Sample records for samples serum samples
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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.
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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.
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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
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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.
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Serum samples can be substituted by plasma samples for the diagnosis of paratuberculosis.
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.
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A comparison of serum and plasma cytokine values using a multiplexed assay in cats.
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.
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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.
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IL8 and IL16 levels indicate serum and plasma quality.
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.
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Comparison of Schmallenberg virus antibody levels detected in milk and serum from individual cows.
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.
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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
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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
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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.
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Specific IgA and IgG antibodies in paired serum and breast milk samples in human strongyloidiasis.
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.
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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.
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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.
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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.
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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
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Comparison of oral fluid collection methods for the molecular detection of hepatitis B virus.
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.
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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
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Use of dried blood spots for the determination of serum concentrations of tamoxifen and endoxifen.
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.
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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.
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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.
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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
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Drug residues in serum of dogs receiving anticancer chemotherapy.
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.
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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
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Serological evidence for the presence of influenza D virus in small ruminants.
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.
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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.
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Serum Dried Samples to Detect Dengue Antibodies: A Field Study.
Maldonado-Rodríguez, Angelica; Rojas-Montes, Othon; Vazquez-Rosales, Guillermo; Chavez-Negrete, Adolfo; Rojas-Uribe, Magdalena; Posadas-Mondragon, Araceli; Aguilar-Faisal, Leopoldo; Cevallos, Ana Maria; Xoconostle-Cazares, Beatriz; Lira, Rosalia
2017-01-01
Dried blood and serum samples are useful resources for detecting antiviral antibodies. The conditions for elution of the sample need to be optimized for each disease. Dengue is a widespread disease in Mexico which requires continuous surveillance. In this study, we standardized and validated a protocol for the specific detection of dengue antibodies from dried serum spots (DSSs). Paired serum and DSS samples from 66 suspected cases of dengue were collected in a clinic in Veracruz, Mexico. Samples were sent to our laboratory, where the conditions for optimal elution of DSSs were established. The presence of anti-dengue antibodies was determined in the paired samples. DSS elution conditions were standardized as follows: 1 h at 4°C in 200 µ l of DNase-, RNase-, and protease-free PBS (1x). The optimal volume of DSS eluate to be used in the IgG assay was 40 µ l. Sensitivity of 94%, specificity of 93.3%, and kappa concordance of 0.87 were obtained when comparing the antidengue reactivity between DSSs and serum samples. DSS samples are useful for detecting anti-dengue IgG antibodies in the field.
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Serum Dried Samples to Detect Dengue Antibodies: A Field Study
Maldonado-Rodríguez, Angelica; Rojas-Montes, Othon; Chavez-Negrete, Adolfo; Rojas-Uribe, Magdalena; Posadas-Mondragon, Araceli; Aguilar-Faisal, Leopoldo; Xoconostle-Cazares, Beatriz
2017-01-01
Background Dried blood and serum samples are useful resources for detecting antiviral antibodies. The conditions for elution of the sample need to be optimized for each disease. Dengue is a widespread disease in Mexico which requires continuous surveillance. In this study, we standardized and validated a protocol for the specific detection of dengue antibodies from dried serum spots (DSSs). Methods Paired serum and DSS samples from 66 suspected cases of dengue were collected in a clinic in Veracruz, Mexico. Samples were sent to our laboratory, where the conditions for optimal elution of DSSs were established. The presence of anti-dengue antibodies was determined in the paired samples. Results DSS elution conditions were standardized as follows: 1 h at 4°C in 200 µl of DNase-, RNase-, and protease-free PBS (1x). The optimal volume of DSS eluate to be used in the IgG assay was 40 µl. Sensitivity of 94%, specificity of 93.3%, and kappa concordance of 0.87 were obtained when comparing the antidengue reactivity between DSSs and serum samples. Conclusion DSS samples are useful for detecting anti-dengue IgG antibodies in the field. PMID:28630868
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Effect of Repeated Freezing and Thawing on 18 Clinical Chemistry Analytes in Rat Serum
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
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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.
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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
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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
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Measurement of rivaroxaban and apixaban in serum samples of patients
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
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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.
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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.
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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.
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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.
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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.
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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.
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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.
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Evaluation of pharmacokinetics and the stability of daptomycin in serum at various temperatures.
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.
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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.
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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.
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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.
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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.
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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.
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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.
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In vitro production of GHB in blood and serum samples under various storage conditions.
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.
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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
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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.
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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.
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Comparison of serum and plasma measurements of Müllerian inhibiting substance.
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.
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Detection of Serum microRNAs From Department of Defense Serum Repository
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
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Quantitative bioanalysis of strontium in human serum by inductively coupled plasma-mass spectrometry
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
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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.
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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
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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.
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Effect of storage duration on cytokine stability in human serum and plasma.
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.
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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.
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SELDI-TOF-MS proteomic profiling of serum, urine, and amniotic fluid in neural tube defects.
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.
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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.
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Gastrointestinal parasites of feral cats from Christmas Island.
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.
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An enzyme-linked immunosorbent assay for the evaluation of thrombocytopenia induced by heparin.
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.
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METHOD FOR MICRORNA ISOLATION FROM CLINICAL SERUM SAMPLES
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
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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.
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Mimivirus Circulation among Wild and Domestic Mammals, Amazon Region, Brazil
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
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Antibodies against MERS coronavirus in dromedary camels, United Arab Emirates, 2003 and 2013.
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.
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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.
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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
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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
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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.
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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.
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Amoeba-Resisting Bacteria and Ventilator-Associated Pneumonia
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
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Polyfluoroalkyl substance exposure in the Mid-Ohio River Valley, 1991-2012.
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.
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Organic cattle products: Authenticating production origin by analysis of serum mineral content.
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.
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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.
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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.
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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.
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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.
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Correlation Between Iron and alpha and pi Glutathione-S-Transferase Levels in Humans
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
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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.
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Influence of the collection tube on metabolomic changes in serum and plasma.
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.
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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.
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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.
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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.
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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.
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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
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Pilot Metabolome-Wide Association Study of Benzo(a)pyrene in Serum from Military Personnel
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
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Pilot Metabolome-Wide Association Study of Benzo(a)pyrene in Serum From Military Personnel.
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.
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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.
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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.
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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.
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Improvement in the stability of serum samples stored in an automated refrigerated module.
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.
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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
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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.
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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.
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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
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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.
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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.
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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.
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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.
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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
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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.
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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.
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Heterophile antibody interference in qualitative urine/serum hCG devices: Case report.
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.
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Evaluation of an enzyme immunoassay for detection of dengue virus NS1 antigen in human serum.
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.
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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
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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
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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.
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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.
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Feasibility study of feces for noninvasive biomonitoring of brominated flame retardants in toddlers.
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.
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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.
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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.
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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.
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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
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Determination of chlormequat in pig serum and sow milk by LC-MS/MS.
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.
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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.
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Serum and saliva cortisol relations in adolescents during pregnancy and the early postpartum period.
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.
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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
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Organochlorine pesticide residue levels in blood serum of inhabitants from Veracruz, Mexico.
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.
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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).
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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.
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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
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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.
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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.
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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.
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Isolation of Coxiella burnetii from serum of patients with acute Q fever.
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.
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[Determination of ethylene glycol in biological fluids--propylene glycol interferences].
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.
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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
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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.
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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.
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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
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Partition of environmental chemicals between maternal and fetal blood and tissues.
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.
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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.
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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.
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Urine phenobarbital drug screening: potential use for compliance assessment in neonates.
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.
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Occurrence of West Nile virus antibodies in wild birds, horses, and humans in Poland.
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.
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Occurrence of West Nile Virus Antibodies in Wild Birds, Horses, and Humans in Poland
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
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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.
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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.
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Metabolomic Quality Assessment of EDTA Plasma and Serum Samples.
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.
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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.
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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
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Detection of a Serum Siderophore by LC-MS/MS as a Potential Biomarker of Invasive Aspergillosis
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
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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.
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[Presence of west Nile virus in northeast Mexico].
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.
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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
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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.
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Use of Paired Serum Samples for Serodiagnosis of Typhoid Fever
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
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Sipa1l1 is an early biomarker of liver fibrosis in CCl4-treated rats
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
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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
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Serum and urine chromium as indices of chromium status in tannery workers.
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.
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Alemtuzumab: validation of a sensitive and simple enzyme-linked immunosorbent assay.
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.
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NMR Metabolomics Investigates the Influence of Flavonoid-Enriched Rations on Chicken Plasma.
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.
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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.
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Serological evidence of hepatitis E virus infection in dromedary camels in Ethiopia.
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.
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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.
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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.
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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.
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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.
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Yellow Fever Outbreak, Imatong, Southern Sudan
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
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Lack of Serologic Evidence of Neospora caninum in Humans, England
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
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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.
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Indigenous West Nile virus infections in horses in Albania.
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.
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Clinical evaluation of a Mucorales-specific real-time PCR assay in tissue and serum samples.
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.
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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.
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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.
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Serum Albumin Domain Structures in Human Blood Serum by Mass Spectrometry and Computational Biology.
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.
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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.
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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%).
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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.
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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.
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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.
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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
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Hemolysis associated with pneumatic tube system transport for blood samples
Kara, Hasan; Bayir, Aysegul; Ak, Ahmet; Degirmenci, Selim; Akinci, Murat; Agacayak, Ahmet; Marcil, Emine; Azap, Melih
2014-01-01
Objective: The frequency of hemolysis of blood samples may be increased by transport in a pneumatic tube system. The purpose of this study was to evaluate the effect of pneumatic tube system transport on hemolysis of blood samples. Methods: Blood samples were transported from the emergency department to the hospital laboratory manually by hospital staff (49 patients) or with a pneumatic tube system (53 patients). The hemolysis index and serum chemistry studies were performed on the blood samples and compared between the different methods of transport. Results: The blood samples that were transported by the pneumatic tube system had a greater frequency of hemolysis and greater mean serum potassium and median creatinine, aspartate aminotransferase, and lactate dehydrogenase levels than samples transported manually. Conclusion: Blood samples transported from the emergency department to the hospital laboratory by a pneumatic tube system may have a greater frequency of hemolysis than samples transported manually. This may necessitate repeat phlebotomy and cause a delay in completing the laboratory analysis. PMID:24639830
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Polychlorinated biphenyls in umbilical cord serum of newborns from Rio Grande do Sul state, Brazil.
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.
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Nucleic Acid-Based Cross-Linking Assay for Detection and Quantification of Hepatitis B Virus DNA
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
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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.
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Stability of selected serum hormones and lipids after long-term storage in the Janus Serum Bank.
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.
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Lyssavirus Surveillance in Bats, Bangladesh
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
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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.
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Koren, Lee; Ng, Ella S M; Soma, Kiran K; Wynne-Edwards, Katherine E
2012-01-01
Blood samples from wild mammals and birds are often limited in volume, allowing researchers to quantify only one or two steroids from a single sample by immunoassays. In addition, wildlife serum or plasma samples are often lipemic, necessitating stringent sample preparation. Here, we validated sample preparation for simultaneous liquid chromatography--tandem mass spectrometry (LC-MS/MS) quantitation of cortisol, corticosterone, 11-deoxycortisol, dehydroepiandrosterone (DHEA), 17β-estradiol, progesterone, 17α-hydroxyprogesterone and testosterone from diverse mammalian (7 species) and avian (5 species) samples. Using 100 µL of serum or plasma, we quantified (signal-to-noise (S/N) ratio ≥ 10) 4-7 steroids depending on the species and sample, without derivatization. Steroids were extracted from serum or plasma using automated solid-phase extraction where samples were loaded onto C18 columns, washed with water and hexane, and then eluted with ethyl acetate. Quantitation by LC-MS/MS was done in positive ion, multiple reaction-monitoring (MRM) mode with an atmospheric pressure chemical ionization (APCI) source and heated nebulizer (500°C). Deuterated steroids served as internal standards and run time was 15 minutes. Extraction recoveries were 87-101% for the 8 analytes, and all intra- and inter-run CVs were ≤ 8.25%. This quantitation method yields good recoveries with variable lipid-content samples, avoids antibody cross-reactivity issues, and delivers results for multiple steroids. Thus, this method can enrich datasets by providing simultaneous quantitation of multiple steroids, and allow researchers to reimagine the hypotheses that could be tested with their volume-limited, lipemic, wildlife samples.
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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).
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Nipah Virus in Lyle's Flying Foxes, Cambodia
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
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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.
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Stability of Chronic Hepatitis-Related Parameters in Serum Samples After Long-Term Storage.
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.
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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.
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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.
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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.
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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.
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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
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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.
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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.
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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.
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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.
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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.
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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.
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Colon Reference Set Application: Mary Disis - University of Washington (2008) — EDRN Public Portal
The proposed study aims to validate the diagnostic value of a panel of serum antibodies for the early detection of colorectal cancer (CRC). We have developed a serum antibody based assay that shows promise in discriminating sera from CRC patients from healthy donors. We have evaluated two separate sample sets of sera that were available either commercially or were comprised of left over samples from previous studies by our group. Both sample sets showed concordance in discriminatory power. We have not been able to identify investigators with a larger, well defined sample set of early stage colon cancer sera and request assistance from the EDRN in obtaining such samples to help assess the potential diagnostic value of our autoantibody panel
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Avian influenza A (H5N1) virus antibodies in pigs and residents of swine farms, southern China.
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.
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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.
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Traum, Avram Z; Wells, Meghan P; Aivado, Manuel; Libermann, Towia A; Ramoni, Marco F; Schachter, Asher D
2006-03-01
Proteomic profiling with SELDI-TOF MS has facilitated the discovery of disease-specific protein profiles. However, multicenter studies are often hindered by the logistics required for prompt deep-freezing of samples in liquid nitrogen or dry ice within the clinic setting prior to shipping. We report high concordance between MS profiles within sets of quadruplicate split urine and serum samples deep-frozen at 0, 2, 6, and 24 h after sample collection. Gage R&R results confirm that deep-freezing times are not a statistically significant source of SELDI-TOF MS variability for either blood or urine.
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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.
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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
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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.
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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.
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A round robin approach to the analysis of bisphenol a (BPA) in human blood samples
2014-01-01
Background Human exposure to bisphenol A (BPA) is ubiquitous, yet there are concerns about whether BPA can be measured in human blood. This Round Robin was designed to address this concern through three goals: 1) to identify collection materials, reagents and detection apparatuses that do not contribute BPA to serum; 2) to identify sensitive and precise methods to accurately measure unconjugated BPA (uBPA) and BPA-glucuronide (BPA-G), a metabolite, in serum; and 3) to evaluate whether inadvertent hydrolysis of BPA-G occurs during sample handling and processing. Methods Four laboratories participated in this Round Robin. Laboratories screened materials to identify BPA contamination in collection and analysis materials. Serum was spiked with concentrations of uBPA and/or BPA-G ranging from 0.09-19.5 (uBPA) and 0.5-32 (BPA-G) ng/mL. Additional samples were preserved unspiked as ‘environmental’ samples. Blinded samples were provided to laboratories that used LC/MSMS to simultaneously quantify uBPA and BPA-G. To determine whether inadvertent hydrolysis of BPA metabolites occurred, samples spiked with only BPA-G were analyzed for the presence of uBPA. Finally, three laboratories compared direct and indirect methods of quantifying BPA-G. Results We identified collection materials and reagents that did not introduce BPA contamination. In the blinded spiked sample analysis, all laboratories were able to distinguish low from high values of uBPA and BPA-G, for the whole spiked sample range and for those samples spiked with the three lowest concentrations (0.5-3.1 ng/ml). By completion of the Round Robin, three laboratories had verified methods for the analysis of uBPA and two verified for the analysis of BPA-G (verification determined by: 4 of 5 samples within 20% of spiked concentrations). In the analysis of BPA-G only spiked samples, all laboratories reported BPA-G was the majority of BPA detected (92.2 – 100%). Finally, laboratories were more likely to be verified using direct methods than indirect ones using enzymatic hydrolysis. Conclusions Sensitive and accurate methods for the direct quantification of uBPA and BPA-G were developed in multiple laboratories and can be used for the analysis of human serum samples. BPA contamination can be controlled during sample collection and inadvertent hydrolysis of BPA conjugates can be avoided during sample handling. PMID:24690217
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Prevalence and causes of abnormal PSA recovery.
Lautenbach, Noémie; Müntener, Michael; Zanoni, Paolo; Saleh, Lanja; Saba, Karim; Umbehr, Martin; Velagapudi, Srividya; Hof, Danielle; Sulser, Tullio; Wild, Peter J; von Eckardstein, Arnold; Poyet, Cédric
2018-01-26
Prostate-specific antigen (PSA) test is of paramount importance as a diagnostic tool for the detection and monitoring of patients with prostate cancer. In the presence of interfering factors such as heterophilic antibodies or anti-PSA antibodies the PSA test can yield significantly falsified results. The prevalence of these factors is unknown. We determined the recovery of PSA concentrations diluting patient samples with a standard serum of known PSA concentration. Based on the frequency distribution of recoveries in a pre-study on 268 samples, samples with recoveries <80% or >120% were defined as suspect, re-tested and further characterized to identify the cause of interference. A total of 1158 consecutive serum samples were analyzed. Four samples (0.3%) showed reproducibly disturbed recoveries of 10%, 68%, 166% and 4441%. In three samples heterophilic antibodies were identified as the probable cause, in the fourth anti-PSA-autoantibodies. The very low recovery caused by the latter interference was confirmed in serum, as well as heparin- and EDTA plasma of blood samples obtained 6 months later. Analysis by eight different immunoassays showed recoveries ranging between <10% and 80%. In a follow-up study of 212 random plasma samples we found seven samples with autoantibodies against PSA which however did not show any disturbed PSA recovery. About 0.3% of PSA determinations by the electrochemiluminescence assay (ECLIA) of Roche diagnostics are disturbed by heterophilic or anti-PSA autoantibodies. Although they are rare, these interferences can cause relevant misinterpretations of a PSA test result.
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Serum factors and clinical characteristics associated with serum E-Screen activity
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
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Long-term stability of glucose: glycolysis inhibitor vs. gel barrier tubes.
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.
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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.
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Serologic Evidence of Lyssavirus Infection in Bats, Cambodia
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
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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...
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Sequences of Zika Virus Genomes from a Pediatric Cohort in Nicaragua.
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.
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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
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[Applying competitive polymerase chain reaction to the detection of hepatitis B virus DNA].
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.
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Serological tests for detecting Rift Valley fever viral antibodies in sheep from the Nile Delta.
Scott, R M; Feinsod, F M; Allam, I H; Ksiazek, T G; Peters, C J; Botros, B A; Darwish, M A
1986-01-01
To determine the accuracy of serological methods in detecting Rift Valley fever (RVF) viral antibodies, we examined serum samples obtained from 418 sheep in the Nile Delta by using five tests. The plaque reduction neutralization test (PRNT) was considered the standard serological method against which the four other tests were compared. Twenty-four serum samples had RVF viral antibodies detected by PRNT. Hemagglutination inhibition and enzyme-linked immunosorbent assay antibodies to RVF virus were also present in the same 24 serum samples. Indirect immunofluorescence was less sensitive in comparison with PRNT, and complement fixation was the least sensitive. These results extend observations made with laboratory animals to a large field-collected group of Egyptian sheep. PMID:3533977
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Duterme, Sophie; Vanhoof, Raymond; Vanderpas, Jean; Pierard, Denis; Huygen, Kris
2016-04-01
Report on the pitfalls of serodiagnosis of pertussis in Belgium for 2013 by the NRC Bordetella. Determine cases of acute infection using an anti-pertussis toxin (PT) IgG antibody ELISA. A total of 2471 serum samples were received. Clinical information on the duration of cough (at moment of blood sampling) is essential for a reliable interpretation of the results. In order to avoid false negative results, 213 samples for which this information was lacking were not tested. For a total of 2179 patients tested, 520 (23.9%) had antibody levels indicative of an acute infection, 261 (12%) samples were diagnosed as positive (indicative of a pertussis infection or vaccination during the last year), 143 (6.7%) samples were classified as doubtful and 752 (34,5%) (35.5%) were diagnosed as negative. The serodiagnosis of pertussis has limited value for the early diagnosis of the disease and PCR analysis on nasopharyngeal swabs is the method of choice during the first 2 weeks and always for young children <1 year old. For sera collected during the first 2 weeks with anti-PT levels below the threshold for acute infection, a second sample collected 2-3 weeks later is needed a definitive diagnosis. For 503 (23.0%) early samples, a second serum sample was requested but not provided. For 85 patients, for whom a second sample was received, 12.9% were eventually diagnosed as having an acute infection. In order to generate reliable serodiagnostic results for pertussis, serum samples should preferentially be collected 3 weeks after onset of symptoms.
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The effect of sample storage on the performance and reproducibility of the galactomannan EIA test.
Kimpton, George; White, P Lewis; Barnes, Rosemary A
2014-08-01
Galactomannan enzyme immune assay (GM EIA) is a nonculture test for detecting invasive aspergillosis (IA) forming a key part of diagnosis and management. Recent reports have questioned the reproducibility of indices after sample storage. To investigate this, 198 serum samples (72 from cases and 126 from controls) and 61 plasma samples (24 from cases and 37 from controls), initially tested between 2010 and 2013, were retested to determine any change in index. Data were also collected on circulatory protein levels for false-positive serum samples. Serum indices significantly declined on retesting (median: initial, 0.50, retest, 0.23; P < 0.0001). This was shown to be diagnosis dependent as the decline was apparent on retesting of control samples (median: initial 0.50, retest 0.12; P < 0.0001), but was not evident with case samples (median: initial, 0.80, retest, 0.80; P = 0.724). Plasma samples showed little change on reanalysis after long-term storage at 4°C. Retesting after freezing showed a decrease in index values for controls (median: initial 0.40, retest 0.26; P = 0.0505), but no significant change in cases. Circulatory proteins showed a correlation between serum albumin concentration and difference in index value on retesting. Overall, this study suggests that a lack of reproducibility in GM EIA positivity is only significant when disease is absent. Retesting after freezing helps to differentiate false-positive GM EIA results and, with consecutive positivity, could help to improve accuracy in predicting disease status. The freezing of samples prior to testing could potentially reduce false-positivity rates and the need to retest. © The Author 2014. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
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Lorusso, Alessio; Baba, Doumbia; Spedicato, Massimo; Teodori, Liana; Bonfini, Barbara; Marcacci, Maurilia; Di Provvido, Andrea; Isselmou, Katia; Marini, Valeria; Carmine, Irene; Scacchia, Massimo; Di Sabatino, Daria; Petrini, Antonio; Bezeid, Beyatt Ahmed; Savini, Giovanni
2016-06-01
In March 2013, EDTA-blood and serum samples were collected from 119 cattle and 159 dromedaries at the slaughterhouse of Nouakchott, the capital city of the Islamic Republic of Mauritania. Serum samples were screened for the presence of Bluetongue (BT) antibodies by competitive ELISA (cELISA). Positive samples were then tested by serum-neutralization (SN) to determine BTV serotype. RNA from blood samples was first tested by a genus-specific quantitative RT-PCR assay which is able to detect all 27 existing BTV serotypes (RT-qPCR1-27). Positive samples were further screened by a RT-qPCR assay which, instead, is able to detect the classical 24 BTV serotypes only (RT-qPCR1-24). Of the 278 serum samples tested, 177 (mean=63.7%; 95% CI: 57.9%-69.1%) resulted positive by cELISA. Of these, 69 were from cattle (mean=58.0%; 95% CI: 49.0%-66.5%) and 108 from dromedaries (mean=67.9%; 95% CI: 60.3%-74.7%). BTV-26 neutralizing antibodies were by far the most frequently found as they were detected in 146 animals with titres ranging from 1:10 to 1:80. Out of 278 blood samples, 25 (mean=9.0%; 95% CI: 6.2%-12.9%) were found positive for BTV by RT-qPCR1-27, 20 (mean=16.8%; 95% CI: 11.2%-24.6%) were from cattle and 5 (mean=3.1%; 95% CI: 1.4%-7.1%) from dromedaries. When tested by RT-qPCR1-24 the 25 BTV positive samples were negative. Unfortunately, no genetic information by molecular typing or by next generation sequencing has been obtained as for the very low levels of RNA in the blood samples. Copyright © 2016 Elsevier B.V. All rights reserved.
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[Clinical evaluation of a novel HBsAg quantitative assay].
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.
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Giménez-Lirola, Luis G; Mur, Lina; Rivera, Belen; Mogler, Mark; Sun, Yaxuan; Lizano, Sergio; Goodell, Christa; Harris, D L Hank; Rowland, Raymond R R; Gallardo, Carmina; Sánchez-Vizcaíno, José Manuel; Zimmerman, Jeff
2016-01-01
In the absence of effective vaccine(s), control of African swine fever caused by African swine fever virus (ASFV) must be based on early, efficient, cost-effective detection and strict control and elimination strategies. For this purpose, we developed an indirect ELISA capable of detecting ASFV antibodies in either serum or oral fluid specimens. The recombinant protein used in the ELISA was selected by comparing the early serum antibody response of ASFV-infected pigs (NHV-p68 isolate) to three major recombinant polypeptides (p30, p54, p72) using a multiplex fluorescent microbead-based immunoassay (FMIA). Non-hazardous (non-infectious) antibody-positive serum for use as plate positive controls and for the calculation of sample-to-positive (S:P) ratios was produced by inoculating pigs with a replicon particle (RP) vaccine expressing the ASFV p30 gene. The optimized ELISA detected anti-p30 antibodies in serum and/or oral fluid samples from pigs inoculated with ASFV under experimental conditions beginning 8 to 12 days post inoculation. Tests on serum (n = 200) and oral fluid (n = 200) field samples from an ASFV-free population demonstrated that the assay was highly diagnostically specific. The convenience and diagnostic utility of oral fluid sampling combined with the flexibility to test either serum or oral fluid on the same platform suggests that this assay will be highly useful under the conditions for which OIE recommends ASFV antibody surveillance, i.e., in ASFV-endemic areas and for the detection of infections with ASFV isolates of low virulence.
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Analysis of biofluids in aqueous environment based on mid-infrared spectroscopy.
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.
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Virological Survey in free-ranging wildcats (Felis silvestris) and feral domestic cats in Portugal.
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.
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Serum fibroblast growth factor 23 concentrations in dogs with chronic kidney disease.
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.
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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
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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
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NHEXAS PHASE I MARYLAND STUDY--PESTICIDES IN BLOOD ANALYTICAL RESULTS
The Pesticides in Blood Serum data set contains analytical results for measurements of up to 17 pesticides in 358 blood samples over 79 households. Each sample was collected via a venous sample from the primary respondent within each household by a phlebotomist. Samples were ge...
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Analysis of serum and cerebrospinal fluid in clinically normal adult miniature donkeys.
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.
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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
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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
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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.
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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.
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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.
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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
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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.
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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.
