydrazide microparticles, and selective removal with a magnetic separator. Using this protocol sTF was selectively removed from mixtures of CSF and serum containing CSF aTF (β2TF) and serum sTF, respectively. Second, a two-step enzymatic method was developed with neuraminidase and galactose oxidase for generating aldehyde groups in sTF present in CSF and serum mixtures for magnetic hydrazide microparticle capture. After selectively removing sTF from mixtures of CSF and serum, ELISA could detect significant TF signal only in CSF, while the TF signal in serum was negligible. The new approach for selective removal of only sTF in test samples will be promising for the required intervention by a spine surgeon. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Serum endocan levels before and after surgery on low-grade gliomas.

PubMed

Tanriverdi, Taner; Kemerdere, Rahsan; Inal, Berrin B; Yuksel, Odhan; Emre, Humeyra O; Ahmedov, Merdin; Baran, Oguz; Ates, Seda

2017-01-01

Endocan has been shown to be a marker for several cancers and may show degree of malignancy. The aim of this study is to assess serum levels of endocan before and after surgery on low-grade gliomas (LGGs). Endocan was assayed by commercially available enzyme-linked immunosorbent assay (ELISA) kits in a total of 19 patients and 12 controls. Serial serum samples were obtained before and after surgery (1 st day, 1 st week, and 1 st month of surgery). Control samples were collected from cord blood during cesarean section. The results were compared with control brain tissues. Controls showed significantly lower serum endocan levels compared to before and after surgery ( P < 0.05). There is a trend of increase in mean serum levels from before surgery and during the very early period after surgery (during first week); however, in the first month, mean serum levels became lower. Endocan, a vital molecule for angiogenesis, is highly expressed before and after surgery in LGGs, but long-term data is needed. Furthermore, future studies should include high-grade gliomas to discuss whether endocan is associated with recurrence and response to treatment.

Indigenous West Nile virus infections in horses in Albania.

PubMed

Berxholi, K; Ziegler, U; Rexhepi, A; Schmidt, K; Mertens, M; Korro, K; Cuko, A; Angenvoort, J; Groschup, M H

2013-11-01

Serum samples collected from 167 equines of 12 districts in Albania were tested for West Nile virus-specific antibodies by enzyme-linked immunosorbent assay and virus neutralization assay, using WNV lineage 1 and 2. In addition, 95 bird serum samples from Albania and 29 horse samples from Kosovo were tested in ELISA. An overall seroprevalence rate of 22% was found in horses from Albania, whereas no specific antibodies were found in the equine samples from Kosovo and the bird samples. This is the first report indicating WNV infections in animals in Albania, and the first reported seroprevalence study conducted for Kosovo. These results provide evidence for widespread infections of WNV in Albania. © 2013 Blackwell Verlag GmbH.

Impact of sample extraction on the accurate measurement of progesterone in human serum by liquid chromatography tandem mass spectrometry.

PubMed

Ke, Yuyong; Gonthier, Renaud; Labrie, Fernand

2017-05-01

In the present study, the impact of the extraction solvent on the accuracy of endogenous progesterone assay in human serum has been investigated using two selective reaction monitoring (SRM) transitions (315>97 & 315>109). Higher levels of noise and more interference were observed when more polar solvents were used for extraction, thus resulting in serious bias of the measured values of progesterone in serum. This is confirmed by monitoring the ion ratio of 315>97-315>109. This issue could not be easily resolved by changes in MS/MS transitions or chromatography conditions. More bias was observed with the SRM transition 315>109 for the polar solvent extraction. Hexane and 1-chlorobutane (polarity index of 0 and 1, respectively) did provide the cleanest samples with a lower noise level in the chromatograms. Moreover, the measured values of progesterone were not changed with different SRM transitions or longer retention time in search of an improved separation. Recovery tests of progesterone have been performed with 1-chlorobutane in matrices with phosphate buffered saline (PBS) 1x, PBS 1×3% bovine serum albumin (BSA), stripped serum/H 2 O (1:1) and unstripped serum. The recovery (70%∼80%) consistency is observed not only at different levels but also in different matrices. The equivalent recovery between PBS 1x, PBS 1×3% BSA and unstripped serum shows that the impact of progesterone binding to serum proteins on the measurement accuracy can be avoided with this sample preparation procedure. No significant matrix effect on the determination of progesterone was observed with 1-chlorobutane. Within the range of 12.5-2000pg/mL, a good linearity is observed with R>0.99 and weighting factor 1/X. Bias and covariance efficiency of QCs are within 10%. With 1-chlorobutane as the extraction solvent, the concentration of progesterone was measured where the range for postmenopausal serum is 5.74∼91.7pg/mL, which is well below the reported concentrations of 314 pg/mL∼942pg/mL in postmenopausal serum by immunoassay-based techniques, while the range in premenopausal serum is 12.8 pg/mL∼18.6ng/mL. Copyright © 2017 Elsevier B.V. All rights reserved.