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Guthmann, Jean-Paul; Klovstad, Hilde; Boccia, Delia; Hamid, Nuha; Pinoges, Loretxu; Nizou, Jacques-Yves; Tatay, Mercedes; Diaz, Francisco; Moren, Alain; Grais, Rebecca Freeman; Ciglenecki, Iza; Nicand, Elisabeth; Guerin, Philippe Jean
2006-06-15
The conflict in Darfur, Sudan, was responsible for the displacement of 1.8 million civilians. We investigated a large outbreak of hepatitis E virus (HEV) infection in Mornay camp (78,800 inhabitants) in western Darfur. To describe the outbreak, we used clinical and demographic information from cases recorded at the camp between 26 July and 31 December 2004. We conducted a case-cohort study and a retrospective cohort study to identify risk factors for clinical and asymptomatic hepatitis E, respectively. We collected stool and serum samples from animals and performed a bacteriological analysis of water samples. Human samples were tested for immunoglobulin G and immunoglobulin M antibody to HEV (for serum samples) and for amplification of the HEV genome (for serum and stool samples). In 6 months, 2621 hepatitis E cases were recorded (attack rate, 3.3%), with a case-fatality rate of 1.7% (45 deaths, 19 of which involved were pregnant women). Risk factors for clinical HEV infection included age of 15-45 years (odds ratio, 2.13; 95% confidence interval, 1.02-4.46) and drinking chlorinated surface water (odds ratio, 2.49; 95% confidence interval, 1.22-5.08). Both factors were also suggestive of increased risk for asymptomatic HEV infection, although this was not found to be statistically significant. HEV RNA was positively identified in serum samples obtained from 2 donkeys. No bacteria were identified from any sample of chlorinated water tested. Current recommendations to ensure a safe water supply may have been insufficient to inactivate HEV and control this epidemic. This research highlights the need to evaluate current water treatment methods and to identify alternative solutions adapted to complex emergencies.
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Molecular detection of Rift Valley fever virus in serum samples from selected areas of Tanzania.
Chengula, Augustino Alfred; Kasanga, Christopher Jacob; Mdegela, Robinson Hammerthon; Sallu, Raphael; Yongolo, Mmeta
2014-04-01
Rift Valley fever (RVF) is an acute mosquito-borne viral zoonotic disease affecting domestic animals and humans caused by the Rift Valley fever virus (RVFV). The virus belongs to the genus Phlebovirus of the family Bunyaviridae. The main aim of this study was to detect the presence of antibodies to RVFV as well as the virus in the serum samples that were collected from livestock during the 2006/2007 RVF outbreaks in different locations in Tanzania. Analysis of selected samples was done using a RVF-specific inhibition enzyme-linked immunosorbent assay (I-ELISA) and reverse transcription polymerase chain reaction (RT-PCR). Genomic viral RNA was extracted directly from serum samples using a QIAamp Viral RNA Mini Kit (QIAGEN), and a one-step RT-PCR protocol was used to amplify the S segment of RVFV. Positive results were obtained in 39.5% (n = 200) samples using the RVF I-ELISA, and 17.6% (n = 108) of samples were positive by RT-PCR. I-ELISA detected 41 (38.7%), 32 (39.0%), and 6 (50.0%) positive results in cattle, goats, and sheep sera, respectively, whereas the RT-PCR detected 11 (0.2%), 7 (0.2%), and 1 (0.1%) positive results in cattle, goats, and sheep sera, respectively. These findings have demonstrated the presence of RVFV in Tanzania during the 2006/2007 RVF outbreaks. To our knowledge, this is the first report to detect RVFV in serum samples from domestic animals in Tanzania using PCR technique. Therefore, a detailed molecular study to characterize the virus from different geographical locations in order to establish the profile of strains circulating in the country and develop more effective and efficient control strategies should be done.
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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.
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Detection of autoantibodies against thyroid peroxidase in serum samples of hypothyroid dogs.
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.
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Jemima, Ebenezer Angel; Manoharan, Seeralan; Kumanan, Kathaperumal
2014-08-01
The measurement of neutralizing antibodies induced by the glycoprotein of rabies virus is indispensable for assessing the level of neutralizing antibodies in animals or humans. A rapid fluorescent focus inhibition test (RFFIT) has been approved by WHO and is the most widely used method to measure the virus-neutralizing antibody content in serum, but a rapid test system would be of great value to screen large numbers of serum samples. To develop and evaluate a latex agglutination test (LAT) for measuring rabies virus antibodies, a recombinant glycoprotein was expressed in an insect cell system and purified, and the protein was coated onto latex beads at concentrations of 0.1, 0.25, 0.5, 0.75, and 1 mg/ml to find out the optimal concentration for coating latex beads. It was found that 0.5 mg/ml of recombinant protein was optimal for coating latex beads, and this concentration was used to sensitize the latex beads for screening of dog serum samples. Grading of LAT results was done with standard reference serum with known antibody titers. A total of 228 serum samples were tested, out of which 145 samples were positive by both RFFIT and LAT, and the specificity was found to be 100 %. In RFFIT, 151 samples were positive, the sensitivity was found to be 96.03 %, and the accuracy/concordance was found to be 97.39 %. A rapid field test-a latex agglutination test (LAT)-was developed and evaluated for rabies virus antibody assessment using recombinant glycoprotein of rabies virus expressed in an insect cell system.
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Long-term in vitro reactivity for HLA antibodies and comparison of detection using serum vs. plasma
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
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Petrini, Stefano; Pierini, Ilaria; Giammarioli, Monica; Feliziani, Francesco; De Mia, Gian Mario
2017-03-01
We evaluated the use of oral fluid as an alternative to serum samples for Classical swine fever virus (CSFV) detection. Individual oral fluid and serum samples were collected at different times post-infection from pigs that were experimentally inoculated with CSFV Alfort 187 strain. We found no evidence of CSFV neutralizing antibodies in swine oral fluid samples under our experimental conditions. In contrast, real-time reverse transcription-polymerase chain reaction could detect CSFV nucleic acid from the oral fluid as early as 8 d postinfection, which also coincided with the time of initial detection in blood samples. The probability of CSFV detection in oral fluid was identical or even higher than in the corresponding blood sample. Our results support the feasibility of using this sampling method for CSFV genome detection, which may represent an additional cost-effective tool for CSF control.
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NHEXAS PHASE I MARYLAND STUDY--QA ANALYTICAL RESULTS FOR PESTICIDES IN SPIKE SAMPLES
The Pesticides in Spikes data set contains the analytical results of measurements of up to 17 pesticides in 12 control samples (spikes) from 11 households. Measurements were made in samples of blood serum. Controls were used to assess recovery of target analytes from a sample m...
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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
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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.
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Association between serum soluble CD30 and serum creatinine before and after renal transplantation.
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.
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Protein electrophoresis in cranes with presumed insect bite.
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.
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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.
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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.
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Aspergillus colonization in patients with bronchogenic carcinoma.
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.
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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.
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Influenza A Virus Surveillance Based on Pre-Weaning Piglet Oral Fluid Samples.
Panyasing, Y; Goodell, C; Kittawornrat, A; Wang, C; Levis, I; Desfresne, L; Rauh, R; Gauger, P C; Zhang, J; Lin, X; Azeem, S; Ghorbani-Nezami, S; Yoon, K-J; Zimmerman, J
2016-10-01
Influenza A virus (IAV) surveillance using pre-weaning oral fluid samples from litters of piglets was evaluated in four ˜12 500 sow and IAV-vaccinated, breeding herds. Oral fluid samples were collected from 600 litters and serum samples from their dams at weaning. Litter oral fluid samples were tested for IAV by virus isolation, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), RT-PCR subtyping and sequencing. Commercial nucleoprotein (NP) enzyme-linked immunosorbent assay (ELISA) kits and NP isotype-specific assays (IgM, IgA and IgG) were used to characterize NP antibody in litter oral fluid and sow serum. All litter oral fluid specimens (n = 600) were negative by virus isolation. Twenty-five oral fluid samples (25/600 = 4.2%) were qRT-PCR positive based on screening (Laboratory 1) and confirmatory testing (Laboratory 2). No hemagglutinin (HA) and neuraminidase (NA) gene sequences were obtained, but matrix (M) gene sequences were obtained for all qRT-PCR-positive samples submitted for sequencing (n = 18). Genetic analysis revealed that all M genes sequences were identical (GenBank accession no. KF487544) and belonged to the triple reassortant influenza A virus M gene (TRIG M) previously identified in swine. The proportion of IgM- and IgA-positive samples was significantly higher in sow serum and litter oral fluid samples, respectively (P < 0.01). Consistent with the extensive use of IAV vaccine, no difference was detected in the proportion of IgG- and blocking ELISA-positive sow serum and litter oral fluids. This study supported the use of oral fluid sampling as a means of conducting IAV surveillance in pig populations and demonstrated the inapparent circulation of IAV in piglets. Future work on IAV oral fluid diagnostics should focus on improved procedures for virus isolation, subtyping and sequencing of HA and NA genes. The role of antibody in IAV surveillance remains to be elucidated, but longitudinal assessment of specific antibody has the potential to provide information regarding patterns of infection, vaccination status and herd immunity. © 2014 Blackwell Verlag GmbH.
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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.
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Comparison of lacosamide concentrations in cerebrospinal fluid and serum in patients with epilepsy.
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.
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p16 promoter hypermethylation: A useful serum marker for early detection of gastric cancer
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
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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.
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Daily rhythm of salivary and serum urea concentration in sheep
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
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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.
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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.
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NASA Astrophysics Data System (ADS)
Wang, Chun Wei; Manne, Upender; Reddy, Vishnu B.; Oelschlager, Denise K.; Katkoori, Venkat R.; Grizzle, William E.; Kapoor, Rakesh
2010-11-01
A combination tapered fiber-optic biosensor (CTFOB) dip probe for rapid and cost-effective quantification of proteins in serum samples has been developed. This device relies on diode laser excitation and a charged-coupled device spectrometer and functions on a technique of sandwich immunoassay. As a proof of principle, this technique was applied in a quantitative estimation of interleukin IL-6. The probes detected IL-6 at picomolar levels in serum samples obtained from a patient with lupus, an autoimmune disease, and a patient with lymphoma. The estimated concentration of IL-6 in the lupus sample was 5.9 +/- 0.6 pM, and in the lymphoma sample, it was below the detection limit. These concentrations were verified by a procedure involving bead-based xMAP technology. A similar trend in the concentrations was observed. The specificity of the CTFOB dip probes was assessed by analysis with receiver operating characteristics. This analysis suggests that the dip probes can detect 5-pM or higher concentration of IL-6 in these samples with specificities of 100%. The results provide information for guiding further studies in the utilization of these probes to quantify other analytes in body fluids with high specificity and sensitivity.
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NASA Astrophysics Data System (ADS)
Stosnach, Hagen; Mages, Margarete
2009-04-01
In clinical service laboratories, one of the most common analytical tasks with regard to inorganic traces is the determination of the nutrition-relevant elements Fe, Cu, Zn, and Se. Because of the high numbers of samples and the commercial character of these analyses, a time-consuming sample preparation must be avoided. In this presentation, the results of total reflection X-ray fluorescence measurements with a low-power system and different sample preparation procedures are compared with those derived from analysis with common methods like Atomic Absorption Spectroscopy (AAS) and Inductively Coupled Plasma Mass Spectroscopy (ICP-MS). The results of these investigations indicate that the optimal total reflection X-ray fluorescence analysis of the nutrition-relevant elements Fe, Cu, Zn, and Se can be performed by preparing whole blood and serum samples after dilution with ultrapure water and transferring 10 μl of internally standardized sample to an unsiliconized quartz glass sample carrier with subsequent drying in a laboratory oven. Suitable measurement time was found to be 600 s. The enhanced sample preparation by means of microwave or open digestion, in parts combined with cold plasma ashing, led to an improvement of detection limits by a factor of 2 for serum samples while for whole blood samples an improvement was only observed for samples prepared by means of microwave digestion. As the matrix elements P, S, Cl, and for whole blood Fe have a major influence on the detection limits, most probably a further enhancement of analytical quality requires the removal of the organic matrix. However, for the routine analysis of the nutrition-relevant elements, the dilution preparation was found to be sufficient.
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Biagini, R E; Sammons, D L; Smith, J P; Page, E H; Snawder, J E; Striley, C A F; MacKenzie, B A
2004-08-01
To evaluate potential exposure to Bacillis anthracis (Ba) spores in sampling/decontamination workers in the aftermath of an anthrax terror attack. Fifty six serum samples were obtained from workers involved in environmental sampling for Ba spores at the American Media, Inc. (AMI) building in Boca Raton, FL after the anthrax attack there in October 2001. Nineteen sera were drawn from individuals both pre-entry and several weeks after entrance into the building. Nine sera each were drawn from unique individuals at the pre-entry and follow up blood draws. Thirteen donor control sera were also evaluated. Individuals were surveyed for Ba exposure by measurement of serum Ba anti-protective antigen (PA) specific IgG antibodies using a newly developed fluorescent covalent microsphere immunoassay (FCMIA). Four sera gave positive anti-PA IgG results (defined as anti-PA IgG concentrations > or = the mean microg/ml anti-PA IgG from donor control sera (n = 13 plus 2 SD which were also inhibited > or = 85% when the serum was pre-adsorbed with PA). The positive sera were the pre-entry and follow up samples of two workers who had received their last dose of anthrax vaccine in 2000. It appears that the sampling/decontamination workers of the present study either had insufficient exposure to Ba spores to cause the production of anti-PA IgG antibodies or they were exposed to anthrax spores without producing antibody. The FCMIA appears to be a fast, sensitive, accurate, and precise method for the measurement of anti-PA IgG antibodies.
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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.
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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
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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.
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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
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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
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Afridi, Hassan Imran; Kazi, Tasneem Gul; Kazi, Naveed; Kandhro, Ghulam Abbas; Baig, Jameel Ahmed; Shah, Abdul Qadir; Khan, Sumaira; Kolachi, Nida Fatima; Wadhwa, Sham Kumar; Shah, Faheem
2011-01-01
The most common cause of blindness in developing countries is vitamin A deficiency. The World Health Organization (WHO) estimates 13.8 million children have some degree of visual loss related to vitamin A deficiency. The causes of night blindness in children are multifactorial and particular consideration has been given to childhood nutritional deficiency, which is the most common problem found in underdeveloped countries. Such deficiency can result in physiological and pathological processes that in turn influence biological sample composition. Vitamin and mineral deficiency prevents more than two billion people from achieving their full intellectual and physical potential. This study was designed to compare the levels of magnesium (Mg), calcium (Ca), potassium (K), and sodium (Na) in scalp hair, serum, blood, and urine of night blindness children in two age groups, (1-5) and (6-10) years, of both genders comparing them to sex- and age-matched controls. A microwave assisted wet acid digestion procedure was developed as a sample pretreatment for the determination of Mg, Ca, K, and Na in biological samples of children with night blindness. The proposed method was validated by using conventional wet digestion and certified reference samples of hair, serum, blood, and urine. The digests of all biological samples were analysed for Mg, Ca, K, and Na by flame atomic absorption spectrometry (FAAS) using an air/acetylene flame. The results indicated significantly lower levels of Mg, Ca, and K in the biological samples (blood, serum, and scalp hair) of male and female children with night blindness and higher values of Na compared with control subjects of both genders. These data present guidance to clinicians and other professionals investigating deficiency of essential mineral elements in biological samples (scalp hair, serum, and blood) of children with night blindness.
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NHEXAS PHASE I MARYLAND STUDY--QA ANALYTICAL RESULTS FOR PESTICIDE METABOLITES IN SPIKE SAMPLES
The Pesticides in Spikes data set contains the analytical results of measurements of up to 17 pesticides in 12 control samples (spikes) from 11 households. Measurements were made in samples of blood serum. Controls were used to assess recovery of target analytes from a sample m...
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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.
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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.
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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.
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Headspace Solid Phase Micro Extraction Gas Chromatographic Determination of Fenthion in Human Serum
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
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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.
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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.
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A Systematic Evaluation of Blood Serum and Plasma Pre-Analytics for Metabolomics Cohort Studies
Jobard, Elodie; Trédan, Olivier; Postoly, Déborah; André, Fabrice; Martin, Anne-Laure; Elena-Herrmann, Bénédicte; Boyault, Sandrine
2016-01-01
The recent thriving development of biobanks and associated high-throughput phenotyping studies requires the elaboration of large-scale approaches for monitoring biological sample quality and compliance with standard protocols. We present a metabolomic investigation of human blood samples that delineates pitfalls and guidelines for the collection, storage and handling procedures for serum and plasma. A series of eight pre-processing technical parameters is systematically investigated along variable ranges commonly encountered across clinical studies. While metabolic fingerprints, as assessed by nuclear magnetic resonance, are not significantly affected by altered centrifugation parameters or delays between sample pre-processing (blood centrifugation) and storage, our metabolomic investigation highlights that both the delay and storage temperature between blood draw and centrifugation are the primary parameters impacting serum and plasma metabolic profiles. Storing the blood drawn at 4 °C is shown to be a reliable routine to confine variability associated with idle time prior to sample pre-processing. Based on their fine sensitivity to pre-analytical parameters and protocol variations, metabolic fingerprints could be exploited as valuable ways to determine compliance with standard procedures and quality assessment of blood samples within large multi-omic clinical and translational cohort studies. PMID:27929400
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Yılmaz, Cansu; Köksal, Nilgün; Özkan, Hilal; Dorum, Bayram Ali; Bağcı, Onur
2017-01-01
Yılmaz C, Köksal N, Özkan H, Dorum BA, Bağcı O. Low serum IGF-1 and increased cytokine levels in tracheal aspirate samples are associated with bronchopulmonary dysplasia. Turk J Pediatr 2017; 59: 122-129. Despite developments in the perinatal and neonatal care, bronchopulmonary dysplasia (BPD) is still the most frequently seen long-term complication in preterm infants. The aim of this prospective study is to investigate the association between the development of BPD and serial measurements of IGF-1 levels and their relationship with levels of IGF-1 and cytokine in tracheal aspirate fluids. A total of 40 premature infants, born at a gestational age of ≤ 32 weeks, were enrolled in the study. On postnatal day-1, 3, 7, 21 and 28 serum IGF-1 levels and IGF-1 levels, IL-6, IL-8, IL-10 and TNF-alpha levels in tracheal aspirate fluid samples of intubated cases were examined. Mean gestational age of 40 patients included in the study was 29.41 ± 2.23 weeks, and their mean birth weight was 1,256.85 ± 311.48 g. BPD was detected in 35% of cases. Mean gestational week and birth weight of the cases that developed BPD were 30 ± 3 weeks and 1,150 ± 295 g, respectively. Serum IGF-1 levels on postnatal day-1, 3, 7, 21 and 28 in cases who developed BPD were significantly lower when compared with those without BPD (p < 0.01). Levels of IL-6, IL-8, IL-10, and TNF-alpha in tracheal aspirate samples were significantly higher in cases with BPD compared to those without BPD (p < 0.05). IGF-1 levels in tracheal aspirate fluid samples did not differ significantly based on the presence of BPD (p > 0.05). Severity of BPD was associated with decreased serum IGF-1 levels and increased cytokine levels in tracheal aspirate samples.
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Thrombospondin Type-1 Domain-Containing 7A in Idiopathic Membranous Nephropathy
Meyer-Schwesinger, Catherine; Seitz-Polski, Barbara; Ma, Hong; Zahner, Gunther; Dolla, Guillaume; Hoxha, Elion; Helmchen, Udo; Dabert-Gay, Anne-Sophie; Debayle, Delphine; Merchant, Michael; Klein, Jon; Salant, David J.; Stahl, Rolf A.K.; Lambeau, Gérard
2014-01-01
BACKGROUND Idiopathic membranous nephropathy is an autoimmune disease. In approximately 70% of patients, it is associated with autoantibodies against the phospholipase A2 receptor 1 (PLA2R1). Antigenic targets in the remaining patients are unknown. METHODS Using Western blotting, we screened serum samples from patients with idiopathic membranous nephropathy, patients with other glomerular diseases, and healthy controls for antibodies against human native glomerular proteins. We partially purified a putative new antigen, identified this protein by means of mass spectrometry of digested peptides, and validated the results by analysis of recombinant protein expression, immunoprecipitation, and immunohistochemical analysis. RESULTS Serum samples from 6 of 44 patients in a European cohort and 9 of 110 patients in a Boston cohort with anti-PLA2R1–negative idiopathic membranous nephropathy recognized a glomerular protein that was 250 kD in size. None of the serum samples from the 74 patients with idiopathic membranous nephropathy who were sero-positive for anti-PLA2R1 antibodies, from the 76 patients with other glomerular diseases, and from the 44 healthy controls reacted against this antigen. Although this newly identified antigen is clearly different from PLA2R1, it shares some biochemical features, such as N-glycosylation, membranous location, and reactivity with serum only under nonreducing conditions. Mass spectrometry identified this antigen as thrombospondin type-1 domain-containing 7A (THSD7A). All reactive serum samples recognized recombinant THSD7A and immunoprecipitated THSD7A from glomerular lysates. Moreover, immunohistochemical analyses of biopsy samples from patients revealed localization of THSD7A to podocytes, and IgG eluted from one of these samples was specific for THSD7A. CONCLUSIONS In our cohort, 15 of 154 patients with idiopathic membranous nephropathy had circulating autoantibodies to THSD7A but not to PLA2R1, a finding that suggests a distinct subgroup of patients with this condition. (Funded by the French National Center for Scientific Research and others.) PMID:25394321
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Some comments on the effectiveness of the therapy for Β-thalassemia
NASA Astrophysics Data System (ADS)
Jiang, K.; Ma, W. Y.; Ortalli, I.; Pedrazzi, G.; Zhang, X.; Izzi, G. C.
1994-12-01
In the present study, twenty-five samples of red blood cells (RBCs) and two samples of lyophilized serum, drawn from thirteen patients with Β-thanassemia major, have been examined by Mössbauer spectroscopy. All these patients undergo long-term therapy by regular blood transfusion and deferoxamine. The samples were obtained at the end of one cycle of treatments, before the successive blood transfusion. The results show, within the experimental errors, that the ferritin-like iron appears to be absent in the RBCs of the patients but it is still present in the serum.
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Severe reactions from roe without concomitant fish allergy.
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.
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Erythrocyte migration and gap formation in rabbit blood clots in vitro.
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.
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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
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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
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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.
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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.
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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.
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Parathyroid hormone related protein concentration in human serum and CSF correlates with age.
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.
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Hart, Leslie B; Wells, Randall S; Kellar, Nick; Balmer, Brian C; Hohn, Aleta A; Lamb, Stephen V; Rowles, Teri; Zolman, Eric S; Schwacke, Lori H
2015-01-01
Inshore common bottlenose dolphins (Tursiops truncatus) are exposed to a broad spectrum of natural and anthropogenic stressors. In response to these stressors, the mammalian adrenal gland releases hormones such as cortisol and aldosterone to maintain physiological and biochemical homeostasis. Consequently, adrenal gland dysfunction results in disruption of hormone secretion and an inappropriate stress response. Our objective herein was to develop diagnostic reference intervals (RIs) for adrenal hormones commonly associated with the stress response (i.e., cortisol, aldosterone) that account for the influence of intrinsic (e.g., age, sex) and extrinsic (e.g., time) factors. Ultimately, these reference intervals will be used to gauge an individual's response to chase-capture stress and could indicate adrenal abnormalities. Linear mixed models (LMMs) were used to evaluate demographic and sampling factors contributing to differences in serum cortisol and aldosterone concentrations among bottlenose dolphins sampled in Sarasota Bay, Florida, USA (2000-2012). Serum cortisol concentrations were significantly associated with elapsed time from initial stimulation to sample collection (p<0.05), and RIs were constructed using nonparametric methods based on elapsed sampling time for dolphins sampled in less than 30 minutes following net deployment (95% RI: 0.91-4.21 µg/dL) and following biological sampling aboard a research vessel (95% RI: 2.32-6.68 µg/dL). To examine the applicability of the pre-sampling cortisol RI across multiple estuarine stocks, data from three additional southeast U.S. sites were compared, revealing that all of the dolphins sampled from the other sites (N = 34) had cortisol concentrations within the 95th percentile RI. Significant associations between serum concentrations of aldosterone and variables reported in previous studies (i.e., age, elapsed sampling time) were not observed in the current project (p<0.05). Also, approximately 16% of Sarasota Bay bottlenose dolphin aldosterone concentrations were below the assay's detection limit (11 pg/mL), thus hindering the ability to derive 95th percentile RIs. Serum aldosterone concentrations from animals sampled at the three additional sites were compared to the detection limit, and the proportion of animals with low aldosterone concentrations was not significantly different than an expected prevalence of 16%. Although this study relied upon long-term, free-ranging bottlenose dolphin health data from a single site, the objective RIs can be used for future evaluation of adrenal function among individuals sampled during capture-release health assessments.
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Comparative analysis of EV isolation procedures for miRNAs detection in serum samples.
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.
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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.
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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
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The hook effect in calcitonin immunoradiometric assay: a case report.
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.
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Protein microarray analysis reveals BAFF-binding autoantibodies in systemic lupus erythematosus
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
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Introduction of a rapid, simple radioimmunoassay and quality control scheme for thyroxine.
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
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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.
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Johns, Jennifer L.; Moorhead, Kaitlin A.; Hu, Jing; Moorhead, Roberta C.
2018-01-01
Clinical pathology testing of rodents is often challenging due to insufficient sample volume. One solution in clinical veterinary and exploratory research environments is dilution of samples prior to analysis. However, published information on the impact of preanalytical sample dilution on rodent biochemical data is incomplete. The objective of this study was to evaluate the effects of preanalytical sample dilution on biochemical analysis of mouse and rat serum samples utilizing the Siemens Dimension Xpand Plus. Rats were obtained from end of study research projects. Mice were obtained from sentinel testing programs. For both, whole blood was collected via terminal cardiocentesis into empty tubes and serum was harvested. Biochemical parameters were measured on fresh and thawed frozen samples run straight and at dilution factors 2–10. Dilutions were performed manually, utilizing either ultrapure water or enzyme diluent per manufacturer recommendations. All diluted samples were generated directly from the undiluted sample. Preanalytical dilution caused clinically unacceptable bias in most analytes at dilution factors four and above. Dilution-induced bias in total calcium, creatinine, total bilirubin, and uric acid was considered unacceptable with any degree of dilution, based on the more conservative of two definitions of acceptability. Dilution often caused electrolyte values to fall below assay range precluding evaluation of bias. Dilution-induced bias occurred in most biochemical parameters to varying degrees and may render dilution unacceptable in the exploratory research and clinical veterinary environments. Additionally, differences between results obtained at different dilution factors may confound statistical comparisons in research settings. Comparison of data obtained at a single dilution factor is highly recommended. PMID:29497614
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Kataoka, H; Narimatsu, S; Lord, H L; Pawliszyn, J
1999-10-01
The technique of automated in-tube solid-phase microextraction (SPME) coupled with liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) was evaluated for the determination of beta-blockers in urine and serum samples. In-tube SPME is an extraction technique for organic compounds in aqueous samples, in which analytes are extracted from the sample directly into an open tubular capillary by repeated draw/eject cycles of sample solution. LC/MS analyses of beta-blockers were initially performed by liquid injection onto a LC column. Nine beta-blockers tested in this study gave very simple ESI mass spectra, and strong signals corresponding to [M + H]+ were observed for all beta-blockers. The beta-blockers were separated with a Hypersil BDS C18 column using acetonitrile/methanol/water/acetic acid (15:15:70:1) as a mobile phase. To optimize the extraction of beta-blockers, several in-tube SPME parameters were examined. The optimum extraction conditions were 15 draw/eject cycles of 30 microL of sample in 100 mM Tris-HCl (pH 8.5) at a flow rate of 100 microL/min using an Omegawax 250 capillary (Supelco, Bellefonte, PA). The beta-blockers extracted by the capillary were easily desorbed by mobile-phase flow, and carryover of beta-blockers was not observed. Using in-tube SPME/LC/ESI-MS with selected ion monitoring, the calibration curves of beta-blockers were linear in the range from 2 to 100 ng/mL with correlation coefficients above 0.9982 (n = 18) and detection limits (S/N = 3) of 0.1-1.2 ng/mL. This method was successfully applied to the analysis of biological samples without interference peaks. The recoveries of beta-blockers spiked into human urine and serum samples were above 84 and 71%, respectively. A serum sample from a patient administrated propranolol was analyzed using this method and both propranolol and its metabolites were detected.
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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.
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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.
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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
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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.
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Prevalence of hepatitis A viral RNA and antibodies among Chinese blood donors.
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.
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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
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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.
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Fast detection of Piscirickettsia salmonis in Salmo salar serum through MALDI-TOF-MS profiling.
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.
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Reassessment of the Access Testosterone chemiluminescence assay and comparison with LC-MS method.
Dittadi, Ruggero; Matteucci, Mara; Meneghetti, Elisa; Ndreu, Rudina
2018-03-01
To reassess the imprecision and Limit of Quantitation, to evaluate the cross-reaction with dehydroepiandrosterone-sulfate (DHEAS), the accuracy toward liquid chromatography-mass spectrometry (LC-MS) and the reference interval of the Access Testosterone method, performed by DxI immunoassay platform (Beckman Coulter). Imprecision was evaluated testing six pool samples assayed in 20 different run using two reagents lots. The cross-reaction with DHEAS was studied both by a displacement curve and by spiking DHEAS standard in two serum samples with known amount of testosterone. The comparison with LC-MS was evaluated by Passing-Bablock analysis in 21 routine serum samples and 19 control samples from an External Quality Assurance (EQA) scheme. The reference interval was verified by an indirect estimation on 2445 male and 2838 female outpatients. The imprecision study showed a coefficient of variation (CV) between 2.7% and 34.7% for serum pools from 16.3 and 0.27 nmol/L. The value of Limit of Quantitation at 20% CV was 0.53 nmol/L. The DHEAS showed a cross-reaction of 0.0074%. A comparison with LC-MS showed a trend toward a slight underestimation of immunoassay vs LC-MS (Passing-Bablock equations: DxI=-0.24+0.906 LCMS in serum samples and DxI=-0.299+0.981 LCMS in EQA samples). The verification of reference interval showed a 2.5th-97.5th percentile distribution of 6.6-24.3 nmol/L for male over 14 years and <0.5-2.78 nmol/L for female subjects, in accord with the reference intervals reported by the manufacturer. The Access Testosterone method could be considered an adequately reliable tool for the testosterone measurement. © 2017 Wiley Periodicals, Inc.
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Panyasing, Y; Goodell, C K; Wang, C; Kittawornrat, A; Prickett, J R; Schwartz, K J; Ballagi, A; Lizano, S; Zimmerman, J J
2014-04-01
In commercial swine populations, influenza is an important component of the porcine respiratory disease complex (PRDC) and a pathogen with major economic impact. Previously, a commercial blocking ELISA (FlockChek(™) Avian Influenza Virus MultiS-Screen(®) Antibody Test Kit, IDEXX Laboratories, Inc., Westbrook, ME, USA) designed to detect influenza A nucleoprotein (NP) antibodies in avian serum was shown to accurately detect NP antibodies in swine serum. The purpose of this study was to determine whether this assay could detect NP antibodies in swine oral fluid samples. Initially, the procedure for performing the NP-blocking ELISA on oral fluid was modified from the serum testing protocol by changing sample dilution, sample volume, incubation time and incubation temperature. The detection of NP antibody was then evaluated using pen-based oral fluid samples (n = 182) from pigs inoculated with either influenza A virus subtype H1N1 or H3N2 under experimental conditions and followed for 42 days post inoculation (DPI). NP antibodies in oral fluid were detected from DPI 7 to 42 in all inoculated groups, that is, the mean sample-to-negative (S/N) ratio of influenza-inoculated pigs was significantly different (P < 0.0001) from uninoculated controls (unvaccinated or vaccinated-uninoculated groups) through this period. Oral fluid versus serum S/N ratios from the same pen showed a correlation of 0.796 (Pearson's correlation coefficient, P < 0.0001). The results showed that oral fluid samples from influenza virus-infected pigs contained detectable levels of NP antibodies for ≥42 DPI. Future research will be required to determine whether this approach could be used to monitor the circulation of influenza virus in commercial pig populations. © 2012 Blackwell Verlag GmbH.
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Analyte stability during the total testing process: studies of vitamins A, D and E by LC-MS/MS.
Albahrani, Ali A; Rotarou, Victor; Roche, Peter J; Greaves, Ronda F
2016-10-01
There are limited evidence based studies demonstrating the stability of fat-soluble vitamins (FSV) measured in blood. This study aimed to examine the effects of light, temperature and time on vitamins A, D and E throughout the total testing process. Four experiments were conducted. Three investigated the sample matrix, of whole blood, serum and the extracted sample, against the variables of temperature and light; and the fourth experiment investigated the sample during the extraction process against the variable of light. All samples were analysed via our simultaneous FSV method using liquid chromatography-tandem mass spectrometry technology. The allowable clinical percentage change was calculated based on biological variation and desirable method imprecision for each analyte. The total change limit was ±7.3% for 25-OH-vitamin D3, ±11.8% for retinol and ±10.8% for α-tocopherol. Vitamins D and E were stable in the investigated conditions (concentration changes <4%) in the pre-analytical and analytical stages. Vitamin A showed photosensitivity in times >48 h with concentration changes of -6.8% (blood) and -6.5% (serum), both are within the allowable clinical percentage change. By contrast, the extracted retinol sample demonstrated a concentration change of -18.4% after 48 h of light exposure. However, vitamin A in the serum and extracted solution was stable for one month when stored at -20°C. Blood samples for vitamins D and E analyses can be processed in normal laboratory conditions of lighting and temperature. The required conditions for vitamin A analysis are similar when performed within 48 h. For longer-term storage, serum and vitamin A extracts should be stored at -20°C.
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Marques, Sara S.; Magalhães, Luís M.; Tóth, Ildikó V.; Segundo, Marcela A.
2014-01-01
Total antioxidant capacity assays are recognized as instrumental to establish antioxidant status of biological samples, however the varying experimental conditions result in conclusions that may not be transposable to other settings. After selection of the complexing agent, reagent addition order, buffer type and concentration, copper reducing assays were adapted to a high-throughput scheme and validated using model biological antioxidant compounds of ascorbic acid, Trolox (a soluble analogue of vitamin E), uric acid and glutathione. A critical comparison was made based on real samples including NIST-909c human serum certified sample, and five study samples. The validated method provided linear range up to 100 µM Trolox, (limit of detection 2.3 µM; limit of quantification 7.7 µM) with recovery results above 85% and precision <5%. The validated developed method with an increased sensitivity is a sound choice for assessment of TAC in serum samples. PMID:24968275
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Brominated flame retardant (BFRs) and Dechlorane Plus (DP) in paired human serum and segmented hair.
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.
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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.
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Centrifugal microfluidic platform for ultrasensitive detection of botulinum toxin
Koh, Chung -Yan; Schaff, Ulrich Y.; Sandstone Diagnostics, Livermore, CA; ...
2014-12-18
In this study, we present an innovative centrifugal microfluidic immunoassay platform (SpinDx) to address the urgent biodefense and public health need for ultrasensitive point-of-care/incident detection of botulinum toxin. The simple, sample-to-answer centrifugal microfluidic immunoassay approach is based on binding of toxins to antibody-laden capture particles followed by sedimentation of the particles through a density-media in a microfluidic disk and quantification by laser-induced fluorescence. A blind, head-to-head comparison study of SpinDx versus the gold-standard mouse bioassay demonstrates 100-fold improvement in sensitivity (limit of detection = 0.09 pg/mL), while achieving total sample-to-answer time of <30 min with 2-μL required volume of themore » unprocessed sample. We further demonstrate quantification of botulinum toxin in both exogeneous (human blood and serum spiked with toxins) and endogeneous (serum from mice intoxicated via oral, intranasal, and intravenous routes) samples. SpinDx can analyze, without any sample preparation, multiple sample types including whole blood, serum, and food. It is readily expandable to additional analytes as the assay reagents (i.e., the capture beads and detection antibodies) are disconnected from the disk architecture and the reader, facilitating rapid development of new assays. SpinDx can also serve as a general-purpose immunoassay platform applicable to diagnosis of other conditions and diseases.« less
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Thoresen, Stein I; Arnemo, Jon M; Liberg, Olof
2009-06-01
Scandinavian free-ranging wolves (Canis lupus) are endangered, such that laboratory data to assess their health status is increasingly important. Although wolves have been studied for decades, most biological information comes from captive animals. The objective of the present study was to establish reference intervals for 30 clinical chemical and 8 hematologic analytes in Scandinavian free-ranging wolves. All wolves were tracked and chemically immobilized from a helicopter before examination and blood sampling in the winter of 7 consecutive years (1998-2004). Seventy-nine blood samples were collected from 57 gray wolves, including 24 juveniles (24 samples), 17 adult females (25 samples), and 16 adult males (30 samples). Whole blood and serum samples were stored at refrigeration temperature for 1-3 days before hematologic analyses and for 1-5 days before serum biochemical analyses. Reference intervals were calculated as 95% confidence intervals except for juveniles where the minimum and maximum values were used. Significant differences were observed between adult and juvenile wolves for RBC parameters, alkaline phosphatase and amylase activities, and total protein, albumin, gamma-globulins, cholesterol, creatinine, calcium, chloride, magnesium, phosphate, and sodium concentrations. Compared with published reference values for captive wolves, reference intervals for free-ranging wolves reflected exercise activity associated with capture (higher creatine kinase activity, higher glucose concentration), and differences in nutritional status (higher urea concentration).
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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.
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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.
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Serum and urine metabolomic fingerprinting in diagnostics of inflammatory bowel diseases.
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.
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Serum and urine metabolomic fingerprinting in diagnostics of inflammatory bowel diseases
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
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51Cr-EDTA absorption blood test: an easy method for assessing small intestinal permeability in dogs.
Frias, Rafael; Sankari, Satu; Westermarck, Elias
2004-01-01
The 51Cr-EDTA test is a valuable clinical tool for screening intestinal diseases in dogs. The test is performed by calculating the percentage of recovery from urine of a PO-ingested dose of 51Cr-EDTA after 6 or 24 hours. Careful urine collection is a practical limitation of this test in dogs, and our goal was to develop a simpler test that measures 51Cr-EDTA in blood. A 51Cr-EDTA absorption test was simultaneously performed on urine and serum 43 times in healthy Beagle Dogs. Timed blood samples were withdrawn, and urine was collected during a 6-hour period. Percentages of the ingested dose were then calculated in urine and serum. The mean +/- standard deviation (range) percentage in urine after 6 hours was 14.07 +/- 8.72% (3.81-34.18%), whereas results in serum from samples taken at 2, 3, 4, 5, and 6 hours were 0.49 +/- 0.45% (0.02-2.13%), 0.75 +/- 0.52% (0.03-1.89%), 0.82 +/- 0.57% (0.13-2.21%), 0.70 +/- 0.53% (0.12-1.99%), and 0.47 +/- 0.44% (0.11-1.79%), respectively. The results for blood specimens showed good concordance with those for urine, especially for the samples taken at 4 hours (r = 0.89). Moreover, the correlation between urine and blood was better when the sum of the percentages of the recovered analyte from various blood samples was compared with urine. The correlation coefficient when summing 4 blood samples was excellent (r = 0.97) and remained excellent when summing only 2 blood samples taken at 3 and 5 hours (r = 0.95) or at 3 and 4 hours (r = 0.94). We conclude that a serum 51Cr-EDTA test determined by summing successive blood samples provides an easier means of estimating small intestinal permeability in dogs and gives results comparable to those of the 6-hour urine test.
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Liu, Ruyu; Rao, Huiying; Wang, Jianghua; Xie, Xingwang; Jiang, Dong; Pan, Xiaoben; Zhao, Ping; Zhang, Henghui; Wei, Lai
2013-01-01
It has been reported that monoclonal antibodies (MAbs) to the E1E2 glycoproteins may have the potential to prevent hepatitis C virus (HCV) infection. The protective epitopes targeted by these MAbs have been mapped to the regionsencompassing amino acids 313–327 and 432–443. In this study, we synthesized these two peptides and tested the reactivity of serum samples from 336 patients, 210 of whichwere from Chronic Hepatitis C (CHC) patients infected with diverse HCV genotypes.The remaining 126 samples were isolated from patients who had spontaneously clearedHCV infection.In the chronic HCV-infected group (CHC group), the prevalence of human serum antibodies reactive to epitopes 313–327 and 432–443was 24.29%(51 of 210) and4.76%(10 of 210),respectively. In thespontaneousclearance group (SC group),the prevalence was 0.79%(1 of 126) and 12.70%(16 of 126), respectively.The positive serum samples that contained antibodies reactive to epitope 313–327 neutralizedHCV pseudoparticles (HCVpp) bearing the envelope glycoproteins of genotypes 1a or 1b and/or 4, but genotypes 2a, 3a, 5 and 6 were not neutralized. The neutralizing activity of these serum samples could not be inhibited by peptide 313–327. Six samples (SC17, SC38, SC86, SC92, CHC75 and CHC198) containing antibodies reactive to epitope 432–443 had cross-genotype neutralizing activities. Theneutralizing activityof SC38, SC86, SC92 and CHC75waspartiallyinhibited by peptide 432–443. However,the neutralizing activity of sample SC17 for genotype 4HCVpp and sample CHC198 for genotype 1b HCVppwere notinhibited by the peptide.This study identifies the neutralizing ability of endogenous anti-HCV antibodies and warrants the exploration of antibodies reactive to epitope432–443as sources for future antibody therapies. PMID:23826163
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Höllig, Anke; Stoffel-Wagner, Birgit; Clusmann, Hans; Veldeman, Michael; Schubert, Gerrit A; Coburn, Mark
2017-01-01
Aneurysmal subarachnoid hemorrhage triggers an intense inflammatory response, which is suspected to increase the risk for secondary complications such as delayed cerebral ischemia (DCI). However, to date, the monitoring of the inflammatory response to detect secondary complications such as DCI has not become part of the clinical routine diagnostic. Here, we aim to illustrate the time courses of inflammatory parameters after aneurysmal subarachnoid hemorrhage (aSAH) and discuss the problems of inflammatory parameters as biomarkers but also their possible relevance for deeper understanding of the pathophysiology after aSAH and sophisticated planning of future studies. In this prospective cohort study, 109 patients with aSAH were initially included, n = 28 patients had to be excluded. Serum and-if possible-cerebral spinal fluid samples ( n = 48) were retrieved at days 1, 4, 7, 10, and 14 after aSAH. Samples were analyzed for leukocyte count and C-reactive protein (CRP) (serum samples only) as well as matrix metallopeptidase 9 (MMP9), intercellular adhesion molecule 1 (ICAM1), and leukemia inhibitory factor (LIF) [both serum and cerebrospinal fluid (CSF) samples]. Time courses of the inflammatory parameters were displayed and related to the occurrence of DCI. We illustrate the time courses of leukocyte count, CRP, MMP9, ICAM1, and LIF in patients' serum samples from the first until the 14th day after aSAH. Time courses of MMP9, ICAM1, and LIF in CSF samples are demonstrated. Furthermore, no significant difference was shown relating the time courses to the occurrence of DCI. We estimate that the wide range of the measured values hampers their interpretation and usage as a biomarker. However, understanding the inflammatory response after aSAH and generating a multicenter database may facilitate further studies: realistic sample size calculations on the basis of a multicenter database will increase the quality and clinical relevance of the acquired results.
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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.
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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.
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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.
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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.
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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.
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Meyer, Golo M; Maurer, Hans H; Meyer, Markus R
2016-01-01
This paper reviews MS approaches applied to metabolism studies, structure elucidation and qualitative or quantitative screening of drugs (of abuse) and/or their metabolites. Applications in clinical and forensic toxicology were included using blood plasma or serum, urine, in vitro samples, liquids, solids or plant material. Techniques covered are liquid chromatography coupled to low-resolution and high-resolution multiple stage mass analyzers. Only PubMed listed studies published in English between January 2008 and January 2015 were considered. Approaches are discussed focusing on sample preparation and mass spectral settings. Comments on advantages and limitations of these techniques complete the review.
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Early Whole Blood for Patients Requiring Massive Transfusion after Major Trauma. Addendum
2013-10-01
process of determining which of the 45 patients had an abdominal CT. Measure serum (adipokines, leptin, adiponectin, and ghrelin ). We obtained serial...samples thru 24 hours on 60 patients and samples thru five days on 23 patients. We measured serum adiponectin, leptin, ghrelin , and resistin, and...adiponectin, leptin, and ghrelin were all lower in injured patients than in healthy volunteers at all timepoints. Adiponectin slowly decreased over time
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Dittmer, K E; Hinkson, J A; Dwyer, C; Adlington, B; van Andel, M
2018-01-01
To determine the prevalence of infection with Candidatus Mycoplasma haemolamae (Mhl), antibodies to bovine viral diarrhoea virus (BVDV), and BVDV antigen, and the prevalence of animals with elevated faecal nematode egg counts (FEC) in a sample of adult New Zealand alpaca (Vicugna pacos). Blood samples were obtained from 175 alpaca, collected from 15 farms around New Zealand, and from 31 samples sent to a diagnostic laboratory for routine haematology. Blood smears (n=170) were examined microscopically for the presence of haemoplasma, and DNA was extracted from whole blood (n=206) for real-time PCR testing for Mhl. Packed cell volume (PCV) was determined for 193 samples. Serum samples (n=195) were tested for BVDV antibody using ELISA, and for BVDV antigen using a real-time PCR assay. Faecal samples were collected from 143 animals; FEC were measured, and samples pooled for larval culture. No haemoplasma organisms were present on blood smear examination. Of the 206 blood samples, two (from the same farm) were positive for Mhl by real-time PCR testing, giving a prevalence of infection with Mhl of 0.97%. Of the 195 serum samples tested, four (2.1%) were positive for antibodies to BVDV; animals with BVDV antibodies were from 3/15 (20%) farms, none of which farmed cattle. None of the serum samples were positive by PCR for BVDV antigen. The median FEC was 50 epg (min 0, max 4,700), with 55/143 (38.5%) samples having 0 epg, and 33/143 (23.1%) having ≥250 epg. Haemonchus spp. were the most common nematodes present in faecal larval cultures from the North Island. Log 10 FEC was negatively associated with PCV (p=0.02), and was higher in males than females (p<0.001), and in animals that were positive compared with negative for Mhl (p=0.022). The number of alpaca infected with Mhl was low, as was the seroprevalence of BVDV. Gastrointestinal parasitism was, however, a common finding in this sample of New Zealand alpaca.
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Herrera, Michael; Ding, Haiqing; McClanahan, Robert; Owens, Jane G; Hunter, Robert P
2007-09-15
A highly sensitive and quantitative LC/MS/MS assay for the determination of tilmicosin in serum has been developed and validated. For sample preparation, 0.2 mL of canine serum was extracted with 3 mL of methyl tert-butyl ether. The organic layer was transferred to a new vessel and dried under nitrogen. The sample was then reconstituted for analysis by high performance liquid chromatography-tandem mass spectrometry. A Phenomenex Luna C8(2) analytical column was used for the chromatographic separation. The eluent was subsequently introduced to the mass spectrometer by electrospray ionization. A single range was validated for 50-5000 ng/mL for support of toxicokinetic studies. The inter-day relative error (inaccuracy) for the LLOQ samples ranged from -5.5% to 0.3%. The inter-day relative standard deviations (imprecision) at the respective LLOQ levels were < or =10.1%.
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NASA Astrophysics Data System (ADS)
Youssef, A. O.
2018-04-01
A novel, simple, sensitive and selective spectrofluorimetric method was developed for the determination of Acetazolamide in pharmaceutical tablets and serum samples using photo probe Tb3+-ACAC. The Acetazolamide can remarkably quench the luminescence intensity of Tb3+-ACAC complex in DMSO at pH 6.8 and λex = 350 nm. The quenching of luminescence intensity of Tb3+-ACAC complex especially the electrical band at λem = 545 nm is used for the assessment of Acetazolamide in the pharmaceutical tablet and serum samples. The dynamic range found for the determination of Acetazolamide concentration is 4.49 × 10-9-1.28 × 10-7 mol L-1, and the limit of detection (LOD) and limit of quantification (LOQ) are (4.0 × 10-9 and 1.21 × 10-8) mol L-1, respectively.
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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.
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Schoolchildren with Learning Difficulties Have Low Iron Status and High Anemia Prevalence
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
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Schoolchildren with Learning Difficulties Have Low Iron Status and High Anemia Prevalence.
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.
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Fecal corticosterone reflects serum corticosterone in Florida sandhill cranes.
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.
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Rota, J S; Turner, J C; Yost-Daljev, M K; Freeman, M; Toney, D M; Meisel, E; Williams, N; Sowers, S B; Lowe, L; Rota, P A; Nicolai, L A; Peake, L; Bellini, W J
2009-10-01
Following the clinical diagnosis of the first case of mumps on September 22, 2006 at the University of Virginia (UVA), 52 suspected cases were identified through active surveillance for mumps by the end of December 2006. Samples were collected from 47 students who presented with parotitis despite a documented history of two doses of measles, mumps, and rubella (MMR) vaccine. Six of 47 serum samples (13%) were positive for mumps IgM, and 46/47 specimens were positive for mumps IgG. Endpoint titration of acute phase serum samples from laboratory-confirmed cases did not provide evidence that elevated serum IgG is a consistent marker for infection among cases due to secondary vaccine failure. Buccal swab samples from 39 of the 47 students were tested by real-time reverse transcription-polymerase chain reaction (RT-PCR) and/or viral culture. Mumps virus or mumps RNA was detected in 12 of 39 buccal samples (31%). Genetic analysis of the virus from the outbreak at UVA indicated that the outbreak was not linked to the large mumps outbreak in the Midwestern US that occurred earlier in 2006. Our findings support the use of viral detection to improve laboratory diagnosis of mumps among persons who have received two doses of MMR.
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Sharipova, I N; Khodak, N M; Puzirev, V F; Burkov, A N; Ulanova, T I
2015-03-01
The detection of false positive serological reactions (FPSR) on HIV-infection under screening examination of pregnant women is an actual problem of practical health care. The original observations testify that under analysis of the same samples of blood serum of pregnant women using screening immune enzyme test-systems of various manufacturers the unmatched data concerning FPSR can be obtained. The purpose of this study was to implement comparative evaluation of specificity of immune enzyme test-systems of three different manufacturers: "DS-IFA-HIV-AGAT-SCREEN" ("Diagnostic Systems"), "Genscreen Ultra HIV Ag-Ab" "Bio Rad" France) and "The CombiBest HIV-1,2 AG/AT" ("Vector-Best" Novosibirsk). The sampling of 440 samples of blood serums of pregnant women from various medical institutions of Nizhnii Novgorod was analyzed. The results of the study demonstrated that FPSR were detected in all test-systems and at that spectrum of samples differed. The identical specificity of compared test-systems amounted to 98.64%. The alternative approach to FPSR to HIV issue under screening examinations of pregnant women was proposed. The proposed mode consisted of consistent application of two test-systems of fourth generation with different format of setup of reaction.
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NHEXAS PHASE I MARYLAND STUDY--LIPIDS IN BLOOD ANALYTICAL RESULTS
The Lipids in Blood data set presents concentrations of cholesterol and total triglycerides in blood serum. The data set presents measurements for up to 2 lipids in 358 blood samples over 79 households. Each sample was collected via a venous sample from the primary respondent w...
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NHEXAS PHASE I MARYLAND STUDY--QA ANALYTICAL RESULTS FOR PESTICIDES IN BLANKS
The Pesticides in Blanks data set contains the analytical results of measurements of up to 20 pesticides in 70 blank samples from 46 households. Measurements were made in blank samples of indoor air, dust, soil, drinking water, food, beverages, and blood serum. Blank samples we...
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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.
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Achaintre, David; Sacerdote, Carlotta; Vineis, Paolo; Key, Timothy J.; Onland Moret, N. Charlotte; Scalbert, Augustin; Rinaldi, Sabina; Ferrari, Pietro
2015-01-01
Objective Although metabolic profiles have been associated with chronic disease risk, lack of temporal stability of metabolite levels could limit their use in epidemiological investigations. The present study aims to evaluate the reliability over a two-year period of 158 metabolites and compare reliability over time in fasting and non-fasting serum samples. Methods Metabolites were measured with the AbsolueIDQp180 kit (Biocrates, Innsbruck, Austria) by mass spectrometry and included acylcarnitines, amino acids, biogenic amines, hexoses, phosphatidylcholines and sphingomyelins. Measurements were performed on repeat serum samples collected two years apart in 27 fasting men from Turin, Italy, and 39 non-fasting women from Utrecht, The Netherlands, all participating in the European Prospective Investigation into Cancer and Nutrition (EPIC) study. Reproducibility was assessed by estimating intraclass correlation coefficients (ICCs) in multivariable mixed models. Results In fasting samples, a median ICC of 0.70 was observed. ICC values were <0.50 for 48% of amino acids, 27% of acylcarnitines, 18% of lysophosphatidylcholines and 4% of phosphatidylcholines. In non-fasting samples, the median ICC was 0.54. ICC values were <0.50 for 71% of acylcarnitines, 48% of amino acids, 44% of biogenic amines, 36% of sphingomyelins, 34% of phosphatidylcholines and 33% of lysophosphatidylcholines. Overall, reproducibility was lower in non-fasting as compared to fasting samples, with a statistically significant difference for 19–36% of acylcarnitines, phosphatidylcholines and sphingomyelins. Conclusion A single measurement per individual may be sufficient for the study of 73% and 52% of the metabolites showing ICCs >0.50 in fasting and non-fasting samples, respectively. ICCs were higher in fasting samples that are preferable to non-fasting. PMID:26274920
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Increased Serum Al Levels in Hemodialysis Patients Kept Enhanced during a 2-Year Prospective Study.
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.
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Stanek, J F; Stich, R W; Dubey, J P; Reed, S M; Njoku, C J; Lindsay, D S; Schmall, L M; Johnson, G K; LaFave, B M; Saville, W J A
2003-11-28
Equine protozoal myeloencephalitis (EPM) is a serious neurologic disease in the horse most commonly caused by Sarcocystis neurona. The domestic cat (Felis domesticus) is an intermediate host for S. neurona. In the present study, nine farms, known to have prior clinically diagnosed cases of EPM and a resident cat population were identified and sampled accordingly. In addition to the farm cats sampled, samples were also collected from a mobile spay and neuter clinic. Overall, serum samples were collected in 2001 from 310 cats, with samples including barn, feral and inside/outside cats. Of these 310 samples, 35 were from nine horse farms. Horse serum samples were also collected and traps were set for opossums at each of the farms. The S. neurona direct agglutination test (SAT) was used for both the horse and cat serum samples (1:25 dilution). Fourteen of 35 (40%) cats sampled from horse farms had circulating S. neurona agglutinating antibodies. Twenty-seven of the 275 (10%) cats from the spay/neuter clinic also had detectable S. neurona antibodies. Overall, 115 of 123 (93%) horses tested positive for anti-S. neurona antibodies, with each farm having greater than a 75% exposure rate among sampled horses. Twenty-one opossums were trapped on seven of the nine farms. Eleven opossums had Sarcocystis sp. sporocysts, six of them were identified as S. neurona sporocysts based on bioassays in gamma-interferon gene knockout mice with each opossum representing a different farm. Demonstration of S. neurona agglutinating antibodies in domestic and feral cats corroborates previous research demonstrating feral cats to be naturally infected, and also suggests that cats can be frequently infected with S. neurona and serve as one of several natural intermediate hosts for S. neurona.