A Rapid Dot Immunoassay for the Detection of Serum Antibodies to Eastern Equine Encephalomyelitis and Saint Louis Encephalitis Viruses in Sentinel Chickens

DTIC Science & Technology

1988-01-01

tion inhibition (HI) tests .6 uses a novel membrane, polyvinylydine difluor- EEE and SLE viruses are significant public ide (PVDF), and yields a...only 3 aged values between the two tests (Fig. 3). Vari- of 6 SLE virus-inoculated birds developed an- ability in EIA titer was observed with samples... SLE samples negative by EIA were location. Four serum samples from each of 3 also negative by DotIA. EEE-positive chickens were tested by EIA and A

Serum Ropivacaine Concentrations and Systemic Local Anesthetic Toxicity in Trauma Patients Receiving Long-Term Continuous Peripheral Nerve Block Catheters

DTIC Science & Technology

2010-02-01

concentrations were calculated from the following 2 methods: 1. Using peak area ratio of ropivacaine/ bupivacaine of samples versus peak area ratio of...ropivacaine/ bupivacaine of standards (0–3000 ng/mL) using the following formula: RopS (PA of Rop)/(PA of Bup)S /(PA of Rop)/(PA of Bup)STD Bup...1/mL serum used) where Rop ropivacaine, Bup bupivacaine , PA peak area, S sample, and STD standard. 2. Samples of ropivacaine concentrations

Use of cerebrospinal fluid and serum samples impregnated on FTATM Elute filter paper for the diagnosis of infections caused by Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae

PubMed Central

Gonçalves, Maria Gisele; Higa, Fábio Takenori; Castilho, Euclides Ayres; Ibarz-Pavón, Ana Belén; Sacchi, Claudio Tavares

2017-01-01

Background The lack of information regarding the burden of acute bacterial meningitis in Latin America leads to a reduction in the estimated incidence rates of the disease, and impairs public health decisions on the use and follow-up of preventive interventions, particularly, the evaluation of existing vaccination policies. The use of the real-time PCR in diagnostic routine procedures has resulted in a substantial increase in confirmed bacterial meningitis cases. However, in resource-poor countries, these assays are only available in reference laboratories. Sample transportation to these laboratories is a critical constraint, as it requires specialized, high cost courier services. To overcome this barrier we evaluated the use of FTATM Elute filter paper cards for the conservation and processing of samples under normal environmental conditions, as they would be when transported from remote and under-equipped healthcare facilities to the reference centers. A total of 401 samples received in 2015 as part of Sao Paulo’s national surveillance for routine diagnosis were selected for this study. Methods The sensitivity and specificity of real-time PCR were evaluated using fresh serum and cerebrospinal fluid (CSF) samples processed using our laboratory’s standard DNA extraction, and processing the same samples after being dried and stored on FTATM card, and DNA extracted following the manufacturer’s instructions. Results The sensitivities for detection of Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae from CSF dried and stored on FTATM cards were 98%, 92%, and 100%, respectively, and with serum samples were 73%, 88%, and 100%, respectively. When compared to our laboratory’s standard methodology, results showed high concordance, with Kappa index ranges of 0.9877–1.00 for CSF, and 0.8004–1.00 for serum samples. Conclusion The use of FTATM cards for CSF and serum conservation and transport represents a rapid, reliable, and cost-effective alternative that will allow obtaining valuable epidemiological information that would otherwise be lost. PMID:28235065

Analysis of biofluids in aqueous environment based on mid-infrared spectroscopy.

PubMed

Fabian, Heinz; Lasch, Peter; Naumann, Dieter

2005-01-01

In this study we describe a semiautomatic Fourier transform infrared spectroscopic methodology for the analysis of liquid serum samples, which combines simple sample introduction with high sample throughput. The applicability of this new infrared technology to the analysis of liquid serum samples from a cohort of cattle naturally infected with bovine spongiform encephalopathy and from controls was explored in comparison to the conventional approach based on transmission infrared spectroscopy of dried serum films. Artifical neural network analysis of the infrared data was performed to differentiate between bovine spongiform encephalopathy-negative controls and animals in the late stage of the disease. After training of artifical neural network classifiers, infrared spectra of sera from an independent external validation data set were analyzed. In this way, sensitivities between 90 and 96% and specificities between 84 and 92% were achieved, respectively, depending upon the strategy of data collection and data analysis. Based on these results, the advantages and limitations of the liquid sample technique and the dried film approach for routine analysis of biofluids are discussed. 2005 Society of Photo-Optical Instrumentation Engineers.

Virological Survey in free-ranging wildcats (Felis silvestris) and feral domestic cats in Portugal.