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Focant, Jean-François; Eppe, Gauthier; Massart, Anne-Cécile; Scholl, Georges; Pirard, Catherine; De Pauw, Edwin
2006-10-13
We report on the use of a state-of-the-art method for the measurement of selected polychlorinated dibenzo-p-dioxins, polychlorinated dibenzofurans and polychlorinated biphenyls in human serum specimens. The sample preparation procedure is based on manual small size solid-phase extraction (SPE) followed by automated clean-up and fractionation using multi-sorbent liquid chromatography columns. SPE cartridges and all clean-up columns are disposable. Samples are processed in batches of 20 units, including one blank control (BC) sample and one quality control (QC) sample. The analytical measurement is performed using gas chromatography coupled to isotope dilution high-resolution mass spectrometry. The sample throughput corresponds to one series of 20 samples per day, from sample reception to data quality cross-check and reporting, once the procedure has been started and series of samples keep being produced. Four analysts are required to ensure proper performances of the procedure. The entire procedure has been validated under International Organization for Standardization (ISO) 17025 criteria and further tested over more than 1500 unknown samples during various epidemiological studies. The method is further discussed in terms of reproducibility, efficiency and long-term stability regarding the 35 target analytes. Data related to quality control and limit of quantification (LOQ) calculations are also presented and discussed.
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Comparison of Proteins in Whole Blood and Dried Blood Spot Samples by LC/MS/MS
NASA Astrophysics Data System (ADS)
Chambers, Andrew G.; Percy, Andrew J.; Hardie, Darryl B.; Borchers, Christoph H.
2013-09-01
Dried blood spot (DBS) sampling methods are desirable for population-wide biomarker screening programs because of their ease of collection, transportation, and storage. Immunoassays are traditionally used to quantify endogenous proteins in these samples but require a separate assay for each protein. Recently, targeted mass spectrometry (MS) has been proposed for generating highly-multiplexed assays for biomarker proteins in DBS samples. In this work, we report the first comparison of proteins in whole blood and DBS samples using an untargeted MS approach. The average number of proteins identified in undepleted whole blood and DBS samples by liquid chromatography (LC)/MS/MS was 223 and 253, respectively. Protein identification repeatability was between 77 %-92 % within replicates and the majority of these repeated proteins (70 %) were observed in both sample formats. Proteins exclusively identified in the liquid or dried fluid spot format were unbiased based on their molecular weight, isoelectric point, aliphatic index, and grand average hydrophobicity. In addition, we extended this comparison to include proteins in matching plasma and serum samples with their dried fluid spot equivalents, dried plasma spot (DPS), and dried serum spot (DSS). This work begins to define the accessibility of endogenous proteins in dried fluid spot samples for analysis by MS and is useful in evaluating the scope of this new approach.
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[Identification of hepatitis B virus YMDD point mutation using peptide nucleic acid clamping PCR].
Zhang, Yingying; He, Haitang; Yang, Jie; Hou, Jinlin
2013-06-01
To establish a peptide nucleic acid clamping PCR assay for detecting hepatitis B virus (HBV) drug resistance mutation. RtM204I (ATT) mutant, rtM204V (GTG) mutant and rtM204 (ATG) wild-type plasmids mixed at different ratios were detected for mutations by PNA clamping PCR assay and direct sequencing, and the sensitivity and specificity of the two methods were compared. Serum samples from 85 patients with chronic HBV infection were detected for drug resistance using the two methods. The sensitivity of PNA-PCR assay was 0.001% in a 10(5)-fold excess of wild-type HBV DNA with a detection limit of 10(1) copies. The sensitivity of direct sequencing was 10% with a detection limit of 10(4) copies. Mutants were detected in 73 of the 85 serum samples (85.9%), including YIDD in 40 samples, YVDD in 23 samples, and YIDD+YVDD in 10 samples. The agreement of PNA-PCR assay with direct sequencing was only 40% (34/85, YIDD in 21 samples, YVDD in 11 samples, and YIDD+YVDD in 2 samples). Neither of the two methods yielded positive results for the negative control samples, suggesting their good specificity. PNA-PCR assay appears to be a more sensitive and rapid assay for detection of HBV genotypic resistance.
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Exposure of Piglets to Enteroviruses Investigated by an Immunoassay Based on the EV-G1 VP4 Peptide.
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.
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Polyfluoroalkyl Chemicals in the Serum and Milk of ...
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
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Biernacka, Kinga; Karbowiak, Paweł; Wróbel, Paweł; Charęza, Tomasz; Czopowicz, Michał; Balka, Gyula; Goodell, Christa; Rauh, Rolf; Stadejek, Tomasz
2016-12-01
Recently oral fluid has become a novel sample type for pathogen nucleic acid and antibody detection, as it is easy to obtain with non-invasive procedures. The objective of the study was to analyze porcine reproductive and respiratory syndrome virus (PRRSV) and influenza A virus (IAV) circulation in growing pigs from three Polish production farms, using Real Time PCR and ELISA testing of oral fluid and serum. Oral fluids were collected every 2weeks, in the same 3-4 pens of pigs aged between 5 and 17weeks. Additionally, blood samples were collected every 4weeks from 4 pigs corresponding to the same pens as oral fluid and tested for the presence of PRRSV nucleic acid (pooled by 4) and antibodies. In farm A no PRRSV circulation was detected and only maternal antibodies were present. In farm B and farm C antibodies to PRRSV in serum and oral fluid were detected in most samples. In farm B PRRSV Type 1 was detected in 80.9% of oral fluid samples and in 58.3% of serum pools, and in farm C in 92.8% of oral fluid samples and 75% serum pools. Striking differences were observed between different pens in PRRSV detection patterns. In farms B and C ORF5 sequence analysis showed the presence of wild type strains which were about 84-85% identical to the modified live vaccine used. In all three farms two waves of IAV shedding with oral fluid were detected, in weaners and fatteners. Copyright © 2016 Elsevier Ltd. All rights reserved.
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Browne, J L; Schielen, P C J I; Belmouden, I; Pennings, J L A; Klipstein-Grobusch, K
2015-06-01
The objectives of the article is to compare pregnancy-associated plasma protein A (PAPP-A) and free β-subunit of human chorionic gonadotropin (β-hCG) concentrations in dried blood spots (DBSs) with serum of samples obtained from a public hospital in a low-resource setting and to evaluate their stability. Serum and DBS samples were obtained by venipuncture and finger prick from 50 pregnant participants in a cohort study in a public hospital in Accra, Ghana. PAPP-A and β-hCG concentrations from serum and DBS were measured with an AutoDELFIA® (PerkinElmer, PerkinElmer, Turku, Finland) automatic immunoassay. Correlation and Passing-Bablok regression analyses were performed to compare marker levels. High correlation (>0.9) was observed for PAPP-A and β-hCG levels between various sampling techniques. The β-hCG concentration was stable between DBS and serum, PAPP-A concentration consistently lower in DBS. Our findings suggest that β-hCG can be reliably collected from DBS in low-resource tropical settings. The exact conditions of the clinical workflow necessary for reliable PAPP-A measurement in these settings need to be further developed in the future. These findings could have implications for prenatal screening programs feasibility in low-income and middle-income countries, as DBS provides an alternative minimally invasive sampling method, with advantages in sampling technique, stability, logistics, and potential application in low-resource settings. © 2015 John Wiley & Sons, Ltd.
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Depciuch, J; Parlinska-Wojtan, M
2018-02-20
Depression is a serious mental illness. To study the mechanism of depression and search for new, more effective therapies, animal models are often used. Unfortunately, none of the available models reflects all the symptoms of depression. Therefore researchers are looking for new tools to diagnose depression. Unfortunately, the nowadays-available depression diagnosis methods are only psychological tests. However, it is known, that the amount of phospholipids, proteins and lipids decreases during depression. Raman and FTIR (Fourier Transform Infra Red) spectroscopies provide information on the chemical compounds in the measured sample e.g. blood serum. These spectroscopic techniques may thus become reliable and accurate tools for evaluating changes in the amount of phospholipids and proteins in depression disease. In this study differences between dried and liquid blood serum samples of healthy and depressed individuals measured by Raman (range 0-3000cm -1 ) and FTIR (Fourier Transform Infrared) (range 900-3000cm -1 ) spectroscopy were evaluated. The resulting spectra and accurate analysis led to the conclusion that an appropriate measurement of the background and the elimination of peaks from water had the greatest impact on the reliability of the results. Furthermore, after detailed studies of FTIR and Raman spectra of dried and liquid blood serum samples, including a complete analysis of peaks after Kramers-Kröning (KK) transformation, it was found that the sample preparation did not affect the results obtained by Raman spectroscopy. In FTIR measurements only a minimal effect on peak intensity was observed. Copyright © 2017 Elsevier B.V. All rights reserved.
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How the reference values for serum parathyroid hormone concentration are (or should be) established?
Souberbielle, J-C; Brazier, F; Piketty, M-L; Cormier, C; Minisola, S; Cavalier, E
2017-03-01
Well-validated reference values are necessary for a correct interpretation of a serum PTH concentration. Establishing PTH reference values needs recruiting a large reference population. Exclusion criteria for this population can be defined as any situation possibly inducing an increase or a decrease in PTH concentration. As recommended in the recent guidelines on the diagnosis and management of asymptomatic primary hyperparathyroidism, PTH reference values should be established in vitamin D-replete subjects with a normal renal function with possible stratification according to various factors such as age, gender, menopausal status, body mass index, and race. A consensus about analytical/pre-analytical aspects of PTH measurement is also needed with special emphasis on the nature of the sample (plasma or serum), the time and the fasting/non-fasting status of the blood sample. Our opinion is that blood sample for PTH measurement should be obtained in the morning after an overnight fast. Furthermore, despite longer stability of the PTH molecule in EDTA plasma, we prefer serum as it allows to measure calcium, a prerequisite for a correct interpretation of a PTH concentration, on the same sample. Once a consensus is reached, we believe an important international multicentre work should be performed to recruit a very extensive reference population of apparently healthy vitamin D-replete subjects with a normal renal function in order to establish the PTH normative data. Due to the huge inter-method variability in PTH measurement, a sufficient quantity of blood sample should be obtained to allow measurement with as many PTH kits as possible.
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Tarasidis, C G; Garnett, W R; Kline, B J; Pellock, J M
1986-01-01
The influence of storage conditions on the total and free concentration of valproic acid (VPA) was studied in six normal male subjects who ingested 750 mg of VPA (3 X 250 mg Depakene capsules; Abbott Laboratories). Blood samples were collected in various types of Vacutainer tubes (red top, no additives; green top, sodium heparin; blue top, sodium citrate; and purple top, EDTA) 2 h post administration of VPA. Either these samples were centrifuged immediately or stored for various periods of time at room temperature or refrigerated, or the supernate was frozen prior to analysis. Free VPA samples were obtained utilizing the Amicon ultrafiltration system. All VPA samples were analyzed by gas-liquid chromatography. Total VPA concentrations obtained from plasma collected with sodium citrate were lower (p less than 0.05) than either serum or plasma collected with other anticoagulants. There were no differences (p greater than 0.05) in total or free VPA concentrations between samples collected in serum or in plasma collected with heparin or EDTA. Storing samples for 96 h at room temperature did not alter the total VPA concentrations but was found to increase the free fraction of VPA (p less than 0.05). The refrigeration or freezing of the supernate from the blood samples for 7 days did not alter (p greater than 0.05) the total or the free fraction of VPA. The results of this study demonstrate that total and/or free VPA may be collected from either serum or plasma, provided sodium citrate is not used to collect plasma.(ABSTRACT TRUNCATED AT 250 WORDS)
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Sajid, Muhammad; Basheer, Chanbasha
2016-07-15
In present work, a new configuration of micro-solid phase extraction was introduced and termed as stir-bar supported micro-solid-phase extraction (SB-μ-SPE). A tiny stir-bar was packed inside the porous polypropylene membrane along with sorbent material and the edges of membrane sheet were heat sealed to secure the contents. The packing of stir-bar inside the μ-SPE device does not allow the device to stick with the wall or any corner of the sample vial during extraction, which is, however, a frequent observation in routine μ-SPE. Moreover, it enhances effective surface area of the sorbent exposed to sample solution through continuous agitation (motion and rotation). It also completely immerses the SB-μ-SPE device in the sample solution even for non-polar sorbents. Polychlorinated biphenyls (PCBs) were selected as model compounds and the method performance was evaluated in human serum samples. After extraction, samples were analyzed by gas chromatography mass spectrometry (GC-MS). The factors that affect extraction efficiency of SB-μ-SPE were optimized. Under optimum conditions, a good linearity (0.1-100ngmL(-1)) with coefficients of determinations ranging from 0.9868 to 0.9992 was obtained. Limits of detections were ranged between 0.003 and 0.047ngmL(-1). Acceptable values for inter-day (3.2-9.1%) and intra-day (3.1-7.2%) relative standard deviations were obtained. The optimized method was successfully applied to determine the concentration of PCB congeners in human serum samples. Copyright © 2016 Elsevier B.V. All rights reserved.
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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
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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.
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Serum prolactin in coeliac disease: a marker for disease activity
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
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Hart, Leslie B.; Wells, Randall S.; Kellar, Nick; Balmer, Brian C.; Hohn, Aleta A.; Lamb, Stephen V.; Rowles, Teri; Zolman, Eric S.; Schwacke, Lori H.
2015-01-01
Inshore common bottlenose dolphins (Tursiops truncatus) are exposed to a broad spectrum of natural and anthropogenic stressors. In response to these stressors, the mammalian adrenal gland releases hormones such as cortisol and aldosterone to maintain physiological and biochemical homeostasis. Consequently, adrenal gland dysfunction results in disruption of hormone secretion and an inappropriate stress response. Our objective herein was to develop diagnostic reference intervals (RIs) for adrenal hormones commonly associated with the stress response (i.e., cortisol, aldosterone) that account for the influence of intrinsic (e.g., age, sex) and extrinsic (e.g., time) factors. Ultimately, these reference intervals will be used to gauge an individual’s response to chase-capture stress and could indicate adrenal abnormalities. Linear mixed models (LMMs) were used to evaluate demographic and sampling factors contributing to differences in serum cortisol and aldosterone concentrations among bottlenose dolphins sampled in Sarasota Bay, Florida, USA (2000–2012). Serum cortisol concentrations were significantly associated with elapsed time from initial stimulation to sample collection (p<0.05), and RIs were constructed using nonparametric methods based on elapsed sampling time for dolphins sampled in less than 30 minutes following net deployment (95% RI: 0.91–4.21 µg/dL) and following biological sampling aboard a research vessel (95% RI: 2.32–6.68 µg/dL). To examine the applicability of the pre-sampling cortisol RI across multiple estuarine stocks, data from three additional southeast U.S. sites were compared, revealing that all of the dolphins sampled from the other sites (N = 34) had cortisol concentrations within the 95th percentile RI. Significant associations between serum concentrations of aldosterone and variables reported in previous studies (i.e., age, elapsed sampling time) were not observed in the current project (p<0.05). Also, approximately 16% of Sarasota Bay bottlenose dolphin aldosterone concentrations were below the assay’s detection limit (11 pg/mL), thus hindering the ability to derive 95th percentile RIs. Serum aldosterone concentrations from animals sampled at the three additional sites were compared to the detection limit, and the proportion of animals with low aldosterone concentrations was not significantly different than an expected prevalence of 16%. Although this study relied upon long-term, free-ranging bottlenose dolphin health data from a single site, the objective RIs can be used for future evaluation of adrenal function among individuals sampled during capture-release health assessments. PMID:25993341
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Molecular diagnosis of Toxoplasma gondii infection in Libya.
Gashout, Aisha; Amro, Ahmad; Erhuma, Mabruk; Al-Dwibe, Hamida; Elmaihub, Eanas; Babba, Hamouda; Nattah, Nabil; Abudher, Abdalhafid
2016-04-16
Toxoplasma gondii infections are prevalent in humans and animals throughout Libya. Current diagnosis is based on detection of Toxoplasma-specific IgM and IgG. In this study, we established and optimized a diagnostic PCR assay for molecular diagnosis of T. gondii in Libya. From January to December, 2010, 177 blood and serum samples were collected from suspected patients. This includes: 140 women who have had spontaneous abortions, 26 HIV-positive patients, nine patients with leukemia and lymphoma, and two infants with ocular infection. Samples were screened for anti-Toxoplasma IgG and IgM antibodies before DNA extraction. The surface antigen gene 2 (SAG2) was targeted in a semi-nested PCR to amplify a 999 bp and a 614 bp fragment in the first and the second run respectively. A total of 54/140 (38.5 %) women who have had spontaneous abortions, 23/26 (88 %) HIV patients, 6/9 (66.6 %) of the leukaemia and lymphoma patients, and one child with ocular infection were seropositive for anti-Toxoplasma IgG and/or IgM. Genomic DNA was extracted from 38 selected seropositive samples. The PCR was sensitive enough to detect DNA concentration of 12 ng/μL. PCR analysis was performed for 38 selected seropositive patients (16 women who have had spontaneous abortions, 15 positive HIV patients, six leukaemia patients and one child with ocular infection). Our designed primers were successfully amplified in 22/38 (57.9 %) samples; 5/12 (35.7 %) from serum and 17/26 (65.8 %) from whole blood samples. All PCR positive samples were IgG-positive except two samples which were IgM and IgG & IgM-positive serum samples respectively. The semi-nested PCR confirmed five more samples. These included two leukaemia and two HIV-positive whole blood samples and one serum sample from an aborted woman. The ability of PCR to diagnose active toxoplasmosis is needed in immunocompromised patients and congenital toxoplasmosis cases, especially when serological techniques fail. For the first time in Libya, we established and optimized semi-nested PCR of SAG2 gene. The developed PCR method was able to detect as little as 12 ng/μL of T. gondii DNA and was useful to diagnose the diseases in women who have had spontaneous abortions, HIV-positive patients, patients with leukemia and lymphoma, and infants with ocular infection.
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Twenty-four cases of imported zika virus infections diagnosed by molecular methods.
Alejo-Cancho, Izaskun; Torner, Nuria; Oliveira, Inés; Martínez, Ana; Muñoz, José; Jane, Mireia; Gascón, Joaquim; Requena-Méndez, Ana; Vilella, Anna; Marcos, M Ángeles; Pinazo, María Jesús; Gonzalo, Verónica; Rodriguez, Natalia; Martínez, Miguel J
2016-10-01
Zika virus is an emerging flavivirus widely spreading through Latin America. Molecular diagnosis of the infection can be performed using serum, urine and saliva samples, although a well-defined diagnostic algorithm is not yet established. We describe a series of 24 cases of imported zika virus infection into Catalonia (northeastern Spain). Based on our findings, testing of paired serum and urine samples is recommended. Copyright © 2016 Elsevier Inc. All rights reserved.
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Serum antileptospiral agglutinins in freshwater turtles from Southern Brazil
Silva, Éverton F; Seyffert, Núbia; Cerqueira, Gustavo M.; Leihs, Karl P.; Athanazio, Daniel A.; Valente, Ana L. S.; Dellagostin, Odir A.; Brod, Claudiomar S.
2009-01-01
In this study, we observed the presence of antileptospiral agglutinins in freshwater turtles of two urban lakes of Pelotas, Southern Brazil. Forty animals (29 Trachemys dorbigny and 11 Phrynops hilarii) were captured and studied. Attempts to isolate leptospires from blood and urine samples were unsuccessful. Serum samples (titer > 100) reactive to pathogenic strains were observed in 11 animals. These data encourage surveys of pet turtles to evaluate the risk of transmission of pathogenic leptospires to humans. PMID:24031348
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NASA Astrophysics Data System (ADS)
Thuy, Tran Thi; Thorsén, Gunnar
2013-07-01
The serum clearance rate of therapeutic antibodies is important as it affects the clinical efficacy, required dose, and dose frequency. The glycosylation of antibodies has in some studies been shown to have an impact on the elimination rates in vivo. Monitoring changes to the glycan profiles in pharmacokinetics studies can reveal whether the clearance rates of the therapeutic antibodies depend on the different glycoforms, thereby providing useful information for improvement of the drugs. In this paper, a novel method for glycosylation analysis of therapeutic antibodies in serum samples is presented. A microfluidic compact-disc (CD) platform in combination with MALDI-MS was used to monitor changes to the glycosylation profiles of samples incubated in vitro. Antibodies were selectively purified from serum using immunoaffinity capture on immobilized target antigens. The glycans were enzymatically released, purified, and finally analyzed by MALDI-TOF-MS. To simulate changes to glycan profiles after administration in vivo, a therapeutic antibody was incubated in serum with the enzyme α1-2,3 mannosidase to artificially reduce the amount of the high mannose glycoforms. Glycan profiles were monitored at specific intervals during the incubation. The relative abundance of the high mannose 5 glycoform was clearly found to decrease and, simultaneously, that of high mannose 4 increased over the incubation period. The method can be performed in a rapid, parallel, and automated fashion for glycosylation profiling consuming low amounts of samples and reagents. This can contribute to less labor work and reduced cost of the studies of therapeutic antibodies glycosylation in vitro and in vivo.
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Srinivasan, Balaji; O'Dell, Dakota; Finkelstein, Julia L; Lee, Seoho; Erickson, David; Mehta, Saurabh
2018-01-15
Iron deficiency (ID) is an urgent public health problem that has devastating effects on maternal and child health. However, due to poor access and affordability, screening and diagnosis for ID is often limited to proxy hemoglobin measurements alone. Here, we report the development and validation of ironPhone, a mobile-device coupled portable diagnostics for quantification of serum ferritin concentrations, an iron status biomarker, within a few minutes, from a drop of fingerprick blood. The ironPhone diagnostic platform comprises of a smartphone accessory, an app, and a disposable lateral flow immunoassay test strip to quantify serum ferritin. For initial validation in the lab, we optimized and evaluated the performance of ironPhone with known ferritin concentrations in spiked buffer and serum samples. Following lab validation, we performed a human validation by collecting fingerprick whole blood samples from 20 participants to assess iron status using ironPhone and compared the results with the laboratory standard IMMULITE 2000 analyzer. Findings from the ironPhone for the buffer and spiked serum samples provided a calibration curve with R 2 values of 0.97 (n=27) and 0.93 (n=12), respectively. On comparison with the laboratory standard IMMULITE analyzer in whole blood samples, a correlation of 0.92 (P<0.0001) was observed with a sensitivity of over 90% for predicting ID (ferritin<15.0µg/L) via the ironPhone, demonstrating its promise for iron status assessment at the point-of-care. Copyright © 2017 Elsevier B.V. All rights reserved.
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Chen, Rui; Tan, Yexiong; Wang, Min; Wang, Fangjun; Yao, Zhenzhen; Dong, Liwei; Ye, Mingliang; Wang, Hongyang; Zou, Hanfa
2011-01-01
A robust, reproducible, and high throughput method was developed for the relative quantitative analysis of glycoprotein abundances in human serum. Instead of quantifying glycoproteins by glycopeptides in conventional quantitative glycoproteomics, glycoproteins were quantified by nonglycosylated peptides derived from the glycoprotein digest, which consists of the capture of glycoproteins in serum samples and the release of nonglycopeptides by trypsin digestion of captured glycoproteins followed by two-dimensional liquid chromatography-tandem MS analysis of released peptides. Protein quantification was achieved by comparing the spectrum counts of identified nonglycosylated peptides of glycoproteins between different samples. This method was demonstrated to have almost the same specificity and sensitivity in glycoproteins quantification as capture at glycopeptides level. The differential abundance of proteins present at as low as nanogram per milliliter levels was quantified with high confidence. The established method was applied to the analysis of human serum samples from healthy people and patients with hepatocellular carcinoma (HCC) to screen differential glycoproteins in HCC. Thirty eight glycoproteins were found with substantial concentration changes between normal and HCC serum samples, including α-fetoprotein, the only clinically used marker for HCC diagnosis. The abundance changes of three glycoproteins, i.e. galectin-3 binding protein, insulin-like growth factor binding protein 3, and thrombospondin 1, which were associated with the development of HCC, were further confirmed by enzyme-linked immunosorbent assay. In conclusion, the developed method was an effective approach to quantitatively analyze glycoproteins in human serum and could be further applied in the biomarker discovery for HCC and other cancers. PMID:21474793
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Reference measurement procedure for total glycerides by isotope dilution GC-MS.
Edwards, Selvin H; Stribling, Shelton L; Pyatt, Susan D; Kimberly, Mary M
2012-04-01
The CDC's Lipid Standardization Program established the chromotropic acid (CA) reference measurement procedure (RMP) as the accuracy base for standardization and metrological traceability for triglyceride testing. The CA RMP has several disadvantages, including lack of ruggedness. It uses obsolete instrumentation and hazardous reagents. To overcome these problems the CDC developed an isotope dilution GC-MS (ID-GC-MS) RMP for total glycerides in serum. We diluted serum samples with Tris-HCl buffer solution and spiked 200-μL aliquots with [(13)C(3)]-glycerol. These samples were incubated and hydrolyzed under basic conditions. The samples were dried, derivatized with acetic anhydride and pyridine, extracted with ethyl acetate, and analyzed by ID-GC-MS. Linearity, imprecision, and accuracy were evaluated by analyzing calibrator solutions, 10 serum pools, and a standard reference material (SRM 1951b). The calibration response was linear for the range of calibrator concentrations examined (0-1.24 mmol/L) with a slope and intercept of 0.717 (95% CI, 0.7123-0.7225) and 0.3122 (95% CI, 0.3096-0.3140), respectively. The limit of detection was 14.8 μmol/L. The mean %CV for the sample set (serum pools and SRM) was 1.2%. The mean %bias from NIST isotope dilution MS values for SRM 1951b was 0.7%. This ID-GC-MS RMP has the specificity and ruggedness to accurately quantify total glycerides in the serum pools used in the CDC's Lipid Standardization Program and demonstrates sufficiently acceptable agreement with the NIST primary RMP for total glyceride measurement.
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Ren, Yao; Walczyk, Thomas
2014-09-01
Ferritin is a hollow sphere protein composed of 24 subunits that can store up to 4500 iron atoms in its inner cavity. It is mainly found in the liver and spleen but also in serum at trace levels. Serum ferritin is considered as the best single indicator in assessing body iron stores except liver or bone marrow biopsy. However, it is confounded by other disease conditions. Ferritin bound iron (FBI) and ferritin saturation have been suggested as more robust biomarkers. The current techniques for FBI determination are limited by low antibody specificity, low instrument sensitivity and possible analyte losses during sample preparation. The need for a highly sensitive and reliable method is widely recognized. Here we describe a novel technique to detect serum FBI using species-specific isotope dilution mass spectrometry (SS-IDMS). [(57)Fe]-ferritin was produced by biosynthesis and in vitro labeling with the (57)Fe spike in the form of [(57)Fe]-citrate after cell lysis and heat treatment. [(57)Fe]-ferritin for sample spiking was further purified by fast liquid protein chromatography. Serum ferritin and added [(57)Fe]-ferritin were separated from other iron species by ultrafiltration followed by isotopic analysis of FBI using negative thermal ionization mass spectrometry. Repeatability of our assay is 8% with an absolute detection limit of 18 ng FBI in the sample. As compared to other speciation techniques, SS-IDMS offers maximum control over sample losses and species conversion during analysis. The described technique may therefore serve as a reference technique for clinical applications of FBI as a new biomarker for assessing body iron status.
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Acute phase proteins in the diagnosis of bovine subclinical mastitis.
Safi, Shahabeddin; Khoshvaghti, Ameneh; Jafarzadeh, Seyed Reza; Bolourchi, Mahmoud; Nowrouzian, Iradj
2009-12-01
The California mastitis test (CMT) and somatic cell count (SCC) are commonly used for diagnosis of subclinical mastitis in cattle. Acute phase proteins (APPs), as alternative biomarkers of mastitis, may increase in concentration in the absence of macroscopic changes in the milk, or may precede the onset of clinical signs. The aim of this study was to compare the accuracy of APPs measured in milk and in serum with bacterial culture for the diagnosis of bovine subclinical mastitis. One hundred and seventy-five Holstein cows were randomly selected from 7 dairy farms. Quarter milk and serum samples were taken from all cows. Milk samples were analyzed using a CMT and SCC, and for haptoglobin (MHp) and amyloid A (MAA) concentrations, and were also submitted for bacterial culture. Serum samples obtained concurrently were analyzed for haptoglobin (SHp) and amyloid A (SAA). Two-sample Wilcoxon (Mann-Whitney) test was used to compare SCC, MAA, MHp, SAA, and SHp concentrations between culture-positive and culture-negative animals. Receiver-operating characteristic analysis was used to assess the performance of each test using bacterial culture as the reference method. MAA concentration was the most accurate of the 5 tests, with a sensitivity of 90.6% and specificity of 98.3% at concentrations >16.4 mg/L. MAA and MHp had significantly larger areas under the curve than the respective serum proteins, SAA and SHp. The results suggest that measuring haptoglobin and amyloid A in milk is more accurate than serum analysis for the diagnosis of subclinical mastitis in Holstein cows.
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Li, Jian; Chen, Tian; Wang, Yuwei; Shi, Zhixiong; Zhou, Xianqing; Sun, Zhiwei; Wang, Dejun; Wu, Yongning
2017-02-01
Two simplified sample preparation procedures for simultaneous extraction and clean-up of tetrabromobisphenol A, α-, β-, and γ-hexabromocyclododecane and polybrominated diphenyl ethers in human serum were developed and validated. The first procedure was based on solid-phase extraction. Sample extraction, purification, and lipid removal were carried out directly on an Oasis HLB cartridge. The second procedure was a quick, easy, cheap, effective, rugged, and safe-based approach using octadecyl-modified silica particles as a sorbent. After sample extraction and cleanup, tetrabromobisphenol A/hexabromocyclododecane was separated from polybrominated diphenyl ethers by using a Si-based cartridge. Tetrabromobisphenol A and hexabromocyclododecane were then detected by high-performance liquid chromatography coupled to tandem mass spectrometry, while polybrominated diphenyl ethers were detected by gas chromatography coupled to tandem mass spectrometry. The results of the spike recovery test using fetal bovine serum showed that the average recoveries of the analytes ranged from 87.3 to 115.3% with relative standard deviations equal to or lower than 13.4 %. Limits of detection of the analytes were in the range of 0.4-19 pg/mL except for decabromodiphenyl ether. The developed method was successfully applied to routine analysis of human serum samples from occupational workers and the general population. Extremely high serum polybrominated diphenyl ethers levels up to 3.32 × 10 4 ng/g lipid weight were found in occupational workers. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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Bagchus, Wilma M; Smeets, Jean M W; Verheul, Herman A M; De Jager-Van Der Veen, Suzanne M; Port, Andreas; Geurts, T B Paul
2005-05-01
The pharmacokinetics of nandrolone in serum and urine were investigated in healthy young men after a single im injection of 50 mg (n = 20), 100 mg (n = 17), or 150 mg (n = 17) nandrolone decanoate. Blood samples were collected before treatment and for up to 32 d after dosing. In addition, in the 50- and 150-mg groups, 24-h urine samples were collected before treatment and on d 1, 7, and 33 after treatment; in the 150-mg group, additional samples were collected after 3 and 6 months. Serum concentrations and the area under the curve of nandrolone increased proportionally with the dose administered. The peak serum concentration ranged from 2.14 ng/ml in the 50-mg group to 4.26 ng/ml in the 100-mg group and 5.16 ng/ml in the 150-mg group. The peak serum concentration was reached after 30 h (50 and 100 mg) and 72 h (150 mg), whereas the terminal half-life was 7-12 d. In urine, pretreatment concentrations of 19-norandrosterone (19-NA) and/or 19-noretiocholanolone (19-NE) were detected in five of 37 subjects (14%). In the 50-mg group, 19-NA and/or 19-NE could be detected at least until 33 d after injection in 16 of 17 subjects (94%). In the 150-mg group, who were presumed to have not previously used nandrolone, nandrolone metabolites could be detected for up to 6 months in eight of 12 subjects (67%) for 19-NE and in 10 of 12 subjects (83%) for 19-NA.
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Abdel Rahman, Mohamed F.; Hashad, Ingy M.; Abdel-Maksoud, Sahar M.; Farag, Nabil M.; Abou-Aisha, Khaled
2012-01-01
Aim: The aim of this study was to detect endothelial nitric oxide synthase (eNOS) Glu298Asp gene variants in a random sample of the Egyptian population, compare it with those from other populations, and attempt to correlate these variants with serum levels of nitric oxide (NO). The association of eNOS genotypes or serum NO levels with the incidence of acute myocardial infarction (AMI) was also examined. Methods: One hundred one unrelated healthy subjects and 104 unrelated AMI patients were recruited randomly from the 57357 Hospital and intensive care units of El Demerdash Hospital and National Heart Institute, Cairo, Egypt. eNOS genotypes were determined by polymerase chain reaction–restriction fragment length polymorphism. Serum NO was determined spectrophotometrically. Results: The genotype distribution of eNOS Glu298Asp polymorphism determined for our sample was 58.42% GG (wild type), 33.66% GT, and 7.92% TT genotypes while allele frequencies were 75.25% and 24.75% for G and T alleles, respectively. No significant association between serum NO and specific eNOS genotype could be detected. No significant correlation between eNOS genotype distribution or allele frequencies and the incidence of AMI was observed. Conclusion: The present study demonstrated the predominance of the homozygous genotype GG over the heterozygous GT and homozygous TT in random samples of Egyptian population. It also showed the lack of association between eNOS genotypes and mean serum levels of NO, as well as the incidence of AMI. PMID:22731641
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Colón, I; Caro, D; Bourdony, C J; Rosario, O
2000-01-01
Premature breast development (thelarche) is the growth of mammary tissue in girls younger than 8 years of age without other manifestations of puberty. Puerto Rico has the highest known incidence of premature thelarche ever reported. In the last two decades since this serious public health anomaly has been observed, no explanation for this phenomenon has been found. Some organic pollutants, including pesticides and some plasticizers, can disrupt normal sexual development in wildlife, and many of these have been widely used in Puerto Rico. This investigation was designed to identify pollutants in the serum of Puerto Rican girls with premature thelarche. A method for blood serum analysis was optimized and validated using pesticides and phthalate esters as model compounds of endocrine-disrupting chemicals. Recovery was > 80% for all compounds. We performed final detection by gas chromatography/mass spectrometry. We analyzed 41 serum samples from thelarche patients and 35 control samples. No pesticides or their metabolite residues were detected in the serum of the study or control subjects. Significantly high levels of phthalates [dimethyl, diethyl, dibutyl, and di-(2-ethylhexyl)] and its major metabolite mono-(2-ethylhexyl) phthalate were identified in 28 (68%) samples from thelarche patients. Of the control samples analyzed, only one showed significant levels of di-isooctyl phthalate. The phthalates that we identified have been classified as endocrine disruptors. This study suggests a possible association between plasticizers with known estrogenic and antiandrogenic activity and the cause of premature breast development in a human female population. PMID:11017896
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Ballinas-Verdugo, Martha; Reyes, Pedro Antonio; Mejia-Dominguez, Ana; López, Ruth; Matta, Vivian; Monteón, Victor M
2011-12-01
Thirteen Trypanosoma cruzi isolates from different geographic regions of Mexico and Guatemala belonging to discrete typing unit (DTU) I and a reference CL-Brener (DTU VI) strain were used to perform enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR). A panel of 57 Mexican serum samples of patients with chronic chagasic cardiopathy and asymptomatic infected subjects (blood bank donors) were used in this study. DNA from the above 14 T. cruzi strains were extracted and analyzed by PCR using different sets of primers designed from minicircle and satellite T. cruzi DNA. The chronic chagasic cardiopathy serum samples were easily recognized with ELISA regardless of the source of antigenic extract used, even with the CL-Brener TcVI, but positive serum samples from blood bank donors in some cases were not recognized by some Mexican antigenic extracts. On the other hand, PCR showed an excellent performance despite the set of primers used, since all Mexican and Guatemalan T. cruzi strains were correctly amplified. In general terms, Mexican, Guatemalan, and CL-Brener T. cruzi strains are equally good sources of antigen when using the ELISA test to detect Mexican serum samples. However, there are some strains with poor performance. The DTU I strains are easily detected using either kinetoplast or satellite DNA target designed from DTU VI strains.
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Cavalleri, A; Colombo, C; Venturelli, E; Miceli, R; Mariani, L; Cornelli, U; Pala, V; Berrino, F; Secreto, G
2004-01-01
Measuring the free radical activity in serum samples from prospective studies is the best way to investigate the association between oxidative stress and human diseases. Prospective studies require the analysis of serum samples that have often been stored for a long time. Our study was designed to determine the effect of storage at -30 degrees C and -80 degrees C for two years on free radical activity. We analyzed the free radical activity by measuring circulating hydroperoxides in a pool of sera at baseline and after one day, one week, one month and 25 months of storage, using a photometric method (d-ROMs test). Measurements were performed in aliquots thawed only once at each time point and in aliquots frozen and thawed repeatedly over the study period. After two years we observed a small but statistically significant 4% decrease in the hydroperoxide concentration, which was substantially unaffected by storage temperatures and repeated freeze-thaw cycles. We also carried out the d-ROMs test in sera from ten apparently healthy volunteers at 2, 8, 24, and 48 hours after collection and storage at 4 degrees C and did not observe any significant variation. In conclusion, the d-ROMs test is a simple method suitable to evaluate the free radical activity in frozen serum samples after long-term storage.
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Sharma, Neekun; Hotta, Akitoyo; Yamamoto, Yoshie; Fujita, Osamu; Uda, Akihiko; Morikawa, Shigeru; Yamada, Akio
2013-01-01
A novel competitive enzyme-linked immunosorbent assay (cELISA) was developed and evaluated for detection of antibodies against Francisella tularensis in humans. The assay is based on the ability of serum antibodies to inhibit the binding of monoclonal antibodies (MAbs) directed against F. tularensis lipopolysaccharide antigens. The assay was evaluated using serum samples of tularemia patients, inactivated F. tularensis-immunized rabbits, and F. tularensis-infected mice. Antibodies against F. tularensis were successfully detected in serum samples of tularemia patients as well as the immunized and infected animals. The cELISA method was compared to indirect ELISA (iELISA) and the commonly used microagglutination test (MA) using serum samples of 19 tularemia patients and 50 healthy individuals. The sensitivity and specificity of cELISA were 93.9 and 96.1%, respectively, in comparison to the iELISA. MA was less sensitive than cELISA with a sensitivity and specificity of only 81.8 and 98.0%, respectively. A high degree of correlation (R2 = 0.8226) was observed between cELISA and iELISA results. The novel cELISA developed in this study appears to be highly sensitive and specific for serodiagnosis of human tularemia. The potential of the MAb-based cELISA to be used in both human and animal samples emphasizes its usefulness for serological survey of tularemia among multiple animal species. PMID:23114700
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El Hage, J; Lorusso, A; Carmine, I; Di Gennaro, A; Portanti, O; Olivieri, S; Casaccia, C; Pisciella, M; Teodori, L; Sghaier, S; Savini, G
2013-10-01
Since 2000, several incursions of bluetongue virus (BTV) occurred in the Mediterranean Basin involving European and surrounding Countries. The Middle East represents one of the most important gateways for the access of BTV in Europe. Limited data on the BTV situation in this area are available. In this perspective, an epidemiological survey on the presence of BTV in Lebanon was conducted. Of the 181 serum samples tested, 97 (mean = 53.6%; 95% CI: 46.3-60.7) resulted positive when tested for the presence of BTV antibodies by c-ELISA, of these 42 (mean = 42%; 95% CI: 32.8-51.8) serum samples were from sheep and 55 (mean = 67.9%; 95% CI: 57.1-77.1) serum samples were from goats. Fourteen blood samples (14/110; mean = 12.7%; 95% CI: 7.8-20.3), 6 (6/66; mean = 9.1%; 95% CI: 4.4-18.5) from sheep and 8 (8/44; mean = 18.2%; 95% CI: 9.6-32.0) from goats, were positive by qRT-PCR. The results with serum-neutralization assay and typing performed by RT-PCR confirmed that six BTV serotypes are currently circulating in Lebanon, and these serotypes are as follows: 1, 4, 6, 8, 16 and 24. This study is the first report that confirms the presence and circulation of BTV in Lebanon. © 2013 Blackwell Verlag GmbH.
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Sub-tesla-field magnetization of vibrated magnetic nanoreagents for screening tumor markers
NASA Astrophysics Data System (ADS)
Chieh, Jen-Jie; Huang, Kai-Wen; Shi, Jin-Cheng
2015-02-01
Magnetic nanoreagents (MNRs), consisting of liquid solutions and magnetic nanoparticles (MNPs) coated with bioprobes, have been widely used in biomedical disciplines. For in vitro tests of serum biomarkers, numerous MNR-based magnetic immunoassay methods or schemes have been developed; however, their applications are limited. In this study, a vibrating sample magnetometer (VSM) was used for screening tumor biomarkers based on the same MNRs as those used in other immunoassay methods. The examination mechanism is that examined tumor biomarkers are typically conjugated to the bioprobes coated on MNPs to form magnetic clusters. Consequently, the sub-Tesla-field magnetization (Msub-T) of MNRs, including magnetic clusters, exceeds that of MNRs containing only separate MNPs. For human serum samples, proteins other than the targeted biomarkers induce the formation of magnetic clusters with increased Msub-T because of weak nonspecific binding. In this study, this interference problem was suppressed by the vibration condition in the VSM and analysis. Based on a referenced Msub-T,0 value defined by the average Msub-T value of a normal person's serum samples, including general proteins and few tumor biomarkers, the difference ΔMsub-T between the measured Msub-T and the reference Msub-T,0 determined the expression of only target tumor biomarkers in the tested serum samples. By using common MNRs with an alpha-fetoprotein-antibody coating, this study demonstrated that a current VSM can perform clinical screening of hepatocellular carcinoma.
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Utility of the Department of Defense Serum Repository in Assessing Deployment Exposure.
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.
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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.
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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).
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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.
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Koestel, Zoe L; Backus, Robert C; Tsuruta, Kaoru; Spollen, William G; Johnson, Sarah A; Javurek, Angela B; Ellersieck, Mark R; Wiedmeyer, Charles E; Kannan, Kurunthachalam; Xue, Jingchuan; Bivens, Nathan J; Givan, Scott A; Rosenfeld, Cheryl S
2017-02-01
Bisphenol A (BPA) is a widely present endocrine disruptor chemical found in many household items. Moreover, this chemical can bioaccumulate in various terrestrial and aquatic sources; thereby ensuring continual exposure of animals and humans. For most species, including humans, diet is considered the primary route of exposure. However, there has been little investigation whether commercial-brands of dog foods contain BPA and potential health ramifications of BPA-dietary exposure in dogs. We sought to determine BPA content within dog food, whether short-term consumption of these diets increases serum concentrations of BPA, and potential health consequences, as assessed by potential hematological, serum chemistry, cortisol, DNA methylation, and gut microbiome changes, in dogs associated with short-term dietary exposure to BPA. Fourteen healthy privately-owned dogs were used in this study. Blood and fecal samples were collected prior to dogs being placed for two-weeks on one of two diets (with one considered to be BPA-free), and blood and fecal samples were collected again. Serum/plasma samples were analyzed for chemistry and hematology profiles, cortisol concentrations, 5-methylcytosine in lymphocytes, and total BPA concentrations. Fecal samples were used for microbiome assessments. Both diets contained BPA, and after two-weeks of being on either diet, dogs had a significant increase in circulating BPA concentrations (pre-samples=0.7±0.15ng/mL, post-samples=2.2±0.15ng/mL, p<0.0001). Elevated BPA concentrations positively correlated with increased plasma bicarbonate concentrations and associated with fecal microbiome alterations. Short-term feeding of canned dog food increased circulating BPA concentrations in dogs comparable to amounts detected in humans, and greater BPA concentrations were associated with serum chemistry and microbiome changes. Dogs, who share our internal and external environments with us, are likely excellent indicators of potential human health concerns to BPA and other environmental chemicals. These findings may also have relevance to aquatic and terrestrial wildlife. Copyright © 2016 Elsevier B.V. All rights reserved.
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Nassan, Feiby L; Chavarro, Jorge E; Mínguez-Alarcón, Lidia; Williams, Paige L; Tanrikut, Cigdem; Ford, Jennifer B; Dadd, Ramace; Perry, Melissa J; Hauser, Russ; Gaskins, Audrey J
2018-06-01
We examined associations of residential distance to major roadways, as a proxy for traffic-related air pollution exposures, with sperm characteristics and male reproductive hormones. The cohort included 797 men recruited from Massachusetts General Hospital Fertility Center between 2000 and 2015 to participate in fertility research studies. Men reported their residential addresses at enrollment and provided 1-6 semen samples and a blood sample during follow-up. We estimated the Euclidean distance to major roadways (e.g. interstates and highways: limited access highways, multi-lane highways (not limited access), other numbered routes, and major roads) using information from the Massachusetts Department of Geographic Information Systems. Semen parameters (1238 semen samples), sperm DNA integrity (389 semen samples), chromosomal disomy (101 semen samples), and serum reproductive hormones (405 serum samples) were assessed following standard procedures. Men in this cohort were primarily Caucasian (86%), not current smokers (92%), with a college or higher education (88%), and had an average age of 36 years and BMI of 27.7 kg/m 2 . The median (interquartile range) residential distance to a major roadway was 111 (37, 248) meters. Residential proximity to major roadways was not associated with semen parameters, sperm DNA integrity, chromosomal disomy, or serum reproductive hormone concentrations. The adjusted percent change (95% CI) in semen quality parameters associated with a 500 m increase in residential distance to a major roadway was -1.0% (-6.3, 4.5) for semen volume, 4.3% (-5.8, 15.7) for sperm concentration, 3.1% (-7.2, 14.5) for sperm count, 1.1% (-1.2, 3.4) for % total motile sperm, and 0.1% (-0.3, 0.5) for % morphologically normal sperm. Results were consistent when we modeled the semen parameters dichotomized according to WHO 2010 reference values. Residential distance to major roadways, as a proxy for traffic-related air pollution exposure, was not related to sperm characteristics or serum reproductive hormones among men attending a fertility clinic in Massachusetts. Copyright © 2018 Elsevier GmbH. All rights reserved.
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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
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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.
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NASA Astrophysics Data System (ADS)
Marcó P., L. M.; Jiménez, E.; Hernández C., E. A.; Rojas, A.; Greaves, E. D.
2001-11-01
The method of quantification using the Compton peak as an internal standard, developed in a previous work, was applied to the routine determination of Fe, Cu, Zn and Se in serum samples from normal individuals and cancer patients by total reflection X-ray fluorescence spectrometry. Samples were classified according to age and sex of the donor, in order to determine reference values for normal individuals. Results indicate that the Zn/Cu ratio and the Cu concentration could prove to be useful tools for cancer diagnosis. Significant differences in these parameters between the normal and cancer group were found for all age ranges. The multielemental character of the technique, coupled with the small amounts of sample required and the short analysis time make it a valuable tool in clinical analysis.
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Tess, D A; Cole, R O; Toler, S M
1995-12-15
A simple and highly sensitive reversed-phase fluorimetric HPLC method for the quantitation of droloxifene from rat, monkey, and human plasma as well as human serum is described. This assay employs solid-phase extraction and has a dynamic range of 25 to 10,000 pg/ml. Sample extraction (efficiencies > 86%) was accomplished using a benzenesulfonic acid (SCX) column with water and methanol rinses. Droloxifene and internal standard were eluted with 1 ml of 3.5% (v/v) ammonium hydroxide (30%) in methanol. Samples were quantitated using post-column UV-photochemical cyclization coupled with fluorimetric detection with excitation and emission wavelengths of 260 nm and 375 nm, respectively. Relative ease of sample extraction and short run times allow for the analysis of approximately 100 samples per day.
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Leptospira and Brucella antibodies in collared anteaters (Tamandua tetradactyla) in Brazilian zoos.
Sales, Indiara dos Santos; Folly, Márcio Manhães; Garcia, Luize Néli Nunes; Ramos, Tatiane Mendes Varela; da Silva, Mariana Cristina; Pereira, Martha Maria
2012-12-01
The presence of Leptospira spp. and Brucella spp. antibodies was investigated in serum samples from 28 collared anteaters (Tamandua tetradactyla) kept in seven Brazilian zoos. Sera were tested against 19 Leptospira serovars using microscopic agglutination. Samples reacted to the following serovars: two (7.14%) to Patoc, three (10.71%) to Tarrasovi, three (10.71%) to both Patoc and Tarrasovi, two (7.14%) to Wolffi, and one (3.57%) to Australis. Two (7.14%) samples reacted to the buffered Brucella antigen test, but no confirmatory reaction occurred using the 2-mercaptoethanol slow slide agglutination test. No sample was reactive in the agar gel immunodiffusion test for rugose species of Brucella. The presence of anti-leptospira agglutinins in captive T. tetradactyla serum indicates that this species may be susceptible to infection by these bacteria.
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Antonucci, FrancescoPaolo; Cento, Valeria; Sorbo, Maria Chiara; Manuelli, Matteo Ciancio; Lenci, Ilaria; Sforza, Daniele; Di Carlo, Domenico; Milana, Martina; Manzia, Tommaso Maria; Angelico, Mario; Tisone, Giuseppe; Perno, Carlo Federico; Ceccherini-Silberstein, Francesca
2017-08-01
We evaluated the performance of a rapid method to quantify HCV-RNA in the hepatic and extrahepatic compartments, by using for the first time the Abbott RealTime HCV-assay. Non-tumoral (NT), tumoral (TT) liver samples, lymph nodes and ascitic fluid from patients undergoing orthotopic-liver-transplantation (N=18) or liver resection (N=4) were used for the HCV-RNA quantification; 5/22 patients were tested after or during direct acting antivirals (DAA) treatment. Total RNA and DNA quantification from tissue-biopsies allowed normalization of HCV-RNA concentrations in IU/μg of total RNA and IU/10 6 liver-cells, respectively. HCV-RNA was successfully quantified with high reliability in liver biopsies, lymph nodes and ascitic fluid samples. Among the 17 untreated patients, a positive and significant HCV-RNA correlation between serum and NT liver-samples was observed (Pearson: rho=0.544, p=0.024). Three DAA-treated patients were HCV-RNA "undetectable" in serum, but still "detectable" in all tested liver-tissues. Differently, only one DAA-treated patient, tested after sustained-virological-response, showed HCV-RNA "undetectability" in liver-tissue. HCV-RNA was successfully quantified with high reliability in liver bioptic samples and extrahepatic compartments, even when HCV-RNA was "undetectable" in serum. Abbott RealTime HCV-assay is a good diagnostic tool for HCV quantification in intra- and extra-hepatic compartments, whenever a bioptic sample is available. Copyright © 2017 Elsevier B.V. All rights reserved.
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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.
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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…
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Klein, Joern; Hussain, Manzoor; Ahmad, Munir; Afzal, Muhammad; Alexandersen, Soren
2008-01-01
Background Foot-and-mouth disease (FMD) is endemic in Pakistan and causes huge economic losses. This work focus on the Landhi Dairy Colony (LDC), located in the suburbs of Karachi. LDC is the largest Buffalo colony in the world, with more than 300,000 animals (around 95% buffaloes and 5% cattle, as well as an unknown number of sheep and goats). Each month from April 2006 to April 2007 we collected mouth-swabs from apparently healthy buffaloes and cattle, applying a convenient sampling based on a two-stage random sampling scheme, in conjunction with participatory information from each selected farm. Furthermore, we also collected epithelium samples from animals with clinical disease, as well as mouth-swabs samples from those farms. In addition, we analysed a total of 180 serum samples randomly collecting 30 samples each month at the local slaughterhouse, from October 2006 to March 2007. Samples have been screened for FMDV by real-time RT-PCR and the partial or full 1D coding region of selected isolates has been sequenced. Serum samples have been analysed by applying serotype-specific antibody ELISA and non-structural proteins (NSP) antibody ELISA. Results FMDV infection prevalence at aggregate level shows an endemic occurrence of FMDV in the colony, with peaks in August 2006, December 2006 and February 2007 to March 2007. A significant association of prevalence peaks to the rainy seasons, which includes the coldest time of the year and the muslimic Eid-festival, has been demonstrated. Participatory information indicated that 88% of all questioned farmers vaccinate their animals. Analysis of the serum samples showed high levels of antibodies for serotypes O, A, Asia 1 and C. The median endpoint-titre for all tested serotypes, except serotype C, in VNT titration is at a serum dilution of equal or above 1/100. All 180 serum samples collected have been tested for antibodies against the non-structural proteins and all but four have been found positive. Out of the 106 swab-samples from apparently healthy and affected animals positive in real-time RT-PCR, we sequenced the partial or full 1D coding region from 58 samples. In addition we sequenced the full 1D coding region of 17 epithelium samples from animals with clinical signs of FMD. From all sequenced samples, swabs and epithelium, 19 belong to the regional PanAsia II lineage of serotype O and 56 to the A/Iran/2005 lineage of serotype A. Conclusion For an effective and realisable FMD control program in LDC, we suggest to introduce a twice annually mass vaccination of all buffaloes and cattle in the colony. These mass vaccinations should optimally take place shortly before the beginning of the two rainy periods, e.g. in June and September. Those vaccinations should, in our opinion, be in addition to the already individually performed vaccinations of single animals, as the latter usually targets only newly introduced animals. This suggested combination of mass vaccination of all large ruminants with the already performed individually vaccination should provide a continuous high level of herd immunity in the entire colony. Vaccines used for this purpose should contain the matching vaccine strains, i.e. as our results indicate antigens for A/Iran/2005 and the regional type of serotype O (PanAsia II), but also antigens of the, in this world region endemic, Asia 1 lineage should be included. In the long term it will be important to control the vaccine use, so that subclinical FMD will be avoided. PMID:18445264
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Massuger, L F; Thomas, C M; Segers, M F; Corstens, F H; Verheijen, R H; Kenemans, P; Poels, L G
1992-11-01
The development of human anti-mouse antibodies (HAMA) may cause problems in radioimmunotargeting studies, but may also improve survival of patients. To identify the presence of HAMA in blood samples from patients intravenously injected with 1 mg of 111In-labeled OV-TL3-F(ab')2, we developed three specific OV-TL 3-based HAMA assays and tested these along with two commercially available nonspecific HAMA assays (Sorin and Immunomedics). The specific assays were positive for HAMA with 10 postinjection serum samples from 7 patients. Eight of the 10 samples were also HAMA positive with one or both nonspecific HAMA assays. Conflicting results were observed with half the number of samples. The two nonspecific assays also reacted positively with another 11 serum samples from 5 patients including their preinjection samples. Despite some contradictory results, the nonspecific HAMA assays identify both pre-existent and Mab-induced HAMA, whereas the specific OV-TL3-based HAMA assays identify specific immune-responses occurring after the OV-TL 3 injection.