PubMed

Duarte, A; Fernandes, M; Santos, N; Tavares, L

2012-08-17

To determine the presence of viral pathogens in natural areas a survey was conducted on an opportunistic sample of fifty eight wild (Felis silvestris silvestris) and feral cats (F. s. catus). The biological materials included serum, lung tissue extract and stool. Feline leukemia virus p27 antigen was detected in 13/50 serum/lung tissue extract samples (26%), canine distemper virus antibodies were detected in 2/26 serum/lung tissue extract samples (7.7%), feline coronavirus RNA was present in 6/29 stool samples (20.7%) and feline parvovirus DNA in 2/29 stool samples (6.9%). Canine distemper virus RNA was not detected. Feline immunodeficiency virus and feline coronavirus antibodies were not detected. Evidence of exposure to feline leukemia virus, canine distemper virus, feline coronavirus and feline parvovirus was found in wild and feral cats raising the importance of performing a comprehensive survey to correctly evaluate the potential threat of infectious diseases to endangered species, namely to the wildcat and to the Iberian lynx, which is meant to be reintroduced after 2012 in Portugal. Copyright © 2012 Elsevier B.V. All rights reserved.

Multicommuted flow system for the determination of glucose in animal blood serum exploiting enzymatic reaction and chemiluminescence detection

PubMed Central

Pires, Cherrine K.; Martelli, Patrícia B.; Lima, José L. F. C.; Saraiva, Maria Lúcia M. F. S.

2003-01-01

An automatic flow procedure based on multicommutation dedicated for the determination of glucose in animal blood serum using glucose oxidase with chemiluminescence detection is described. The flow manifold consisted of a set of three-way solenoid valves assembled to implement multicommutation. A microcomputer furnished with an electronic interface and software written in Quick BASIC 4.5 controlled the manifold and performed data acquisition. Glucose oxidase was immobilized on porous silica beads (glass aminopropyl) and packed in a minicolumn (15 × 5 mm). The procedure was based on the enzymatic degradation of glucose, producing hydrogen peroxide, which oxidized luminol in the presence of hexacyanoferrate(III), causing the chemiluminescence. The system was tested by analysing a set of serum animal samples without previous treatment. Results were in agreement with those obtained with the conventional method (LABTEST Kit) at the 95% confidence level. The detection limit and variation coefficient were estimated as 12.0 mg l−1 (99.7% confidence level) and 3.5% (n = 20), respectively. The sampling rate was about 60 determinations h−1 with sample concentrations ranging from 50 to 600 mg l−1 glucose. The consumptions of serum sample, hexacyanoferrate(III) and luminol were 46 μl, 10.0 mg and 0.2 mg/determination, respectively. PMID:18924619

Detection of Wuchereria bancrofti DNA in paired serum and urine samples using polymerase chain reaction-based systems.

PubMed

Ximenes, Camila; Brandão, Eduardo; Oliveira, Paula; Rocha, Abraham; Rego, Tamisa; Medeiros, Rafael; Aguiar-Santos, Ana; Ferraz, João; Reis, Christian; Araujo, Paulo; Carvalho, Luiz; Melo, Fabio L

2014-12-01

The Global Program for the Elimination of Lymphatic Filariasis (GPELF) aims to eliminate this disease by the year 2020. However, the development of more specific and sensitive tests is important for the success of the GPELF. The present study aimed to standardise polymerase chain reaction (PCR)-based systems for the diagnosis of filariasis in serum and urine. Twenty paired biological urine and serum samples from individuals already known to be positive for Wuchereria bancrofti were collected during the day. Conventional PCR and semi-nested PCR assays were optimised. The detection limit of the technique for purified W. bancrofti DNA extracted from adult worms was 10 fg for the internal systems (WbF/Wb2) and 0.1 fg by using semi-nested PCR. The specificity of the primers was confirmed experimentally by amplification of 1 ng of purified genomic DNA from other species of parasites. Evaluation of the paired urine and serum samples by the semi-nested PCR technique indicated only two of the 20 tested individuals were positive, whereas the simple internal PCR system (WbF/Wb2), which has highly promising performance, revealed that all the patients were positive using both samples. This study successfully demonstrated the possibility of using the PCR technique on urine for the diagnosis of W. bancrofti infection.

Prevalence of hepatitis A viral RNA and antibodies among Chinese blood donors.

PubMed

Sun, P; Su, N; Lin, F Z; Ma, L; Wang, H J; Rong, X; Dai, Y D; Li, J; Jian, Z W; Tang, L H; Xiao, W; Li, C Q

2015-12-09

Like other developing countries, China was reported to have a relatively high seroprevalence of anti-hepatitis A antibodies (anti-HAV). However, no studies have evaluated the prevalence of anti-HAV and HAV RNA among voluntary blood donors with or without elevated serum alanine transaminase (ALT) levels. Anti-HAV antibodies were detected using an enzyme-linked immunosorbent assay, and reverse transcription quantitative polymerase chain reaction was carried out for detection of HAV RNA. In the current study, we analyzed a total of 450 serum samples with elevated ALT levels (≥40 U/L) and 278 serum samples with non-elevated ALT levels. Seroprevalence rates of anti-HAV were 51.6% in donors with elevated ALT and 41.4% in donors with non-elevated ALT; however, none of the samples was positive for HAV RNA. The results of our study showed lower seroprevalence rates of anti-HAV in blood donors (irrespective of ALT levels) than those in published data on Chinese populations. Although donors with elevated ALT had statistically higher prevalence rates of anti- HAV than did those with non-elevated ALT, none of the serum samples had detectable levels of the active virus. In conclusion, our results demonstrate that the transmission of hepatitis A by blood transfusion will occur rarely.

[Effects of psychological stress on performances in open-field test of rats and tyrosine's modulation].

PubMed

Chen, Wei-Qiang; Cheng, Yi-Yong; Li, Shu-Tian; Hong, Yan; Wang, Dong-Lan; Hou, Yue

2009-02-01

To explore the effects of different doses of tyrosine modulation on behavioral performances in open field test of psychological stress rats. The animal model of psychological stress was developed by restraint stress for 21 days. Wistar rats were randomly assigned to five groups (n = 10) as follows: control group (CT), stress control group (SCT), low, medium and high-doses of tyrosine modulation stress groups (SLT, SMT and SIT). The changes of behavioral performances were examined by open-field test. Serum levels of cortisol, norepinephrine and dopamine were also detected. The levels of serum cortisol were all increased obviously in the four stress groups, and their bodyweight gainings were diminished. The behavioral performances of SCT rats in open-field test were changed significantly in contrast to that of CT rats. However, The behavioral performances of SMT and SHT rats were not different from that of CT rats. In addition, the serum levels of norepinephrine and dopamine were downregulated obviously in SCT and SLT groups, and no differences were observed in other groups. Psychological stress can impair body behavioral performances, and moderate tyrosine modulation may improve these abnormal changes. The related mechanisms may be involved with the changes of norepinephrine and dopamine.

A novel dynamic flow immunochromatographic test (DFICT) using gold nanoparticles for the serological detection of Toxoplasma gondii infection in dogs and cats.

PubMed

Jiang, Wei; Liu, Yingchun; Chen, Yongjun; Yang, Qiufeng; Chun, Peter; Yao, Kailing; Han, Xiangan; Wang, Shaohui; Yu, Shengqing; Liu, Yongjie; Wang, Quan

2015-10-15

A novel dynamic flow immunochromatographic test (DFICT) is proposed for rapid assay utilizing Toxoplasma gondii as a model. The test is based on a proprietary technology that combines the principles of immunochromatography and fluid dynamics. Gold nanoparticles conjugated to staphylococcal protein A (SPA) were prepared in liquid form and used as signal vehicles. T. gondii-specific recombinant antigens and SPA were sprayed onto a nitrocellulose membrane in strips at positions designated as T and C, respectively. The DFICT is performed by applying a 100 µL aliquot of liquid gold-SPA conjugate to the reagent hole and a 5 μL aliquot of serum sample to the sample hole. The results were observable within 5 min by the naked eye. The lowest detectable limit of the assay was determined as the highest dilution (1:320) of positive serum. No cross-reaction of the antibodies with other related canine or feline pathogens was observed. The DFICT can be stored for 12 months at 4°C or 6 months with no loss of sensitivity or specificity. A high degree of consistency was observed between the DFICT and the standard ELISA kit, supporting the reliability of the novel test strip. The introduction of a liquid gold nanoparticle conjugate reagent provides this method with several attractive characteristics, such as ease of manufacture, low sample volume requirements, high selectivity and high efficiency. This method opens a novel pathway for rapid diagnostic screening and field analysis. Copyright © 2015 Elsevier B.V. All rights reserved.

Reproducibility of biomarkers in induced sputum and in serum from chronic smokers.

PubMed

Zuiker, Rob G J A; Kamerling, Ingrid M C; Morelli, Nicoletta; Calderon, Cesar; Boot, J Diderik; de Kam, Marieke; Diamant, Zuzana; Burggraaf, Jacobus; Cohen, Adam F

2015-08-01

Soluble inflammatory markers obtained from non-invasive airway sampling such as induced sputum may be useful biomarkers for targeted pharmaceutical interventions. However, before these soluble markers can be used as potential targets, their variability and reproducibility need to be established in distinct study populations. This study aimed to assess the reproducibility of biomarkers obtained from induced sputum and serum in chronic smokers and non-smokers. Sputum and serum samples were obtained from 16 healthy non-smokers and 16 asymptomatic chronic smokers (for both groups: 8M/8F, 30-52 years, FEV1 ≥80% pred.; ≥10 pack years for the smokers) on 2 separate visits 4-10 days apart. Soluble markers in serum and sputum were analysed by ELISA. The differences between smokers vs non-smokers were analysed with a t-test and variability was assessed on log-transformed data by a mixed model ANOVA. Analysable sputum samples could be obtained from all 32 subjects. In both study populations neutrophils and macrophages were the predominant cell types. Serum Pulmonary Surfactant Associated Protein D had favourable reproducibility criteria for reliability ratio (0.99), intra-subject coefficient of variation (11.2%) and the Bland Altman limits of agreement. Furthermore, chronic smokers, compared to non-smokers, had significantly higher sputum concentrations of IL-8 (1094.6 pg/mL vs 460.8 pg/mL, p = 0.006)), and higher serum concentrations of Pulmonary Surfactant Associated Protein D (110.9 pg/mL vs 64.7 pg/mL, p = 0.019), and lower concentrations of Serum Amyloid A (1352.4 pg/mL vs 2297.5 pg/mL, p = 0.022). Serum Pulmonary Surfactant Associated Protein D proved to be a biomarker that fulfilled the criteria for reproducibility in both study groups. Copyright © 2015 Elsevier Ltd. All rights reserved.