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[35 leptospira isolated from the vitreous body of 32 horses with recurrent uveitis (ERU)].
Brem, S; Gerhards, H; Wollanke, B; Meyer, P; Kopp, H
1999-01-01
130 vitreous samples, systematically collected in 1998 from 117 horses during vitrectomy, were cultured for the presence of leptospires. All horses suffered from equine recurrent uveitis (ERU), also known as periodic ophthalmia or moon blindness, and were treated surgically to combat painful attacks, and to preserve vision. In 35 out of 130 vitreous samples (35/130 = 26.9%), leptospires could be isolated. These isolates belong to the grippotyphosa serogroup (n = 31) and to the australis serogroup (n = 4). So, for the first time, leptospires were recovered from eyes in vivo in a large number of horses with ERU. Vitreous samples and one serum sample from each horse were also tested for leptospiral antibodies using the microscopic agglutination test (MAT). In 92 vitreous samples (92/130 = 70.7%) and 96 serum samples (96/117 = 82.0%) leptospiral antibodies were detected at a dilution of > 1:100. The presence of intact leptospires and specific antibodies in eyes affected with ERU demonstrates a local antibody production to leptospiral antigen. These results indicate an important etiological role of leptospires in equine recurrent uveitis.
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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
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Roberts, Michael S; Turner, David C; Owens, Thandranese S; Ramachandran, Abhijit; Wetmore, Cynthia; Throm, Stacy L; Stewart, Clinton F
2013-06-15
A LC-ESI-MS/MS method for the determination of crenolanib (CP-868,596) in human serum was developed and validated employing d4-CP-868,596 as an internal standard (ISTD). In addition to human serum, the method was also partially validated for crenolanib determination in human cerebrospinal fluid (CSF) samples. Sample aliquots (50μl of serum or CSF) were prepared for analysis using liquid-liquid extraction (LLE) with tert-butyl methyl ether. Chromatography was performed using a phenomenex Gemini C18 column (3μm, 100mm×4.6mm I.D.) in a column heater set at 50°C and an isocratic mobile phase (methanol/water/formic acid at a volume ratio of 25/25/0.15, v/v/v). The flow rate was 0.45mL/min, and the retention time for both analyte and ISTD was less than 3.5min. Samples were analyzed with an API-5500 LC-MS/MS system (ESI) in positive ionization mode coupled to a Shimadzu HPLC system. The ion transitions monitored were m/z 444.4→373.1 and m/z 448.2→374.2 for crenolanib and ISTD, respectively. The method was linear over the range of 5-1000ng/mL for serum and 0.5-1000ng/mL for CSF. For human serum, both intra-day and inter-day precision were <4%, while intra-day and inter-day accuracy were within 8% of nominal values. Recovery was greater than 50% for both the analyte and ISTD. For CSF samples, both intra-day and inter-day precision were <9% except at the lower limit of quantification (LLOQ) which was <17%. The intra-day and inter-day accuracy were within 11% of the nominal CSF concentrations. After validation, this method was successfully applied to the analysis of serial pharmacokinetic samples obtained from a child treated with oral crenolanib. Copyright © 2013 Elsevier B.V. All rights reserved.
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Humann-Ziehank, Esther; Tegtmeyer, Philip C; Seelig, Bjoern; Roehrig, Petra; Ganter, Martin
2013-11-19
This study was performed to demonstrate the widespread distribution and severity of selenium (Se) deficiency in sheep flocks and to evaluate the impact of influencing factors. In 150 flocks, ten serum samples of adult ewes were analysed for Se concentration. The farmers were interviewed concerning flock size, provision of mineral supplement, predominant form of husbandry (stationary fenced pasture/transhumance), predominant form of water provision (tap water/well/surface water) and predominant soil (sandy, silty/loamy, clay) in the area. The location of the flock was recorded as well as the production stage/season at the time of sampling. Intra-group variation and the validity to analyse pooled samples were tested. Pools of five samples correlated well with the mean of individually analysed samples. The intra-group range of serum Se concentration varied enormously (mean 45.4 ± 18.8 μg Se/l). About 60% of the flocks showed mean serum Se concentrations below 80 μg/l, 37.4% were below 60 μg Se/l, representing a Se deficient stage. Using mineral supplement in general was no key factor for Se status. Stationary flocks on fenced pasture had constantly higher mean serum Se concentrations during breeding (outdoors, August-November), lambing (mainly indoors, December-March) and lactation (outdoors, April-July), whereas flocks practising transhumance had significantly lower Se status, except during lambing. There was no significant correlation between the soil type and the Se status, but flocks in Southern Germany tend to show a lower Se status compared to Central and Northern Germany. Increasing flock size was associated with lower mean serum Se concentrations. In stationary flocks only, the use of surface water was accompanied by significantly lower Se status. Se deficiency is widespread in German sheep flocks. More than one third of the flocks showed Se deficiency, indicating the need to optimise the nutritional management. Factors raising suspicion of Se deficiency are large flocks, transhumance during lactation and the breeding season as well as surface water provision in stationary flocks.
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Detection of Toxoplasma gondii in the milk of naturally infected goats in the Northeast of Brazil.
Bezerra, M J G; Kim, P C P; Moraes, É P B X; Sá, S G; Albuquerque, P P F; Silva, J G; Alves, B H L S; Mota, R A
2015-08-01
The aim of the present study was to detect the genomic DNA of Toxoplasma gondii in milk samples from naturally infected goats in the state of Pernambuco, (Brazil). In total, 248 blood serum samples were collected and processed from lactating goats and then submitted to a search for antibodies to T. gondii through the indirect immunofluorescence reaction. Samples with a score of 64 or more were considered positive. In total, 248 milk samples were collected and processed from the same group of goats in order to study the DNA of T. gondii using the polymerase chain reaction (PCR) technique. In the serum samples, 56/248 (22.58%) of the animals were positive, whereas the DNA of the parasite was detected in 15/248 (6.05%) of the milk samples. Five of these 15 samples were animals who were also positive in the serology. This study reports the first occurrence of the elimination of T. gondii from the milk of naturally infected goats in the north-east of Brazil. It is suggested that the consumption of in natura goat milk may constitute a potential risk to the health of milk consumers in this region. © 2013 Blackwell Verlag GmbH.
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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.
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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.
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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
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Powell, Rebecca L R; Ouellette, Ian; Lindsay, Ross W; Parks, Christopher L; King, C Richter; McDermott, Adrian B; Morrow, Gavin
2013-06-01
Results from recent HIV-1 vaccine studies have indicated that high serum antibody (Ab) titers may not be necessary for Ab-mediated protection, and that Abs localized to mucosal sites might be critical for preventing infection. Enzyme-linked immunosorbent assay (ELISA) has been used for decades as the gold standard for Ab measurement, though recently, highly sensitive microsphere-based assays have become available, with potential utility for improved detection of Abs. In this study, we assessed the Bio-Plex(®) Suspension Array System for the detection of simian immunodeficiency virus (SIV)-specific Abs in rhesus macaques (RMs) chronically infected with SIV, whose serum or mucosal SIV-specific Ab titers were negative by ELISA. We developed a SIVmac239-specific 4-plex bead array for the simultaneous detection of Abs binding to Env, Gag, Pol, and Nef. The 4-plex assay was used to quantify SIV-specific serum IgG and rectal swab IgA titers from control (SIV-naive) and SIVmac239-infected RMs. The Bio-Plex assay specifically detected anti-SIV Abs in specimens from SIV-infected animals for all four analytes when compared to SIV-naive control samples (p≤0.04). Furthermore, in 70% of Env and 79% of Gag ELISA-negative serum samples, specific Ab was detected using the Bio-Plex assay. Similarly, 71% of Env and 48% of Gag ELISA-negative rectal swab samples were identified as positive using the Bio-Plex assay. Importantly, assay specificity (i.e., probability of true positives) was comparable to ELISA (94%-100%). The results reported here indicate that microsphere-based methods provide a substantial improvement over ELISA for the detection of Ab responses, aid in detecting specific Abs when analyzing samples containing low levels of Abs, such as during the early stages of a vaccine trial, and may be valuable in attempts to link protective efficacy of vaccines with induced Ab responses.
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Abu El-Asrar, Ahmed M; Berghmans, Nele; Al-Obeidan, Saleh A; Mousa, Ahmed; Opdenakker, Ghislain; Van Damme, Jo; Struyf, Sofie
2016-09-01
The purpose of this study was to determine levels of the cytokines IL-1β, IL-6, IL-21, IL-22, and IL-23 and the chemokines CXCL13, CCL19, CCL20, and CCL21 in aqueous humor (AH) samples from patients with specific uveitic entities. Paired serum samples (n = 13) and AH samples (n = 111) from patients with active idiopathic granulomatous uveitis (IGU) or with uveitis associated with HLA-B27-related inflammation, Behçet's disease (BD), Vogt-Koyanagi-Harada (VKH) disease, or sarcoidosis and control patients were analyzed in two different multiplex assays. Cytokines IL-1β, IL-21, IL-22, and IL-23 were not detected in any AH sample. Chemokine CCL21 concentrations in serum were significantly higher than those in AH. CCL19 levels in AH and serum were not significantly different. Levels of CCL20 and CXCL13 in AH were significantly higher than those in serum. IL-6 was not detected in serum samples. IL-6 AH levels were significantly higher in patients with HLA-B27-associated uveitis and in BD patients than in patients with VKH disease, sarcoidosis, and IGU (P < 0.0001). CCL20 AH levels were significantly higher in HLA-B27-associated uveitis than in BD, VKH, sarcoidosis, and IGU (P = 0.001), whereas CXCL13 AH levels were significantly higher in VKH disease and IGU than in HLA-B27-associated uveitis, BD, and sarcoidosis (P = 0.007). IL-6-driven immune responses are more potent in HLA-B27-associated uveitis and BD than in VKH disease, sarcoidosis, and IGU. CCL20 appears to be a specific biomarker of HLA-B27-associated uveitis, whereas CXCL13 appears to be a biomarker of VKH disease and IGU. Our findings suggest that IL-6, CCL20, and CXCL13 could serve as drug targets for treatment of specific clinical entities of endogenous uveitis.
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Serum chemistry comparisons between captive and free-ranging giraffes (Giraffa camelopardalis).
Schmidt, Debra A; Barbiers, Robyn B; Ellersieck, Mark R; Ball, Ray L; Koutsos, Elizabeth A; Griffin, Mark E; Grobler, Douw; Citino, Scott B; Bush, Mitchell
2011-03-01
Serum chemistry analyses were compared between captive and free-ranging giraffes (Giraffa camelopardalis) in an attempt to better understand some of the medical issues seen with captive giraffes. Illnesses, including peracute mortality, energy malnutrition, pancreatic disease, urolithiasis, hoof disease, and severe intestinal parasitism, may be related to zoo nutrition and management issues. Serum samples were collected from 20 captive giraffes at 10 United States institutions. Thirteen of the captive animal samples were collected from animals trained for blood collection; seven were banked samples obtained from a previous serum collection. These samples were compared with serum samples collected from 24 free-ranging giraffes in South Africa. Differences between captive and free-ranging giraffes, males and females, and adults and subadults were analyzed by using a 2 x 2 x 2 factorial and Fisher's least significant difference for mean separation; when necessary variables were ranked and analyzed via analysis of variance. Potassium and bilirubin concentrations and alanine aminotransferase (ALT) activities were different between captive and free-ranging giraffes, but all fell within normal bovid reference ranges. The average glucose concentration was significantly elevated in free-ranging giraffes (161 mg/dl) compared with captive giraffes (113 mg/dl). All giraffes in this study had glucose concentrations higher than bovine (42-75 mg/ dl) and caprine (48-76 mg/dl) reference ranges. Differences were also seen in lipase, chloride, and magnesium though these findings are likely not clinically significant. There were no differences detected between sexes. Adults had higher concentrations of potassium, total protein, globulins, and chloride and higher gamma glutamyltransferase activities, whereas subadults had higher concentrations of phosphorus. Within the captive group, nonimmobilized animals had higher concentrations of total protein and globulins. Captive giraffe diets need further investigation to determine if the differences seen in this study, especially glucose and bilirubin concentrations and ALT activities, may result in some health problems often seen in captive giraffes.
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Cho, Sung E; Kim, Sollip; Kim, Young D; Lee, Hyojung; Seo, Dong H; Song, Junghan; Um, Tae H; Cho, Chong R; Kim, Nam H; Hwang, Jong H
2017-09-01
Background We evaluated the performance of ultra-performance liquid chromatography-tandem mass spectrometry to measure serum 3-epi-25-hydroxyvitamin D 3 , 25-hydroxyvitamin D 3 and 25-hydroxyvitamin D 2 concentrations in 519 infant, paediatric and adolescent serum samples in Korea. Methods We used a Kinetex XB-C18 column and isocratic methanol/water (77.5/22.5, v/v) with 0.025% (v/v) high-performance liquid chromatography solvent additive flowing at 0.25 mL/min, yielding an 11 min/sample run time. A TQD triple quadrupole mass spectrometer in electrospray ionization positive ion mode with multiple reaction monitoring transition via an MSMS vitamin D kit was used to evaluate precision, carryover, ion suppression and linearity. Samples were prepared using the 4-phenyl-1,2,4-triazoline-3,5-dione derivatization method. Results Intra- and inter-run precisions were 1.23-13.28% and 1.02-10.08%, respectively. Group carryovers were -0.27% and 0.10%, respectively. There was no ion suppression. The calibration curve showed good linearity from calibrator Level 1 (11.75 nmol/L) to 6 (375 nmol/L) with R 2 > 0.9999. The 3-epi-25-hydroxyvitamin D 3 and 25-hydroxyvitamin D 3 peaks were clearly separated in the extracted ion chromatogram. Infant serum samples 3-epi-25-hydroxyvitamin D 3 concentrations were significantly higher than paediatric and adolescent concentrations. Conclusions The ultra-performance liquid chromatography-tandem mass spectrometry assay performed acceptably, clearly separating 3-epi-25-hydroxyvitamin D 3 from 25-hydroxyvitamin D 3 . High 3-epi-25-hydroxyvitamin D 3 concentrations were observed in infant but not in paediatric and adolescent serum samples.
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Clustering of Toxoplasma gondii Infections Within Families of Congenitally Infected Infants
Contopoulos-Ioannidis, Despina; Wheeler, Kelsey M.; Ramirez, Raymund; Press, Cindy; Mui, Ernest; Zhou, Ying; Van Tubbergen, Christine; Prasad, Sheela; Maldonado, Yvonne; Withers, Shawn; Boyer, Kenneth M.; Noble, A. Gwendolyn; Rabiah, Peter; Swisher, Charles N.; Heydemann, Peter; Wroblewski, Kristen; Karrison, Theodore; Grigg, Michael E.; Montoya, Jose G.; McLeod, Rima
2015-01-01
Background. Family clusters and epidemics of toxoplasmosis in North, Central, and South America led us to determine whether fathers of congenitally infected infants in the National Collaborative Chicago-Based Congenital Toxoplasmosis Study (NCCCTS) have a high incidence of Toxoplasma gondii infection. Methods. We analyzed serum samples collected from NCCCTS families between 1981 and 2013. Paternal serum samples were tested for T. gondii antibodies with immunoglobulin (Ig) G dye test and IgM enzyme-linked immunosorbent assay. Additional testing of paternal serum samples was performed with differential-agglutination and IgG avidity tests when T. gondii IgG and IgM results were positive and serum samples were collected by the 1-year visit of the congenitally infected child. Prevalence of paternal seropositivity and incidence of recent infection were calculated. We analyzed whether certain demographics, maternal parasite serotype, risk factors, or maternal/infant clinical manifestations were associated with paternal T. gondii infection status. Results. Serologic testing revealed a high prevalence (29 of 81; 36%) of T. gondii infection in fathers, relative to the average seropositivity rate of 9.8% for boys and men aged 12–49 years in the United States between 1994 and 2004 (P < .001). Moreover, there was a higher-than-expected incidence of recent infections among fathers with serum samples collected by the 1-year visit of their child (6 of 45; 13%; P < .001). No demographic patterns or clinical manifestations in mothers or infants were associated with paternal infections, except for sandbox exposure. Conclusions. The high prevalence of chronic and incidence of recent T. gondii infections in fathers of congenitally infected children indicates that T. gondii infections cluster within families in North America. When a recently infected person is identified, family clustering and community risk factors should be investigated for appropriate clinical management. PMID:26405150
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Clustering of Toxoplasma gondii Infections Within Families of Congenitally Infected Infants.
Contopoulos-Ioannidis, Despina; Wheeler, Kelsey M; Ramirez, Raymund; Press, Cindy; Mui, Ernest; Zhou, Ying; Van Tubbergen, Christine; Prasad, Sheela; Maldonado, Yvonne; Withers, Shawn; Boyer, Kenneth M; Noble, A Gwendolyn; Rabiah, Peter; Swisher, Charles N; Heydemann, Peter; Wroblewski, Kristen; Karrison, Theodore; Grigg, Michael E; Montoya, Jose G; McLeod, Rima
2015-12-15
Family clusters and epidemics of toxoplasmosis in North, Central, and South America led us to determine whether fathers of congenitally infected infants in the National Collaborative Chicago-Based Congenital Toxoplasmosis Study (NCCCTS) have a high incidence of Toxoplasma gondii infection. We analyzed serum samples collected from NCCCTS families between 1981 and 2013. Paternal serum samples were tested for T. gondii antibodies with immunoglobulin (Ig) G dye test and IgM enzyme-linked immunosorbent assay. Additional testing of paternal serum samples was performed with differential-agglutination and IgG avidity tests when T. gondii IgG and IgM results were positive and serum samples were collected by the 1-year visit of the congenitally infected child. Prevalence of paternal seropositivity and incidence of recent infection were calculated. We analyzed whether certain demographics, maternal parasite serotype, risk factors, or maternal/infant clinical manifestations were associated with paternal T. gondii infection status. Serologic testing revealed a high prevalence (29 of 81; 36%) of T. gondii infection in fathers, relative to the average seropositivity rate of 9.8% for boys and men aged 12-49 years in the United States between 1994 and 2004 (P < .001). Moreover, there was a higher-than-expected incidence of recent infections among fathers with serum samples collected by the 1-year visit of their child (6 of 45; 13%; P < .001). No demographic patterns or clinical manifestations in mothers or infants were associated with paternal infections, except for sandbox exposure. The high prevalence of chronic and incidence of recent T. gondii infections in fathers of congenitally infected children indicates that T. gondii infections cluster within families in North America. When a recently infected person is identified, family clustering and community risk factors should be investigated for appropriate clinical management. © The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.
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Inoue, K; Yoshimura, Y; Makino, T; Nakazawa, H
2000-11-01
Alkylphenols can affect human health because they disrupt the endocrine system. In this study, an analytical method for determining trace amounts of 4-nonylphenol (NP) and 4-octylphenol (OP) in human blood samples was developed. Reversed-phase HPLC with multi-electrode electrochemical coulometric-array detection was used for the determination of NP and OP in plasma and serum samples prepared with a solid-phase extraction method. The separation was achieved using an isocratic mobile phase of 0.7% phosphoric acid-acetonitrile with a C18 reversed phase column. The detection limits of NP and OP were 1.0 and 0.5 ng ml-1, respectively. The recoveries of NP and OP added to human plasma samples were above 70.0% with a relative standard deviation of less than 15.5%. The method was found to be applicable to the determination of NP and OP in various human blood samples such as serum and plasma.
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Ergünay, Koray; Özkul, Aykut
2011-04-01
West Nile virus (WNV) infections may trigger febrile conditions and/or neuroinvasive disease in a portion of the exposed individuals. Serosurveillance data from various regions of Turkey indicate WNV activity. The aim of this study was to confirm the antibody specificity of the serum samples via virus neutralization assay, previously reported to be reactive for WNV IgM. The samples originated from two individuals with the preliminary diagnosis of aseptic meningitis/encephalitis of unknown etiology in 2009 and had been classified as probable WNV infections. Cerebrospinal fluid and sera samples of these patients had been evaluated as negative for WNV RNA and IgG antibodies. Only one serum sample could be included in the neutralization assay due to the limited amounts in the current investigation. The sample was observed as positive in dilutions of 1/20 and 1/40, thus confirming the diagnosis of WNV-related central nervous system infection in a 62 year-old female patient from Ankara, Central Anatolia, Turkey.
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Feasibility of Using Convalescent Plasma Immunotherapy for MERS-CoV Infection, Saudi Arabia.
Arabi, Yaseen M; Hajeer, Ali H; Luke, Thomas; Raviprakash, Kanakatte; Balkhy, Hanan; Johani, Sameera; Al-Dawood, Abdulaziz; Al-Qahtani, Saad; Al-Omari, Awad; Al-Hameed, Fahad; Hayden, Frederick G; Fowler, Robert; Bouchama, Abderrezak; Shindo, Nahoko; Al-Khairy, Khalid; Carson, Gail; Taha, Yusri; Sadat, Musharaf; Alahmadi, Mashail
2016-09-01
We explored the feasibility of collecting convalescent plasma for passive immunotherapy of Middle East respiratory syndrome coronavirus (MERS-CoV) infection by using ELISA to screen serum samples from 443 potential plasma donors: 196 patients with suspected or laboratory-confirmed MERS-CoV infection, 230 healthcare workers, and 17 household contacts exposed to MERS-CoV. ELISA-reactive samples were further tested by indirect fluorescent antibody and microneutralization assays. Of the 443 tested samples, 12 (2.7%) had a reactive ELISA result, and 9 of the 12 had reactive indirect fluorescent antibody and microneutralization assay titers. Undertaking clinical trials of convalescent plasma for passive immunotherapy of MERS-CoV infection may be feasible, but such trials would be challenging because of the small pool of potential donors with sufficiently high antibody titers. Alternative strategies to identify convalescent plasma donors with adequate antibody titers should be explored, including the sampling of serum from patients with more severe disease and sampling at earlier points during illness.
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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.
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Sukla, Soumi; Ghosh, Anisa; Saha, Rajdeep; De, Abhishek; Adhya, Samit; Biswas, Subhajit
2018-06-13
Zika virus infections have recently been reported in many dengue-endemic areas globally. Both dengue (DENV) and Zika (ZIKV) virus are transmitted by Aedes mosquitoes, raising the possibility of mixed infections in both vector and host. We evaluated DENV and ZIKV prevalence in human and vector samples in Kolkata, a DENV-endemic city. Blood samples were collected from 70 patients presenting dengue-like fever symptoms at a hospital in Kolkata during 2015-16. Serum was obtained and tested for DENV infection by DENV NS1-based ELISA. Adult (n=8) and larval stages (n=12) of Aedes were also collected. A RT-PCR-based screening of both viruses supplemented by amplicon sequencing was performed. Of the 70 samples, 20 DENV NS1-positive serum samples were used for detailed molecular study for DENV infection. Eighteen of these (90 %) were positive by hemi-nested serotype-specific RT-PCR for DENV1/2/3, with four samples showing evidence of DENV2-3 or DENV1-3 mixed infection. None were ZIKV-positive using NS5 or ENV-based PCR, though weak amplification of a DENV1 NS5 sequence was detected in three serum samples indicating cross-reactivity of the primers. All mosquito samples were ZIKV-negative, whereas 5/8 (63 %) of adult mosquitoes and 11/12 (92 %) of larvae were DENV3-positive. Both host and vector samples showed absence of ZIKV but high prevalence of DENV. The high rate of infection of larvae with DENV is suggestive of trans-ovarial transmission that could contribute to the surge of human infections during each post-monsoon season. It would be important to guard against false positives using the available Zika-reporting primer sets.
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Dayre McNally, J; Matheson, Loren A; Sankaran, Koravangattu; Rosenberg, Alan M
2008-11-01
This study compared 25-hydroxyvitamin D [25(OH)D] measurements in capillary and venous blood samples collected, respectively by fingerprick and venipuncture. Capillary blood for measuring 25(OH)D has potential advantages by reducing blood volume required (2mL versus 0.3mL for venipuncture and capillary sampling, respectively), facilitating blood collection for those populations in whom venipuncture is difficult (e.g. infants and children), improving patient convenience and reducing costs associated with phlebotomy. The results demonstrated a highly significant relationship between 25(OH)D levels in serum derived from venous and capillary blood samples (r(2)=0.901). Despite statistically higher 25(OH)D levels in fingerprick samples (108+/-9nmol/L) compared with venipuncture samples (90+/-7nmol/L), the correlation between venous and capillary samples provides support for this approach as a practical alternative to venipuncture for vitamin D determination. However, clinical application may require the incorporation of a correction factor for the assessment of insufficiency, and research studies should avoid using the two methods interchangeably. Studying vitamin D's role in health and disease requires collection techniques and measurement methods that are reliable, reproducible, easily accessible, inexpensive and minimally burdensome to the patient. The option to collect patient samples by fingerprick may facilitate the collection process.
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Frölich, K; Li, H; Müller-Doblies, U
1998-10-01
A total of 486 serum samples collected from several species of both free-living and captive cervids in Germany was examined for antibodies against malignant catarrhal fever (MCF)-associated viruses (MCFV) by a competitive-inhibition enzyme-linked immunosorbent assay (CI-ELISA). Eleven (2%) of these samples were positive for antibodies against MCFV. Among 157 serum samples collected from 16 different species of captive deer including four (7%) of 54 fallow deer and one (7%) of 14 sika deer (Cervus nippon) were seropositive. Among 329 samples from three species of free-ranging deer, including 253 roe deer (Capreolus capreolus), 22 red deer (Cervus elaphus) and 54 fallow deer (Cervus dama), only fallow deer were antibody-positive. Of the 25 fallow deer samples collected between 1990 and 1993, four (16%) were seropositive. Among 29 free-ranging fallow deer samples collected in the hunting period 1996-1997, antibodies to MCFV were detected in two (7%) of these sera. All of these fallow deer samples were collected from a circumscribed area in northern Germany. In the same area a high seroprevalence (72%) to MCFV was observed in domestic sheep (n = 50). Among 20 sheep samples (buffy coat) and 15 fallow deer samples (spleen or lymph nodes) examined for ovine herpesvirus 2 (OvHV-2) by PCR, all 20 sheep samples examined were OvHV-2 positive, but all of the 15 fallow deer samples, including seven seropositive deer, were OvHV-2 negative.
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[Detection of antibodies against enterovirus 71 in the sera of Moscow residents].