Development and application of molecularly imprinted polymer - Mn-doped ZnS quantum dot fluorescent optosensing for cocaine screening in oral fluid and serum.

PubMed

Chantada-Vázquez, María Pilar; de-Becerra-Sánchez, Carolina; Fernández-Del-Río, Alba; Sánchez-González, Juan; Bermejo, Ana María; Bermejo-Barrera, Pilar; Moreda-Piñeiro, Antonio

2018-05-01

A molecularly imprinted polymer - Mn-doped ZnS quantum dot-based fluorescence probe for cocaine abuse screening has been prepared and applied to complex samples such as serum and oral fluid. The fluorescent sensing material was prepared by anchoring a selective MIP for COC on the surface of polyethylene glycol (PEG) modified Mn-doped ZnS quantum dots (QDs). Simple and low cost methods have thus been optimized for assessing cocaine abuse in serum and oral fluid by monitoring fluorescence quenching when cocaine (COC) is present (optimized operating conditions with 1.5mL of 200mgL -1 MIP-coated QDs solution, pH 5.5, and 15min before fluorescence scanning). The matrix effect was found to be important when analyzing oral fluid and serum, and several strategies based on centrifugation for oral fluid and solid phase extraction (SPE) for serum were explored. Two analytical methods were developed for oral fluid. The first one (direct method) requires a centrifugation step (6°C, 4000rpm, 20min) to avoid the matrix effect, and allows for cocaine determination by using an aqueous calibration (1:20 dilution). The second method was developed for oral fluid sampled by Salivette devices, and also requires a further centrifugation (6°C, 4000rpm, 20min) of the recovered oral fluid. This method, however, requires the standard addition technique (1:20 dilution) because of the existence of the matrix effect. Regarding serum samples, a direct method (serum dilution) was not possible, and an SPE procedure was needed to avoid the matrix effect (use of aqueous calibration). The limits of detection and quantification when using the Salivette method were 0.035mgL -1 and 0.117mgL -1, respectively; whereas, 0.015mgL -1 (LOD) and 0.050mgL -1 (LOQ) were obtained for serum. Copyright © 2018 Elsevier B.V. All rights reserved.

Ante mortem identification of BSE from serum using infrared spectroscopy

NASA Astrophysics Data System (ADS)

Schmitt, Jürgen; Lasch, Peter; Beekes, Michael; Udelhoven, Thomas; Eiden, Michael; Fabian, Heinz; Petrich, Wolfgang H.; Naumann, Dieter

2004-07-01

In our former studies a diagnostic approach for the detection of transmissible spongiform encephalopaties (TSE) based on FT-IR spectroscopy in combination with artificial neural networks was described, based on a controlled animal study with terminally ill Syrian hamsters and control animals. As a consequence of the bovine spongiform encephalopathy (BSE) crisis in Europe, the development of a disgnostic ante mortem test for cattle has become a matter of great scientific importance and public interest. Since 1986 more than 180,000 clinical cases of BSE have been observed in the UK alone. Most of these cases were confirmed by post mortem examination of brain tissue. However, BSE-related risk assessment and risk-management would greatly benefit from ante mortem testing on living animals. For example, a serum-based test could allow for screening of the cattle population, thus, even a BSE eradication program would be conceivable. Here we report on a novel method for ante mortem BSE testing, which combines infrared spectroscopy of serum samples with multivariate pattern recognition analysis. A classification algorithm was trained using infrared spectra of sera from more than 800 animals from a field study (including BSE positive, healthy controls and animals suffering from viral or bacterial infections). In two validation studies sensitivities of 85% and 87% and specificities of 84% and 91% were achieved, respectively. The combination of classification algorithms increased sensitivity and specificity to 96% and 92%, respectively.

High VEGF-D and Low MMP-2 Serum Levels Predict Nodal-Positive Disease in Invasive Bladder Cancer

PubMed Central

Benoit, Tobias; Keller, Etienne X.; Wolfsgruber, Pirmin; Hermanns, Thomas; Günthart, Michele; Banzola, Irina; Sulser, Tullio; Provenzano, Maurizio; Poyet, Cédric

2015-01-01

Background To investigate stromal variables including angiogenesis, lymphangiogenesis, and matrix metalloproteinase (MMP) in the serum of patients with urothelial carcinoma of the bladder (UCB) and to evaluate their association with histopathological characteristics and clinical outcome. Material/Methods Protein levels of vascular endothelial growth factors-A, -C, -D (VEGF-A/-C/-D), their receptors- VEGF-R2 and -R3 (VEGF-R2/-R3), and matrix metalloproteinases 2, -3, and -7 (MMP-2, MMP-3, MMP-7) were quantified in the blood serum samples of 71 patients with UCB before radical cystectomy (RC). Samples of patients with non-invasive UCB or no history of UCB were investigated as controls (n=20). Protein levels in the serum were measured using a flow cytometric cytokine assay. Results A positive association for VEGF-D (p<0.001) and an inverse association for MMP-2 (p=0.017) were observed in patients with positive lymph node (LN) status at the time of RC. VEGF-A (p<0.001), VEGF-C (p<0.001), MMP-2 (p<0.001), and MMP-7 (p=0.005) serum levels were different in serum of patients with invasive UCB compared with non-invasive UCB or healthy individuals. None of the serum markers were associated with disease progression. Conclusions High VEGF-D and low MMP-2 serum levels predict LN metastasis in patients with UCB at the time of RC. VEGF-A, VEGF-C, MMP-2, and MMP-7 serum levels varied significantly between invasive and non-invasive disease as well as in comparison with healthy individuals. Clinical implementation of these marker serum measurements may be valuable to select high-risk patients with more invasive or nodal-positive disease. PMID:26241709

Associations of Socioeconomic Status and Processed Food Intake with Serum Phosphorus in Community-Living Adults: the Multi-Ethnic Study of Atherosclerosis (MESA)

PubMed Central

Gutiérrez, Orlando M.; Katz, Ronit; Peralta, Carmen A.; de Boer, Ian H.; Siscovick, David; Wolf, Myles; Roux, Ana Diez; Kestenbaum, Bryan; Nettleton, Jennifer A.; Ix, Joachim H.

2011-01-01

Objective Higher serum phosphorus concentrations are associated with cardiovascular disease events and mortality. Low socioeconomic status is linked with higher serum phosphorus, but the reasons are unclear. Poor individuals disproportionately consume inexpensive processed foods commonly enriched with phosphorus-based food preservatives. Accordingly, we hypothesized that excess intake of these foods accounts for a relationship between lower socioeconomic status and higher serum phosphorus. Design Cross-sectional analysis. Setting and Participants We examined a random cohort of 2,664 participants with available phosphorus measurements in the Multi-Ethnic Study of Atherosclerosis, a community-based sample of individuals free of clinically apparent cardiovascular disease from across the United States. Predictor Variables Socioeconomic status, the intake of foods commonly enriched with phosphorus additives (processed meats, sodas) and frequency of fast food consumption. Outcomes Fasting morning serum phosphorus concentrations. Results In unadjusted analyses, lower income and lower educational achievement categories were associated with modestly higher serum phosphorus (by 0.02 to 0.10 mg/dL, P < 0.05 for all). These associations were attenuated in models adjusted for demographic and clinical factors, almost entirely due to adjustment for female gender. There were no statistically significant associations of processed meat intake or frequency of fast-food consumption with serum phosphorus in multivariable-adjusted analyses. In contrast, each serving per day higher soda intake was associated with 0.02 mg/dl lower serum phosphorus (95% confidence interval, −0.04, −0.01). Conclusions Greater intake of foods commonly enriched with phosphorus additives was not associated with higher serum phosphorus in a community-living sample with largely preserved kidney function. These results suggest that excess intake of processed and fast foods may not impact fasting serum phosphorus concentrations among individuals without kidney disease. PMID:22217539

Diagnostic value of serum Golgi protein 73 for HBV-related primary hepatic carcinoma

PubMed Central

Gao, Guosheng; Dong, Feibo; Xu, Xiaozhen; Hu, Airong; Hu, Yaoren

2015-01-01

Background: Alpha-fetoprotein (AFP) levels are routinely used for diagnosis and monitoring of hepatic diseases, but it has a limited value. Golgi protein 73 (GP73) has been suggested as a new marker for hepatic diseases. Objective: To explore the clinical value of serum GP73 in different diseases associated with hepatitis B virus (HBV) infection. Method: Between January 2010 and August 2014, serum samples from 88 patients with chronic hepatitis B (CHB), 78 patients with HBV-related liver cirrhosis (LC), and 194 patients with HBV-related primary hepatic cancer (PHC) were collected. Serum samples from 30 healthy volunteers were used as controls. ELISA and microparticle enzyme immunoassay were used to measure serum GP73 and AFP levels. Receiver operating characteristic (ROC) curves were used to analyze the diagnostic value of serum GP73 and AFP for PHC. Results: For the diagnosis of PHC, GP73 showed a sensitivity of 65.5% and specificity of 66.3%, while AFP levels showed sensitivity of 64.4% and specificity of 76.5%. Serial testing (both tests are positive) could increase the specificity (sensitivity of 45.9% and specificity of 85.5%) while parallel testing (any single positive test result) could increase the sensitivity (sensitivity of 84.0% and specificity of 57.2%). Serum GP73 and AFP levels were significantly different between Child-Pugh grades (P<0.001 for GP73 and P=0.044 for AFP). Significant differences in serum GP73 and AFP were found between TNM stages (all P<0.001). Conclusion: Serum GP73 had limited diagnostic value for HBV-related PHC. The combined use of serum GP73 and AFP levels improved the diagnostic efficacy. PMID:26617863

Analytical Comparison of In Vitro-Spiked Human Serum and Plasma for PCR-Based Detection of Aspergillus fumigatus DNA: a Study by the European Aspergillus PCR Initiative