Kravchenko, A T; Omel'chenko, T N
1984-03-01
Twenty three out of 55 serum samples obtained from subjects aged 20-55 years have proved to be seropositive, 15 of the positive serum samples (i.e. more than 50%) possessing the titer greater than or equal to 1:16. The presence of specific antibodies in the sera of the inhabitants of Moscow suggests that either enterovirus 71 or some other virus whose antigens are partly identical to those of enterovirus 71 apparently circulates among the population of Moscow.
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Persistence of Immunity Acquired after a Single Dose of Rubella Vaccine in Japan.
Okafuji, Takao; Okafuji, Teruo; Nakayama, Tetsuo
2016-05-20
To date, Takahashi, Matsuura, and TO-336 strains of live-attenuated rubella vaccine have been used in Japan. Japan implemented a single-dose rubella vaccination program until 2006. However, few reports are available on the persistence of immunity after this vaccination program. We collected 276 serum samples from January 2009 to December 2011 at Okafuji Pediatric Clinic and assessed the immune status of these samples against rubella virus during 1-10 years after vaccination with a single dose of Takahashi rubella vaccine. Regional outbreak of rubella did not occur during 1999-2011. The collected serum samples were tested for antibodies against the rubella virus by performing a standard hemagglutination inhibition (HAI) test. Our results showed that all the tested serum samples contained antibodies against the rubella virus 10 years after the vaccination. Geometric mean titer of HAI antibodies was 1:180 and decreased to 1:68 at 10 years after the vaccination. The levels of HAI antibodies decreased logarithmically with time after the vaccination. In conclusion, vaccine-acquired immunity after vaccination with a single dose of live-attenuated Takahashi rubella vaccine was retained for at least 10 years when rubella was under regional control.
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The Antimicrobial Peptide Lysozyme Is Induced after Multiple Trauma
Klüter, Tim; Fitschen-Oestern, Stefanie; Lippross, Sebastian; Weuster, Matthias; Pufe, Thomas; Tohidnezhad, Mersedeh; Beyer, Andreas; Seekamp, Andreas; Varoga, Deike
2014-01-01
The antimicrobial peptide lysozyme is an important factor of innate immunity and exerts high potential of antibacterial activity. In the present study we evaluated the lysozyme expression in serum of multiple injured patients and subsequently analyzed their possible sources and signaling pathways. Expression of lysozyme was examined in blood samples of multiple trauma patients from the day of trauma until 14 days after trauma by ELISA. To investigate major sources of lysozyme, its expression and regulation in serum samples, different blood cells, and tissue samples were analysed by ELISA and real-time PCR. Neutrophils and hepatocytes were stimulated with cytokines and supernatant of Staphylococcus aureus. The present study demonstrates the induction and release of lysozyme in serum of multiple injured patients. The highest lysozyme expression of all tested cells and tissues was detected in neutrophils. Stimulation with trauma-related factors such as interleukin-6 and S. aureus induced lysozyme expression. Liver tissue samples of patients without trauma show little lysozyme expression compared to neutrophils. After stimulation with bacterial fragments, lysozyme expression of hepatocytes is upregulated significantly. Toll-like receptor 2, a classic receptor of Gram-positive bacterial protein, was detected as a possible target for lysozyme induction. PMID:25258475
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Sankar, Surya; Harshan, Hiron M; Somarajan, S R; Srivastava, S K
2010-06-01
A recombinant leptospiral lipoprotein, LigB, was evaluated for use in the diagnosis of bovine leptospirosis by enzyme-linked immunosorbent assay (rLigB IgG ELISA). The standard reference test (Microscopic agglutination test, MAT) of 200 serum samples from cattle suspected of leptospirosis showed that 95 (47.5%) samples had positive agglutination titres, which ranged from 100 to 1600. In rLigB IgG ELISA, 49% of the samples were positive. Sensitivity of IgG ELISA for 95 bovine sera, which had MAT titres of greater than or equal to 100, were 100%. ELISA showed a specificity of 97.1% with 105 bovine sera, which were negative at a 1:50 dilution in MAT for Leptospira interrogans serovars. The results of ELISA and MAT correspond very good. When analytical specificity of IgG ELISA was evaluated using bovine serum samples from animals showing the serum antibodies to other pathogens, no cross-reaction was observed. Thus the recombinant LigB IgG ELISA can be used instead of the MAT as an aid to the diagnosis of bovine leptospirosis. Copyright 2009. Published by Elsevier India Pvt Ltd.
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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...
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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...
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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...
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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...
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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...
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Higher serum 25-hydroxyvitamin D concentrations are related to a reduced risk of depression.
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.
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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.
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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.
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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
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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.
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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)
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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.
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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
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Sampling and analysis techniques for monitoring serum for trace elements.
Ericson, S P; McHalsky, M L; Rabinow, B E; Kronholm, K G; Arceo, C S; Weltzer, J A; Ayd, S W
1986-07-01
We describe techniques for controlling contamination in the sampling and analysis of human serum for trace metals. The relatively simple procedures do not require clean-room conditions. The atomic absorption and atomic emission methods used have been applied in studying zinc, copper, chromium, manganese, molybdenum, selenium, and aluminum concentrations. Values obtained for a group of 16 normal subjects agree with the most reliable values reported in the literature, obtained by much more elaborate techniques. All of these metals can be measured in 3 to 4 mL of serum. The methods may prove especially useful in monitoring concentrations of essential trace elements in blood of patients being maintained on total parenteral nutrition.
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Xie, Gui-lin; Yin, Jun-xia; Duan, Xin-yu; Feng, Xiao-hui; Wang, Yang; Jiang, Kai; Chen, Xin-mei
2015-06-01
One hundred and fifty-nine serum samples from hydatid disease patients and 80 serum samples from patients with other liver diseases were detected by gold-immunochromatographic assay, and read by naked eyes and the gold-immunochromatographic test strip reader. The sensitivity, specificity, and accuracy of eye-based method was 92.4% (147/159), 85.0% (68/80), and 89.9% (215/239), which was lower than that of the reader detection (95.6%, 93.7%, 95.0%, respectively). While, its false negative rate (7.5%, 12/159) and false positive rate (15.0%, 12/80) was higher than that of the reader detection (4.4% and 6.3%, respectively).
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Salgado, R; Benoy, I; Vermeulen, P; van Dam, P; Van Marck, E; Dirix, L
2004-01-01
In order to investigate whether the high bFGF serum levels encountered in cancer patients are derived from the tumour, we analysed serum bFGF levels in 18 untreated randomly selected patients with operable colorectal, cervical and ovarian cancer in the blood draining the tumour, i.e., in mesenteric and uterine veins, and compared these with arterial samples. No significantly elevated bFGF levels were found in the veins draining the tumours compared with arterial samples in our patient population. This suggests that, in contrast to what is generally presumed, serum bFGF levels might also be derived from other sources besides the tumour, e.g., platelets.
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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.
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Jha, Ashish Kumar
2015-01-01
Glomerular filtration rate (GFR) estimation by plasma sampling method is considered as the gold standard. However, this method is not widely used because the complex technique and cumbersome calculations coupled with the lack of availability of user-friendly software. The routinely used Serum Creatinine method (SrCrM) of GFR estimation also requires the use of online calculators which cannot be used without internet access. We have developed user-friendly software "GFR estimation software" which gives the options to estimate GFR by plasma sampling method as well as SrCrM. We have used Microsoft Windows(®) as operating system and Visual Basic 6.0 as the front end and Microsoft Access(®) as database tool to develop this software. We have used Russell's formula for GFR calculation by plasma sampling method. GFR calculations using serum creatinine have been done using MIRD, Cockcroft-Gault method, Schwartz method, and Counahan-Barratt methods. The developed software is performing mathematical calculations correctly and is user-friendly. This software also enables storage and easy retrieval of the raw data, patient's information and calculated GFR for further processing and comparison. This is user-friendly software to calculate the GFR by various plasma sampling method and blood parameter. This software is also a good system for storing the raw and processed data for future analysis.
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Moraes, Claudia Trigo Pedroso; Oliveira, Danielle Bruna Leal; Campos, Angelica Cristine Almeida; Bosso, Patricia Alves; Lima, Hildener Nogueira; Stewien, Klaus Eberhard; Gilio, Alfredo Elias; Vieira, Sandra Elisabete; Botosso, Viviane Fongaro; Durigon, Edison Luiz
2015-01-01
Human respiratory syncytial virus (HRSV) is an important respiratory pathogens among children between zero-five years old. Host immunity and viral genetic variability are important factors that can make vaccine production difficult. In this work, differences between biological clones of HRSV were detected in clinical samples in the absence and presence of serum collected from children in the convalescent phase of the illness and from their biological mothers. Viral clones were selected by plaque assay in the absence and presence of serum and nucleotide sequences of the G2 and F2 genes of HRSV biological clones were compared. One non-synonymous mutation was found in the F gene (Ile5Asn) in one clone of an HRSV-B sample and one non-synonymous mutation was found in the G gene (Ser291Pro) in four clones of the same HRSV-B sample. Only one of these clones was obtained after treatment with the child's serum. In addition, some synonymous mutations were determined in two clones of the HRSV-A samples. In conclusion, it is possible that minor sequences could be selected by host antibodies contributing to the HRSV evolutionary process, hampering the development of an effective vaccine, since we verify the same codon alteration in absence and presence of human sera in individual clones of BR-85 sample. PMID:25742274
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Moraes, Claudia Trigo Pedroso; Oliveira, Danielle Bruna Leal; Campos, Angelica Cristine Almeida; Bosso, Patricia Alves; Lima, Hildener Nogueira; Stewien, Klaus Eberhard; Gilio, Alfredo Elias; Vieira, Sandra Elisabete; Botosso, Viviane Fongaro; Durigon, Edison Luiz
2015-02-01
Human respiratory syncytial virus (HRSV) is an important respiratory pathogens among children between zero-five years old. Host immunity and viral genetic variability are important factors that can make vaccine production difficult. In this work, differences between biological clones of HRSV were detected in clinical samples in the absence and presence of serum collected from children in the convalescent phase of the illness and from their biological mothers. Viral clones were selected by plaque assay in the absence and presence of serum and nucleotide sequences of the G2 and F2 genes of HRSV biological clones were compared. One non-synonymous mutation was found in the F gene (Ile5Asn) in one clone of an HRSV-B sample and one non-synonymous mutation was found in the G gene (Ser291Pro) in four clones of the same HRSV-B sample. Only one of these clones was obtained after treatment with the child's serum. In addition, some synonymous mutations were determined in two clones of the HRSV-A samples. In conclusion, it is possible that minor sequences could be selected by host antibodies contributing to the HRSV evolutionary process, hampering the development of an effective vaccine, since we verify the same codon alteration in absence and presence of human sera in individual clones of BR-85 sample.
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Perez-Guaita, David; Kuligowski, Julia; Quintás, Guillermo; Garrigues, Salvador; Guardia, Miguel de la
2013-03-30
Locally weighted partial least squares regression (LW-PLSR) has been applied to the determination of four clinical parameters in human serum samples (total protein, triglyceride, glucose and urea contents) by Fourier transform infrared (FTIR) spectroscopy. Classical LW-PLSR models were constructed using different spectral regions. For the selection of parameters by LW-PLSR modeling, a multi-parametric study was carried out employing the minimum root-mean square error of cross validation (RMSCV) as objective function. In order to overcome the effect of strong matrix interferences on the predictive accuracy of LW-PLSR models, this work focuses on sample selection. Accordingly, a novel strategy for the development of local models is proposed. It was based on the use of: (i) principal component analysis (PCA) performed on an analyte specific spectral region for identifying most similar sample spectra and (ii) partial least squares regression (PLSR) constructed using the whole spectrum. Results found by using this strategy were compared to those provided by PLSR using the same spectral intervals as for LW-PLSR. Prediction errors found by both, classical and modified LW-PLSR improved those obtained by PLSR. Hence, both proposed approaches were useful for the determination of analytes present in a complex matrix as in the case of human serum samples. Copyright © 2013 Elsevier B.V. All rights reserved.
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Xiao, Qin; Hu, Bin; Duan, Jiankun; He, Man; Zu, Wanqing
2007-10-01
A novel method for the analysis of four polybrominated diphenyl ethers (PBDEs) in environmental and human serum samples based on hollow fiber-liquid phase microextraction (HF-LPME) followed by gas chromatography-inductively coupled plasma mass spectrometric (GC-ICP-MS) detection has been developed. The organic solvent in the porous hollow fiber was first dipped into the sample for extraction at a given time, and the retracted organic phase was introduced into the GC-ICP-MS for analysis. The addition of methanol has a strong effect on the HF-LPME extraction efficiency. Other significant parameters affecting the extraction efficiency of HF-LPME were also studied. HF-LPME was effective to isolate the analytes from the complex matrix. Under the optimized conditions, the detection limits of the proposed method varied from 15.2 to 40.5 ng/L. In general, the relative standard deviations (RSDs) were less than 10%. Good linearity was obtained with the correlation coefficients all better than 0.999. The proposed method is simple, quick, few microliters of organic solvent required, and is especially suitable for the analysis of the real sample with small amount available. The overall process of HF-LPME with GC-ICP-MS was applied successfully for the determination of polybrominated diphenyl ethers (PBDEs) in environmental and spiked human serum samples, and the results were satisfactory.
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Acute and chronic impact of smoking on salivary and serum total antioxidant capacity.
Kurku, Huseyin; Kacmaz, Murat; Kisa, Ucler; Dogan, Ozlem; Caglayan, Osman
2015-02-01
To investigate increased oxidative stress in saliva of smokers along with their serum. The case-control study was conducted from September to December 2008 in the Department of Biochemistry of the Medical School, Kirikkale University, Kirikkale, Turkey. A blood sample and saliva samples before and after smoking were collected from the smokers, while blood and saliva samples were taken from the controls. All samples were taken concurrently. The samples were measured for total antioxidant capacity, total oxidant stress, oxidative stress index, malondialdehyde, nitric oxide and total sulfhydryl groups. Levels of superoxide dismutase and glutathione peroxidase were also measured on saliva samples. SPSS 13 was used for statistical analysis. Of the 54 subjects in the study, 27(50%) were smokers with a mean age of 28.4 ± 5.42 years, and 27 (50%) were controls with a mean age of 29.7 ± 8.03 years. Total oxidant stress, oxidative stress index, malondialdehyde and nitric oxide levels were found higher in the serum samples of smokers (p < 0.05), and the levels of total sulfhydryl groups in smokers were lower compared with the controls (p < 0.05). Among the smokers, salivary malondialdehyde levels were higher before and after smoking (p < 0.05), glutathione peroxidase levels were lower than the controls, and salivary nitric oxide levels after smoking were higher than both those of the control group and the levels before smoking (p < 0.05). Both acute and chronic increased oxidative conditions may be a significant sign of the destructive effects of smoking. The investigation of disorders in smokers concerning oxidative stress will be beneficial in terms of novel approaches and treatment modalities.
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Mayer, Horst; Brümmer, Jens; Brinkmann, Thomas
2011-01-01
To implement Lean Six Sigma in our central laboratory we conducted a project to measure single pre-analytical steps influencing turnaround time (TAT) of emergency department (ED) serum samples. The traditional approach of extracting data from the Laboratory Information System (LIS) for a retrospective calculation of a mean TAT is not suitable. Therefore, we used radiofrequency identification (RFID) chips for real time tracking of individual samples at any pre-analytical step. 1,200 serum tubes were labelled with RFID chips and were provided to the emergency department. 3 RFID receivers were installed in the laboratory: at the outlet of the pneumatic tube system, at the centrifuge, and in the analyser area. In addition, time stamps of sample entry at the automated sample distributor and communication of results from the analyser were collected from LIS. 1,023 labelled serum tubes arrived at our laboratory. 899 RFID tags were used for TAT calculation. The following transfer times were determined (median 95th percentile in min:sec): pneumatic tube system --> centrifuge (01:25/04:48), centrifuge --> sample distributor (14:06/5:33), sample distributor --> analysis system zone (02:39/15:07), analysis system zone --> result communication (12:42/22:21). Total TAT was calculated at 33:19/57:40 min:sec. Manual processes around centrifugation were identified as a major part of TAT with 44%/60% (median/95th percentile). RFID is a robust, easy to use, and error-free technology and not susceptible to interferences in the laboratory environment. With this study design we were able to measure significant variations in a single manual sample transfer process. We showed that TAT is mainly influenced by manual steps around the centrifugation process and we concluded that centrifugation should be integrated in solutions for total laboratory automation.
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Opriessnig, Tanja; Hemann, Michelle; Johnson, John K; Heinen, Sheila; Giménez-Lirola, Luis G; O'Neill, Kevin C; Hoang, Hai; Yoon, Kyoung-Jin; Gottschalk, Marcelo; Halbur, Patrick G
2013-01-01
Accurate diagnosis of exposure to Actinobacillus pleuropneumoniae is important for maintaining negative farms. In the present study, the ability of a dual-plate complement fixation (CF) assay and 3 commercially available enzyme-linked immunosorbent assays (ELISAs; quad-plate ELISA-1, single-plate ELISA-2, and single-plate ELISA-3) in detecting serological evidence of A. pleuropneumoniae exposure was compared using serum samples of experimentally infected or vaccinated pigs, or field samples from the United States. Forty-two pigs were divided into groups of 2 pigs and were inoculated with 1 of 15 A. pleuropneumoniae strains representing all known serovars of A. pleuropneumoniae, or with Actinobacillus suis, or were vaccinated with a bacterin containing A. pleuropneumoniae serovar 1, 3, 5, or 7. Serum samples collected at the day of inoculation or vaccination and 7, 14, 21, and 28 days later were used to compare the assays. On samples from experimentally infected pigs, the dual-plate CF assay, quad-plate ELISA-1, single-plate ELISA-2, and single-plate ELISA-3 had sensitivities of 0.46, 0.74, 0.13, and 0.13 and specificities of 0.90, 1.0, 1.0, and 1.0, respectively. Vaccinated pigs were identified only by the dual-plate CF assay and the quad-plate ELISA-1. In addition, 90 serum samples with unknown A. pleuropneumoniae exposure collected under field conditions were tested with all assays. The agreement of the 4 assays on field samples was slight to fair. While several assays are available for demonstration of A. pleuropneumoniae exposure, differences in assay targets complicate test choices. Decisions on which assay or combination of assays to use depend on the specific reasons for running the assays.
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Krämer, Michael; Heese, Peter; Banger, Markus; Madea, Burkhard; Hess, Cornelius
2018-06-01
Due to a lack of reference blood concentrations in the literature, the forensic evaluation of prothipendyl findings in blood samples is difficult. Interpretations with regard to the assessment of blood concentrations as well as an estimation of the ingested prothipendyl amounts were often vague. To describe a concentration range in clinical samples, prothipendyl and prothipendyl sulfoxide concentrations were determined in serum samples of 50 psychiatric patients receiving 40 mg, 80 mg, or 160 mg doses of prothipendyl. The analyses of prothipendyl and prothipendyl sulfoxide were carried out using validated methods of high performance liquid chromatography coupled to triple quadrupole mass spectrometry (LC-QQQ-MS), respectively. 40 mg doses caused average prothipendyl serum concentrations of 18.0 ng/mL (1 hour after intake) and 7.9 ng/mL (10.5 hours after intake), while 80 mg doses caused averages of 42.6 ng/mL and 15.2 ng/mL at the mentioned times of sampling. Irrespective of the given dose, prothipendyl concentrations below 30 ng/mL were observed in 80% of the patient samples taken 1 hour after ingestion as well as in 90% of the samples collected 10.5 hours after administration. Serum concentrations of the Phase I metabolite prothipendyl sulfoxide averaged 4.3 ng/mL (1 hour after intake) and 3.6 ng/mL (10.5 hours after intake). Possible drug-drug interactions regarding absorption and metabolism of prothipendyl are discussed. Results of the herein presented study are useful for the interpretation of analytical prothipendyl findings in forensic toxicology. The utility of the described concentration range is demonstrated by discussing two death cases involving prothipendyl findings. Copyright © 2017 John Wiley & Sons, Ltd.
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Halpern, Micah D.; Molins, Claudia R.; Schriefer, Martin
2014-01-01
A serology-based tiered approach has, to date, provided the most effective means of laboratory confirmation of clinically suspected cases of Lyme disease, but it lacks sensitivity in the early stages of disease and is often dependent on subjectively scored immunoblots. We recently demonstrated the use of immuno-PCR (iPCR) for detecting Borrelia burgdorferi antibodies in patient serum samples that were positive for Lyme disease. To better understand the performance of the Lyme disease iPCR assay, the repeatability and variability of the background of the assay across samples from a healthy population (n = 36) were analyzed. Both of these parameters were found to have coefficients of variation of <3%. Using eight antigen-specific iPCR assays and positive call thresholds established for each assay, iPCR IgM and/or IgG diagnosis from Lyme disease patient serum samples (n = 12) demonstrated a strong correlation with that of 2-tier testing. Furthermore, a simplified iPCR approach using a single hybrid antigen and detecting only IgG antibodies confirmed the 2-tier diagnosis in the Lyme disease patient serum samples (n = 12). Validation of the hybrid antigen IgG iPCR assay using a blinded panel of Lyme disease and non-Lyme disease patient serum samples (n = 92) resulted in a sensitivity of 69% (95% confidence interval [CI], 50% to 84%), compared to that of the 2-tier analysis at 59% (95% CI, 41% to 76%), and a specificity of 98% (95% CI, 91% to 100%) compared to that of the 2-tier analysis at 97% (95% CI, 88% to 100%). A single-tier hybrid antigen iPCR assay has the potential to be an improved method for detecting host-generated antibodies against B. burgdorferi. PMID:24899074
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Alizadeh, Naader; Mohammadi, Abdorreza; Tabrizchi, Mahmoud
2008-03-07
A simple, rapid and highly sensitive method for simultaneous analysis of methamphetamine (MA) and 3,4-methylenedioxy methamphetamine (MDMA) in human serum was developed using the solid-phase microextraction (SPME) combined with ion mobility spectrometry (IMS). A dodecylsulfate-doped polypyrrole (PPy-DS) was applied as a new fiber for SPME. Electrochemically polymerized PPy is formed on the surface of a platinum wire and will contain charge-compensating anion (dodecylsulfate) incorporated during synthesis using cyclic voltammetry (CV) technique. The extraction properties of the fiber to MA and MDMA were examined, using a headspace-SPME (HS-SPME) device and thermal desorption in injection port of IMS. The results show that PPy-DS as a SPME fiber coating is suitable for the successful extraction of these compounds. This method is suitable for the identification and determination of MAs, is not time-consuming, requires small quantities of sample and does not require any derivatization. Parameters like pH, extraction time, ionic strength, and temperature of the sample were studied and optimized to obtain the best extraction results. The HS-SPME-IMS method provided good repeatability (RSDs<7.8 %) for spiked serum samples. The calibration graphs were linear in the range of 20-4000 ng ml(-1) (R(2)>0.99) and detection limits for MDMA and MA were 5 and 8 ng ml(-1), respectively. HS-SPME-IMS of non-spiked serum sample provided a spectrum without any peak from the matrix, supporting an effective sample clean-up. Finally, the proposed method was applied for analysis one of the ecstasy tablet.
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Paz, Gustavo F; Rugani, Jeronimo M N; Marcelino, Andreza P; Gontijo, Célia M F
2018-06-01
The aim of this study was to evaluate the relationship between naturally occurring Leishmania spp. infections in dogs (Canis familiaris) and the practical implications of the use of serological and molecular methods to confirm diagnoses. The study population consisted of 96 domestic dogs in southeastern Brazil. Serum samples were tested for the presence of anti-Leishmania immunoglobulin G (IgG) antibodies using four commercial canine visceral leishmaniasis kits. Dogs confirmed positive by immunofluorescence antibody test (IFAT) were culled and samples from mesenteric lymph nodes, spleen border, bone marrow and ear skin were taken and submitted to DNA extraction. PCR reactions were performed using primers that amplify a 300-350 bp fragment of the Leishmania ribosomal internal transcribed spacer 1 (ITS1) region. The ITS1 amplified products were analyzed by PCR-RFLP using Hae III restriction endonuclease. To confirm the Leishmania species detected by PCR, each purified sample was sequenced in duplicate. Of the 96 serum samples submitted to serological assays, 8 (8.3%) tested positive for Leishmania by IFAT, 4 (4.1%) by ELISA, 2 (2.1%) by rK39 RDT and 7 (7.3%) by DPP. Four of these infected dogs (50%) were found to be infected only by Leishmania braziliensis or Leishmania amazonensis, and their serum samples tested positive by IFAT and DPP. These findings demonstrate for the first time that cross-reactivity of L. braziliensis and L. amazonensis infection in dogs can be found using the DPP serum test. This is the first record of Leishmania (Leishmania) amazonensis confirmed by a specific molecular marker in dogs (Canis familiaris) from Belo Horizonte, Brazil. Copyright © 2018 Elsevier B.V. All rights reserved.
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Frías-De-León, María Guadalupe; Ramírez-Bárcenas, José Antonio; Rodríguez-Arellanes, Gabriela; Velasco-Castrejón, Oscar; Taylor, Maria Lucia; Reyes-Montes, María Del Rocío
2017-03-01
Histoplasmosis is considered the most important systemic mycosis in Mexico, and its diagnosis requires fast and reliable methodologies. The present study evaluated the usefulness of PCR using Hcp100 and 1281-1283 (220) molecular markers in detecting Histoplasma capsulatum in occupational and recreational outbreaks. Seven clinical serum samples of infected individuals from three different histoplasmosis outbreaks were processed by enzyme-linked immunosorbent assay (ELISA) to titre anti-H. capsulatum antibodies and to extract DNA. Fourteen environmental samples were also processed for H. capsulatum isolation and DNA extraction. Both clinical and environmental DNA samples were analysed by PCR with Hcp100 and 1281-1283 (220) markers. Antibodies to H. capsulatum were detected by ELISA in all serum samples using specific antigens, and in six of these samples, the PCR products of both molecular markers were amplified. Four environmental samples amplified one of the two markers, but only one sample amplified both markers and an isolate of H. capsulatum was cultured from this sample. All PCR products were sequenced, and the sequences for each marker were analysed using the Basic Local Alignment Search Tool (BLASTn), which revealed 95-98 and 98-100 % similarities with the reference sequences deposited in the GenBank for Hcp100 and 1281-1283 (220) , respectively. Both molecular markers proved to be useful in studying histoplasmosis outbreaks because they are matched for pathogen detection in either clinical or environmental samples.
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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
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Evidence to Support a Contribution of Polyreactive Antibodies to HLA Serum Reactivity.
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.
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Massive glutamine cyclization to pyroglutamic acid in human serum discovered using NMR spectroscopy.
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.
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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.
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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.
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Transfer of perfluoroalkyl substances from mother to fetus in a Spanish birth cohort.
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.
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Stability of selected serum proteins after long-term storage in the Janus Serum Bank.
Gislefoss, Randi E; Grimsrud, Tom K; Mørkrid, Lars
2009-01-01
Human serum from biobanks is frequently used in prospective epidemiological studies. Long-term storage may modify its composition. A better understanding of the stability of the serum components may improve the interpretation of future studies. The concentrations of selected proteins; immunoglobulins, carrier proteins and enzymes in samples stored at -25 degrees C for 25 years and 2 years were compared with 1-month-old samples. For each length of storage time, 130 specimens were randomly selected from apparently healthy male blood donors aged 40-49 years. We examined the distribution of values, compared dispersion and localization of central tendency, and established reference intervals for each component. The study demonstrated non-significant or numerically small group differences in the concentrations of albumin, aspartate amino transferase, cystatin C, immunoglobulin E, immunoglobulin G, and sex hormone binding globulin. Mean values between fresh and 25-year-old samples suggested larger differences during storage for alanine amino transferase (-73.4%), creatinine kinase (-96.1%), insulin C-peptide (-98.7%), ferritin (-18.5%) and transferrin (+8.2%). The findings showed that long-term storage can introduce a considerable bias for vulnerable components.