PubMed Central

Mengoli, Carlo; Springer, Jan; Bretagne, Stéphane; Cuenca-Estrella, Manuel; Klingspor, Lena; Lagrou, Katrien; Melchers, Willem J. G.; Morton, C. Oliver; Barnes, Rosemary A.; Donnelly, J. Peter; White, P. Lewis

2015-01-01

The use of serum or plasma for Aspergillus PCR testing facilitates automated and standardized technology. Recommendations for serum testing are available, and while serum and plasma are regularly considered interchangeable for use in fungal diagnostics, differences in galactomannan enzyme immunoassay (GM-EIA) performance have been reported and are attributed to clot formation. Therefore, it is important to assess plasma PCR testing to determine if previous recommendations for serum are applicable and also to compare analytical performance with that of serum PCR. Molecular methods testing serum and plasma were compared through multicenter distribution of quality control panels, with additional studies to investigate the effect of clot formation and blood fractionation on DNA availability. Analytical sensitivity and time to positivity (TTP) were compared, and a regression analysis was performed to identify variables that enhanced plasma PCR performance. When testing plasma, sample volume, preextraction-to-postextraction volume ratio, PCR volume, duplicate testing, and the use of an internal control for PCR were positively associated with performance. When whole-blood samples were spiked and then fractionated, the analytical sensitivity and TTP were superior when testing plasma. Centrifugation had no effect on DNA availability, whereas the presence of clot material significantly lowered the concentration (P = 0.028). Technically, there are no major differences in the molecular processing of serum and plasma, but the formation of clot material potentially reduces available DNA in serum. During disease, Aspergillus DNA burdens in blood are often at the limits of PCR performance. Using plasma might improve performance while maintaining the methodological simplicity of serum testing. PMID:26085614

Per- and polyfluoroalkyl substances in human serum and urine samples from a residentially exposed community.

PubMed

Worley, Rachel Rogers; Moore, Susan McAfee; Tierney, Bruce C; Ye, Xiaoyun; Calafat, Antonia M; Campbell, Sean; Woudneh, Million B; Fisher, Jeffrey

2017-09-01

Per- and polyfluoroalkyl substances (PFAS) are considered chemicals of emerging concern, in part due to their environmental and biological persistence and the potential for widespread human exposure. In 2007, a PFAS manufacturer near Decatur, Alabama notified the United States Environmental Protection Agency (EPA) it had discharged PFAS into a wastewater treatment plant, resulting in environmental contamination and potential exposures to the local community. To characterize PFAS exposure over time, the Agency for Toxic Substances and Disease Registry (ATSDR) collected blood and urine samples from local residents. Eight PFAS were measured in serum in 2010 (n=153). Eleven PFAS were measured in serum, and five PFAS were measured in urine (n=45) from some of the same residents in 2016. Serum concentrations were compared to nationally representative data and change in serum concentration over time was evaluated. Biological half-lives were estimated for perfluorooctanoic acid (PFOA), perfluorooctane sulfonic acid (PFOS), and perfluorohexane sulfonic acid (PFHxS) using a one-compartment pharmacokinetic model. In 2010 and 2016, geometric mean PFOA and PFOS serum concentrations were elevated in participants compared to the general U.S. In 2016, the geometric mean PFHxS serum concentration was elevated compared to the general U.S. Geometric mean serum concentrations of PFOA, PFOS, and perfluorononanoic acid (PFNA) were significantly (p≤0.0001) lower (49%, 53%, and 58%, respectively) in 2016 compared to 2010. Half-lives for PFOA, PFOS, and PFHxS were estimated to be 3.9, 3.3, and 15.5years, respectively. Concentrations of PFOA in serum and urine were highly correlated (r=0.75) in males. Serum concentrations of some PFAS are decreasing in this residentially exposed community, but remain elevated compared to the U.S. general population. Published by Elsevier Ltd.

Serum levels of inhibin A and inhibin B in women with normal and abnormal luteal function.

PubMed

Yamoto, M; Imai, M; Otani, H; Nakano, R

1997-05-01

To determine whether serum inhibin A and inhibin B concentrations are lower in patients with luteal dysfunction than in women with normal luteal function. Serum samples were collected from seven healthy women with regular menstrual cycles. Serum samples on days +5 to +9 after the LH surge were collected from patients with luteal dysfunction. The diagnosis of luteal dysfunction was based on a luteal phase duration less than 11 days and a single midluteal progesterone level below 10 ng/mL. Serum levels of inhibin A, inhibin B, progesterone, estradiol (E2), FSH, and LH were measured. The serum inhibin A levels were increased toward the late follicular phase. The levels reached a maximum during the midluteal phase, followed by a fall during the late luteal phase. The serum inhibin B levels were high during the follicular phases and the early luteal phase. The levels decreased during the midluteal and late luteal phases. Serum levels (mean +/- standard error of the mean) of inhibin A in patients with luteal dysfunction were significantly lower than those in women during the midluteal phase (26.2 +/- 2.9 compared to 41.9 +/- 2.8 pg/mL; P < .01) in addition to the expected decrease in serum progesterone levels (6.3 +/- 0.7 compared to 14.7 +/- 1.2 ng/mL; P < .01). Serum inhibin B levels did not differ significantly between normal women and those with luteal dysfunction. There also were no significant differences in the E2, FSH, and LH levels. Levels of inhibin A, but not of inhibin B, may reflect the human luteal function.

Analytical Comparison of In Vitro-Spiked Human Serum and Plasma for PCR-Based Detection of Aspergillus fumigatus DNA: a Study by the European Aspergillus PCR Initiative.

PubMed

Loeffler, Juergen; Mengoli, Carlo; Springer, Jan; Bretagne, Stéphane; Cuenca-Estrella, Manuel; Klingspor, Lena; Lagrou, Katrien; Melchers, Willem J G; Morton, C Oliver; Barnes, Rosemary A; Donnelly, J Peter; White, P Lewis

2015-09-01

The use of serum or plasma for Aspergillus PCR testing facilitates automated and standardized technology. Recommendations for serum testing are available, and while serum and plasma are regularly considered interchangeable for use in fungal diagnostics, differences in galactomannan enzyme immunoassay (GM-EIA) performance have been reported and are attributed to clot formation. Therefore, it is important to assess plasma PCR testing to determine if previous recommendations for serum are applicable and also to compare analytical performance with that of serum PCR. Molecular methods testing serum and plasma were compared through multicenter distribution of quality control panels, with additional studies to investigate the effect of clot formation and blood fractionation on DNA availability. Analytical sensitivity and time to positivity (TTP) were compared, and a regression analysis was performed to identify variables that enhanced plasma PCR performance. When testing plasma, sample volume, preextraction-to-postextraction volume ratio, PCR volume, duplicate testing, and the use of an internal control for PCR were positively associated with performance. When whole-blood samples were spiked and then fractionated, the analytical sensitivity and TTP were superior when testing plasma. Centrifugation had no effect on DNA availability, whereas the presence of clot material significantly lowered the concentration (P = 0.028). Technically, there are no major differences in the molecular processing of serum and plasma, but the formation of clot material potentially reduces available DNA in serum. During disease, Aspergillus DNA burdens in blood are often at the limits of PCR performance. Using plasma might improve performance while maintaining the methodological simplicity of serum testing. Copyright © 2015 Loeffler et al.

Field evaluation of two point-of-care tests for syphilis among men who have sex with men, Verona, Italy.

PubMed

Zorzi, Antonella; Cordioli, Maddalena; Gios, Lorenzo; Del Bravo, Paola; Toskin, Igor; Peeling, Rosanna W; Blondeel, Karel; Cornaglia, Giuseppe; Kiarie, James; Ballard, Ronald; Mirandola, Massimo

2017-12-01

The incidence of HIV and syphilis among men who have sex with men (MSM) in Europe has recently increased. Rapid point-of-care tests (POCTs) for syphilis can improve access to screening. The purpose of this study was to evaluate the performance of two syphilis POCTs compared with laboratory tests among MSM. The study was undertaken in Verona, Italy. Asymptomatic MSM, potentially exposed to syphilis, were enrolled prospectively. The POCTs evaluated were SD Bioline Syphilis 3.0 and Chembio DPP Syphilis Screen & Confirm Assay on both serum and fingerprick blood. The results of the POCTs were read by the naked eye by two independent readers and their concordance assessed. A total of 289 MSM were enrolled in the study. Based on laboratory tests, 35 MSM (12.1%) were TPPA-positive alone and 16 (5.5%) were both Treponema pallidum particle agglutination test (TPPA) and rapid plasma reagin (RPR)-positive. The specificities of both POCTs were above 99% on both serum and fingerstick blood specimens, while sensitivities varied considerably. The sensitivity of the SD Bioline test was lower on fingerprick blood (51.4% and 54.3%, readers 1 and 2, respectively) compared with that on serum (80.0% and 82.9%). In contrast, the Chembio test exhibited similar sensitivity values for serum and fingerprick samples (57.7% and 64.0% on serum vs 65.4% and 69.2% on fingerprick for the treponemal component; 63.6% on both samples by both readers for the non-treponemal component). The positive predictive value ranged between 100% and 93.9% for the treponemal component of both syphilis POCTs, but was lower (76.3%-100%)%) for the non-treponemal component of the Chembio POCT. The negative predictive value surpassed 90% for both tests on both samples. The agreement between readers was very high (>99%). The diagnostic performance of the syphilis POCTs was lower than expected; however, considering the prevalence of syphilis among MSM, POCTs should be recommended to improve syphilis detection among MSM. © World Health Organization [2017]. Licensee BMJ Publishing Group Limited. This is an open access article distributed under the terms of the Creative Commons Attribution IGO License (CC BY 4.0), which permits use, distribution, and reproduction for non-commercial purposes in any medium, provided the original work is properly cited. In any reproduction of this article there should not be any suggestion that WHO or this article endorse any specific organization or products. The use of the WHO logo is not permitted. This notice should be preserved along with the article’s original URL.