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Expression of microRNAs of C19MC in Different Histological Types of Testicular Germ Cell Tumour.
Flor, Inga; Spiekermann, Meike; Löning, Thomas; Dieckmann, Klaus-Peter; Belge, Gazanfer; Bullerdiek, Jörn
2016-01-01
Testicular germ cell tumours (TGCTs) are the most common tumours in men aged from 20 to 40 years, with a steadily increasing incidence. This study aimed to characterize the expression of the miRNA cluster C19MC in TGCT and to evaluate the suitability of a C19MC miRNA as a serum biomarker. By quantitative reverse transcription PCR, we measured the expression of miR-517a-3p, miR-519a-3p, and miR-519c 3p in tissue samples of 25 TGCTs and the level of miR-517a-3p in serum samples obtained pre- and postoperatively from the same patients. We detected a significantly higher expression of C19MC miRNAs in non-seminomas than in seminomas and in clinical stages 2 and 3 than in stage 1 in both tissue and serum samples. miRNAs of C19MC are overexpressed in more aggressive types of TGCT, suggesting they contribute to malignancy. Furthermore, they might serve as serum biomarkers for these types of TGCT. Copyright© 2016, International Institute of Anticancer Research (Dr. John G. Delinasios), All rights reserved.
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A macro-enzyme cause of an isolated increase of alkaline phosphatase.
Cervinski, Mark A; Lee, Hong Kee; Martin, Isabella W; Gavrilov, Dimitar K
2015-02-02
Macroenzyme complexes of serum enzymes and antibody can increase the circulating enzymatic activity and may lead to unnecessary additional testing and procedures. Laboratory physicians and scientists need to be aware of techniques to identify macroenzyme complexes when suspected. To investigate the possibility of a macro-alkaline phosphatase in the serum of a 74 year old male with persistently increased alkaline phosphatase we coupled a protein A/G agarose affinity chromatography technique with isoenzyme electrophoresis to look for the presence of macro-alkaline phosphatase. The majority of the alkaline phosphatase activity in the patient's serum sample was bound to the column and only a minor fraction (25%) of alkaline phosphatase activity was present in the column flow-through. The alkaline phosphatase activity was also found to co-elute with the immunoglobulins in the patient sample. The alkaline phosphatase activity in a control serum sample concurrently treated in the same manner did not bind to the column and was found in the column flow-through. The use of protein A/G agarose affinity chromatography is a rapid and simple method that can be applied to the investigation of other macro-enzyme complexes. Copyright © 2014 Elsevier B.V. All rights reserved.
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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.
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NASA Technical Reports Server (NTRS)
Whiitson, Peggy A. (Inventor); Clift, Vaughan L. (Inventor)
1999-01-01
The present invention provides a method and apparatus for separating a blood sample having a volume of up to about 20 milliliters into cellular and acellular fractions. The apparatus includes a housing divided by a fibrous filter into a blood sample collection chamber having a volume of at least about 1 milliliter and a serum sample collection chamber. The fibrous filter has a pore size of less than about 3 microns, and is coated with a mixture including between about 1-40 wt/vol % mannitol and between about 0.1-15 wt/vol % of plasma fraction protein (or an animal or vegetable equivalent thereof). The coating causes the cellular fraction to be trapped by the small pores, leaving the cellular fraction intact on the fibrous filter while the acellular fraction passes through the filter for collection in unaltered form from the serum sample collection chamber.
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Chakraborty, A K; Karam, A; Mukherjee, P; Barkalita, L; Borah, P; Das, S; Sanjukta, R; Puro, K; Ghatak, S; Shakuntala, I; Sharma, I; Laha, R G; Sen, A
2018-03-01
The present study focused on the detection and genetic characterisation of 5' untranslated region (5'UTR) and E2 gene of classical swine fever virus (CSFV, family Flaviviridae , genus Pestivirus ) from bovine population of the northeastern region of India. A total of 134 cattle serum samples were collected from organised cattle farms and were screened for CSFV antigen with a commercial antigen capture enzyme linked immunosorbent assay (Ag-ELISA) and reverse transcription-polymerase chain reaction (RT-PCR). A total of 10 samples were positive for CSFV antigen by ELISA, while all of them were positive in PCR for 5'UTR region. Full length E2 region of CSFV were successfully amplified from two positive samples and used for subsequent phylogenetic analysis and determination of protein 3D structure which showed similarity with reported CSFV isolate from Assam of sub-genogroup 2.1, with minor variations in protein structure.
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Ivanova, Stefka Kr; Angelova, Svetla G; Stoyanova, Asya P; Georgieva, Irina L; Nikolaeva-Glomb, Lubomira K; Mihneva, Zafira G; Korsun, Neli St
2016-12-01
Inflammatory diseases of the heart (myocarditis, pericarditis) are commonly caused by viruses. Among the human cardiotropic viruses, parvovirus B19, Coxsackie B viruses, and adenoviruses play a leading role. The aim of the present study was to determine the presumptive causative role of parvovirus B19, Coxsackie B viruses, and adenoviruses in the development of myocarditis, pericarditis and dilated cardiomyopathy by demonstrating the presence of specific antiviral antibodies or viral DNA in patients' serum samples. We tested serum samples collected between 2010 and 2014 from 235 patients with myocarditis (n=108), pericarditis (n=79), myopericarditis (n=19), dilated cardiomyopathy (n=7), and fever of unknown origin accompanied by cardiac complaints (n=22). The mean age of patients with the standard deviation was 33 ± 18 years. Serological and molecular methods (ELISA for specific IgM/IgG antibodies to parvovirus B19 and IgM antibodies to Coxsackie B viruses and adenoviruses, and PCR for detection of parvovirus B19 in serum samples, respectively) were used in the study. Of all tested 235 serum samples, in 60 (25.5%) positive results for at least one of the three tested viruses were detected. Forty out of these 235 serum samples (17%) were Coxsackie B virus IgM positive. They were found in 17% (18/108) of the patients with myocarditis, in 15% (12/79) of those with pericarditis, in 16% (3/19) of those with myopericarditis and in 32% (7/22) in those with fever of unknown origin. The 63 Coxsackie B virus IgM negative patient's serum samples were tested by ELISA for presence of adenovirus IgM antibodies. Such were found in 4 patients with pericarditis and in 2 patients with fever of unknown origin. Every IgM negative sample (n=189) for Coxsackie B and adenovirus was further tested by ELISA for parvovirus B19 IgM/IgG antibodies. B19-IgM antibodies were detected in 14 patients (7.4%). The percentages for B19-IgM antibodies was 8% (7/90), 5% (3/63) and 31% (4/13) in the patients affected with myocarditis, pericarditis, and fever of unknown origin, respectively. Protective B19-IgG antibodies were found in 108 (57%) of the samples. A B19-PCR signal was detected in all the patients who were B19-IgM positive, and in only 1 patient with positive B19-IgG result, the latter presenting with dilated cardiomyopathy. The present study shows the involvement of Coxsackie B, parvovirus B19 and adenoviruses in the development of inflammatory diseases of the heart (myocarditis and pericarditis). It is the first ever study in the country that simultaneously analyzes the prevalence of the three major human cardiotropic viruses.
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Serum lipid concentrations in six canid and four ursid species in four zoos.
Crissey, Susan D; Ange, Kimberly D; Slifka, Kerri A; Sadler, William; Kahn, Stephen; Ward, Ann M
2004-03-01
Serum lipid levels were measured in healthy captive wild canids and ursids, and the values were compared with previously published data. Serum lipid levels were evaluated in blood samples collected from eight African wild dogs (Lycaon pictus), three arctic foxes (Alopex lagopus), nine gray wolves (Canis lupus), four maned wolves (Chrysocyon brachyurus), two Mexican wolves (Canis lupus baleiyi), nine red wolves (Canis rufus), two brown bears (Ursus arctos), six polar bears (Ursus maritimus), six spectacled bears (Tremarctos ornatus), and five sun bears (Ursus malayanus). Samples were analyzed for total cholesterol, triacylglycerides, high-density lipoprotein-cholesterol, and low-density lipoprotein-cholesterol. Although the results showed a great variation among species, circulating lipids appeared especially high, sometimes extremely so, in the spectacled bears, polar bears, sun bears, and maned wolves compared with all other species sampled. The study provides a substantial basis for comparing lipid levels in presumed healthy animals and indicates a need for controlled study of the effects of diet on circulating lipid levels.
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Seo, Dongmin; Paek, Sung-Ho; Oh, Sangwoo; Seo, Sungkyu; Paek, Se-Hwan
2016-09-24
The incidence of diabetes is continually increasing, and by 2030, it is expected to have increased by 69% and 20% in underdeveloped and developed countries, respectively. Therefore, glucose sensors are likely to remain in high demand in medical device markets. For the current study, we developed a needle-type bio-layer interference (BLI) sensor that can continuously monitor glucose levels. Using dialysis procedures, we were able to obtain hypoglycemic samples from commercial human serum. These dialysis-derived samples, alongside samples of normal human serum were used to evaluate the utility of the sensor for the detection of the clinical interest range of glucose concentrations (70-200 mg/dL), revealing high system performance for a wide glycemic state range (45-500 mg/dL). Reversibility and reproducibility were also tested over a range of time spans. Combined with existing BLI system technology, this sensor holds great promise for use as a wearable online continuous glucose monitoring system for patients in a hospital setting.
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Quantitative assessment of anthrax vaccine immunogenicity using the dried blood spot matrix.
Schiffer, Jarad M; Maniatis, Panagiotis; Garza, Ilana; Steward-Clark, Evelene; Korman, Lawrence T; Pittman, Phillip R; Mei, Joanne V; Quinn, Conrad P
2013-03-01
The collection, processing and transportation to a testing laboratory of large numbers of clinical samples during an emergency response situation present significant cost and logistical issues. Blood and serum are common clinical samples for diagnosis of disease. Serum preparation requires significant on-site equipment and facilities for immediate processing and cold storage, and significant costs for cold-chain transport to testing facilities. The dried blood spot (DBS) matrix offers an alternative to serum for rapid and efficient sample collection with fewer on-site equipment requirements and considerably lower storage and transport costs. We have developed and validated assay methods for using DBS in the quantitative anti-protective antigen IgG enzyme-linked immunosorbent assay (ELISA), one of the primary assays for assessing immunogenicity of anthrax vaccine and for confirmatory diagnosis of Bacillus anthracis infection in humans. We have also developed and validated high-throughput data analysis software to facilitate data handling for large clinical trials and emergency response. Published by Elsevier Ltd.
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Dengue-yellow fever sera cross-reactivity; challenges for diagnosis.
Houghton-Triviño, Natalia; Montaña, Diana; Castellanos, Jaime
2008-01-01
The Flavivirus genera share epitopes inducing cross-reactive antibodies leading to great difficulty in differentially diagnosing flaviviral infections. This work was aimed at evaluating the complexity of dengue and yellow fever serological differential diagnosis. Dengue antibody capture ELISA and a yellow fever neutralisation test were carried out on 13 serum samples obtained from yellow fever patients, 20 acute serum samples from dengue patients and 19 voluntary serum samples pre- and post-vaccination with YF vaccine. Dengue ELISA revealed IgM reactivity in 46,2 % of yellow fever patients and 42 % of vaccinees. Sixteen out of 20 dengue patients (80 %) had high YF virus neutralisation titres. Such very high cross-reactivity data challenged differential laboratory diagnosis of dengue and yellow fever in areas where both flaviviruses co-circulate. New laboratory strategies are thus needed for improving the tests and providing a specific laboratory diagnosis. Cross-reactivity between Flaviviruses represents a great difficulty for epidemiological surveillance and preventing dengue, both of which demand urgent attention.
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Ito, Hiroaki; Hasegawa, Katsuyuki; Hasegawa, Yuuki; Nishimaki, Tadashi; Hosomichi, Kazuyoshi; Kimura, Satoshi; Ohba, Motoi; Yao, Hiroshi; Onimaru, Manabu; Inoue, Ituro; Inoue, Haruhiro
2015-05-21
Blood tests, which are commonly used for cancer screening, generally have low sensitivity. Here, we developed a novel rapid and simple method to generate silver nanoscale hexagonal columns (NHCs) for use in surface-enhanced Raman scattering (SERS). We reported that the intensity of SERS spectra of clinical serum samples obtained from gastrointestinal cancer patients is was significantly higher than that of SERS spectra of clinical serum samples obtained from non-cancer patients. We estimated the combined constituents on silver NHCs by using a field emission-type scanning electron microscope, Raman microscopes, and a 3D laser scanning confocal microscope. We obtained the Raman scattering spectra of samples of physically fractured cells and clinical serum. No spectra were obtained for chemically lysed cultured cells and DNA, RNA, and protein extracted from cultured cells. We believe that our method, which uses SERS with silver NHCs to detect circulating nucleosomes bound by methylated cell-free DNA, may be successfully implemented in blood tests for cancer screening.
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Willard, M D; Simpson, R B; Fossum, T W; Cohen, N D; Delles, E K; Kolp, D L; Carey, D P; Reinhart, G A
1994-04-15
Sixteen German Shepherd Dogs were found, via quantitative microbial culture of intestinal fluid samples, to have small intestinal bacterial overgrowth (IBO) over an 11-month period. All dogs were deficient in serum IgA. Consistent clinical signs suggestive of an alimentary tract disorder were not observed. Serum cobalamin determinations were not helpful in detecting IBO. Serum folate concentrations had variable sensitivity and specificity for detecting dogs from which we could culture > or = 1 x 10(5) bacterial/ml from intestinal fluid samples in the nonfed state. Histologic and intestinal mucosal cytologic examinations were not useful in detecting IBO. Substantial within-dog and between-dog variation was found in the numbers and species of bacteria in the intestines. The difficulty in diagnosing IBO, the variability in organisms found in individual dogs on repeated sampling, the likelihood that intestinal fluid microbial cultures failed to diagnose IBO in some dogs, and the potential of IBO to be clinically inapparent were the most important findings in this study.
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Gu, Jianli; Li, Jitian; Huang, Manyu; Zhang, Zhiyong; Li, Dongsheng; Song, Guoying; Ding, Xingpo; Li, Wuyin
2014-01-01
Osteosarcoma (OS) is the most common malignant bone tumor. To identify OS-related specific proteins for early diagnosis of OS, a novel approach, surface-enhanced laser desorption/ionization-time-of-flight mass spectrometry (SELDI-TOF-MS) to serum samples from 25 OS patients, 16 osteochondroma, and 26 age-matched normal human volunteers as controls, was performed. Two proteins showed a significantly different expression in OS serum samples from control groups. Proteomic profiles and external leave-one-out cross-validation analysis showed that the correct rate of allocation, the sensitivity, and the specificity of diagnosis were 100%. These two proteins were further identified by searching the EPO-KB database, and one of the proteins identified as Serine rich region profile is involved in various cellular signaling cascades and tumor genesis. The presence of these two proteins in OS patients but absence from premalignant and normal human controls implied that they can be potential biomarkers for early diagnosis of OS.
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Armstrong, Nicholas; Richez, Magali; Raoult, Didier; Chabriere, Eric
2017-08-15
A fast UHPLC-UV method was developed for the simultaneous analysis of Hydroxychloroquine, Minocycline and Doxycycline drugs from 100μL of human serum samples. Serum samples were extracted by liquid-liquid extraction and injected into a phenyl hexyl reverse phase column. Compounds were separated using a mobile phase linear gradient and monitored by UV detection at 343nm. Chloroquine and Oxytetracycline were used as internal standards. Lower and upper limits of quantifications, as well as the other levels of calibration, were validated with acceptable accuracy (<15% deviation) and precision (<15% coefficient of variation) according to the European Medicines Agency guidelines. This new method enables cost and time reduction and was considered suitable for the clinical laboratory. It is the first published assay for the therapeutic drug monitoring of patients diagnosed with Q fever or Whipple's disease. Copyright © 2017 Elsevier B.V. All rights reserved.
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Kitchen, A D; Newham, J A
2011-05-01
Whilst some of the assays used for serological screening of post-mortem blood samples from deceased tissue donors in some countries have been specifically validated by the manufacturer for this purpose, a significant number of those currently in use globally have not. Although specificity has previously been considered a problem in the screening of such samples, we believe that ensuring sensitivity is more important. The aim of this study was to validate a broader range of assays for the screening of post-mortem blood samples from deceased tissue donors. Six microplate immunoassays currently in use within National Health Service Blood and Transplant (NHSBT) for the screening of blood, tissue and stem cell donations were included. Representative samples from confirmed positive donors were titrated in screen negative post-mortem samples in parallel with normal pooled negative serum to determine if there was any inhibition with the post-mortem samples. There were no significant differences seen (P < 0.005) between the dilution curves obtained for the positive samples diluted in post-mortem samples and normal pooled sera. Although small numbers of samples were studied, it can be surmised that the post-mortem blood samples from deceased tissue donors, collected according to United Kingdom guidelines, are a suitable substrate for the assays evaluated. No diminution of reactivity was seen when dilution with sera from deceased donors was compared to dilution using pooled serum from live donors. In the absence of genuine low titre positive post-mortem samples, the use of samples spiked with various levels of target material provides a means of qualifying serological screening assays used by NHSBT for the screening of post-mortem blood samples from deceased tissue donors.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Nydegger, U.E.; Corvetta, A.; Spaeth, P.J.
1983-01-01
During solubilization of immune complexes C3b becomes fixed to the immunoglobulin part and serves as a receptor for the alternative complement pathway control protein H. The H-C3b immune complex interaction can be made detectable using 4% polyethyleneglycol to separate free from bound /sup 125/I-H. Tetanus toxoid (Te)/anti-Te complexes kept soluble with fresh serum and containing 125 IU of specific antibody bound 18% of /sup 125/I-H; when fresh serum was chelated with 10 mM EDTA, /sup 125/I-H binding was only 5%. On sucrose density gradients, the H-binding material sedimented in the range of 12 to 30 S. In 36 serum samplesmore » from rheumatoid arthritis (RA) patients and in 12 serum samples from patients with systemic lupus erythematosus (SLE), /sup 125/I-H binding was significantly elevated to 9.5 +/- 4.7% (mean +/- 1 SD) and 13.3 +/- 5.6%, respectively, while /sup 125/I-H binding by 36 normal human sera was 4 +/- 2%. RA samples (17/36, 47%) and SLE samples (9/12, 75%) had H-binding values increased by more than 2 SD above the normal mean. The serum samples were also assessed for conglutinin- and C1q-binding activities; a significant correlation between H and C1q binding was observed (P less than 0.001); there was no correlation between H and conglutinin binding. Although binding to immune complexes through its interaction with C3b, H clearly detects a population of complexes other than conglutinin, thus expanding the possibilities of further characterizing pathological complexes.« less
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Estrogen levels in serum and urine of vegetarian and omnivore premenopausal women
Harmon, Brook; Morimoto, Yukiko; Beckford, Fanchon; Franke, Adrian A.; Stanczyk, Frank Z.; Maskarinec, Gertraud
2014-01-01
Objective Based on the hypothesis that high-meat diets may increase breast cancer risk through hormonal pathways, this analysis compared estrogens in serum and urine by meat-eating status. Design Intervention with repeated measures. Setting Two randomized soy trials (BEAN1 and BEAN2) among premenopausal healthy women. Subjects BEAN1 participants completed 7 unannounced 24-hour dietary recalls and donated 5 blood and urine samples over 2 years. BEAN2 women provided 7 recalls and 3 samples over 13 months. Serum samples were analyzed for estrone (E1) and estradiol (E2) using radioimmunoassays. Nine estrogen metabolites were measured in urine by liquid chromatography mass spectrometry. Semi-vegetarians included women who reported <30 g/day of red meat, poultry, and fish and pescatarians who consumed <20 g/day of meat/poultry but >10 g/day of fish. All other women were classified as non-vegetarians. We applied mixed models to compute least-square means by vegetarian status adjusted for potential confounders. Results The mean age of the 272 participants was 41.9±4.5 years. Serum E1 (85 vs 100 pg/mL, p=0.04) and E2 (140 vs 154 pg/mL, p=0.04) levels were lower in the 37 semi-vegetarians than in the 235 non-vegetarians. The sum of the 9 urinary estrogen metabolites (183 vs 200 pmol/mg creatinine, p=0.27) and the proportions of individual estrogens and pathways did not differ by meat-eating status. Restricting the models to the samples collected during the luteal phase strengthened the associations. Conclusions Given the limitations of this study, the lower levels of serum estrogens in semi-vegetarians than non-vegetarians need confirmation in larger populations. PMID:24050121
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Kania, Dramane; Truong, Tam Nguyen; Montoya, Ana; Nagot, Nicolas; Van de Perre, Philippe; Tuaillon, Edouard
2015-01-01
Point-of-care testing and diagnosis of HIV acute infections play important roles in preventing transmission, but HIV rapid diagnosis tests have poor capacity to detect early infections. Filter paper can be used for capillary blood collection and HIV testing using 4th generation immunoassays. Antigen/antibody combined immunoassays were evaluated for their capacity to identify early HIV infections using filter paper in comparison with rapid test. Thirty nine serum samples collected from HIV seroconverters were spotted onto filter paper and tested by the Roche Elecsys(®) HIV Combi PT test and the DiaSorin Liaison XL Murex HIV Ab/Ag assay. Fourth generation immunoassays identified 34 out of 39 HIV early infections using dried serum spot, whereas the Determine™ HIV-1/2 rapid test detected 24 out of 39 HIV positive serum (87.2% vs 61.5% respectively, p = 0.009). p24 antigen was detected by the Liaison XL in 19 dried serum samples (48.7%). In the group characterized by a negative western blot, 7 out of 8 (87.5%) and 6 out of 8 (75.0%) samples were found positive for HIV using the Elecsys and the Liaison XL, respectively. None of these eight samples classified in this group of early acute infections were found positive by the rapid test. Fourth generation Ag/Ab immunoassays performed on dried serum spot had good performance for HIV testing during the early phases of HIV infection. This method may be useful to detect HIV early infections in hard-to-reach populations and individuals living in remote areas before rapid tests become positive. Copyright © 2014 Elsevier B.V. All rights reserved.
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Lam, Virginie; Albrecht, Matthew A; Takechi, Ryusuke; Prasopsang, Prachya; Lee, Ya Ping; Foster, Jonathan K; Mamo, John C L
2016-06-01
There is increasing evidence supporting an association of higher serum vitamin D concentration with better cognitive performance in older individuals. However, to date, consideration of the putative association between vitamin D and cognition has been based principally on studies investigating clinical participant samples manifesting vitamin D deficiency, particularly in older people. Moreover, relationships between vitamin D and cognition are typically not considered in the context of counter-regulatory calcium-modulating hormones or calcium homeostasis. Serum vitamin D/bioactive (ionised) calcium/parathyroid hormone homeostasis was considered in the context of cognitive performance in healthy, middle-aged and older individuals. A cross-sectional sample of 179 participants between the ages of 47-84 years was recruited for this study (114 females, 65 males). Participants provided fasting blood samples for analysis of serum 25-hydroxyvitamin D levels, ionised calcium (iCa) and parathyroid hormone (PTH) and completed cognitive measures of verbal episodic learning and memory. Serum 25-hydroxyvitamin D concentrations were negatively associated (with and without covariates of age, gender, depression and NART scores, iCa, and PTH) with measures of verbal episodic learning and memory, in particular with trial 5 of the Rey Auditory Verbal Learning Test (RAVLT) and long-delay free recall on the RAVLT. Overall, the findings from this study suggest an association between higher vitamin D status and poorer performance on verbal episodic memory in middle-aged and older individuals with normal vitamin D-calcium-PTH homeostasis. Despite requiring replication in other participant samples, this is a potentially important finding as it indicates that it may not be beneficial from a cognitive perspective to provide vitamin D supplements in individuals with already adequate vitamin D status.
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Espina, Juan Gómez; Montes-Bayón, Maria; Blanco-González, Elisa; Sanz-Medel, Alfredo
2015-10-01
Analytical methods allowing sensitive determination of reduced homocysteine (rHcy), one of the so-called biothiols, in human serum is a topic of growing interest due to its close relation to several human disorders, mainly cardiovascular diseases. Although most widely used analytical strategies to determine total Hcy involve derivatization by means of fluorescent labels, this work proposes the use of ebselen, a Se-containing labelling agent to derivatize the reactive sulfhydryl group of the Hcy molecule in its "free" reduced form, which is more likely to play different roles in disease pathogenesis. Optimization of the derivatization and separation conditions by high-performance liquid chromatography (HPLC) to isolate the excess of derivatizing reagent is carried out here using UV/VIS detection. Further, the study of the Se labelling reaction by electrospray ionization tandem mass spectrometry (ESI-MS/MS) provides a stoichiometry of the derivative of 1:1. The main advantage of using ebselen as a labelling agent is the presence of the Se atom in the molecule that allows the use of inductively coupled plasma mass spectrometry (ICP-MS) as a sensitive and selective Se detector. The coupling of HPLC with ICP-MS provided excellent features for the determination of Se-derivatized rHcy (detection limit of 9.6 nM) in real samples. Quantification was accomplished by using post-column isotope dilution (ID) of Se in serum samples, after precipitation of the main serum proteins. Quantitative results for "free" rHcy turned out to be around 0.18-0.22 μM in serum samples from healthy individuals that could be directly analyzed without sample preconcentration.
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Nguyen, Han Christine Ngoc; Xie, Wanling; Yang, Ming; Hsieh, Chen-Lin; Drouin, Sarah; Lee, Gwo-Shu Mary; Kantoff, Philip W
2013-03-01
Recent studies show that microRNAs (miRNAs), small non-coding RNAs that negatively regulate gene expression, may have potential for monitoring cancer status. We investigated circulating miRNAs in prostate cancer that may be associated with the progression of hormone-sensitive primary tumors to metastatic castration resistant prostate cancer (CRPC) after androgen deprivation therapy. Using genome-wide expression profiling by TaqMan Human MicroRNA Arrays (Applied Biosystems) and/or quantitative real-time polymerase chain reaction, we compared the expression levels of miRNAs in serum samples from 28 patients of low-risk localized disease, 30 of high-risk localized disease and 26 of metastatic CRPC. We demonstrated that serum samples from patients of low risk, localized prostate cancer and metastatic CRPC patients exhibit distinct circulating miRNA signatures. MiR-375, miR-378*, and miR-141 were significantly over-expressed in serum from CRPC patients compared with serum from low-risk localized patients, while miR-409-3p was significantly under-expressed. In prostate primary tumor samples, miR-375 and miR-141 also had significantly higher expression levels compared with those in normal prostate tissue. Circulating miRNAs, particularly miR-375, miR-141, miR-378*, and miR-409-3p, are differentially expressed in serum samples from prostate cancer patients. In the search for improved minimally invasive methods to follow cancer pathogenesis, the correlation of disease status with the expression patterns of circulating miRNAs may indicate the potential importance of circulating miRNAs as prognostic markers for prostate cancer progression. Copyright © 2012 Wiley Periodicals, Inc.
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Rafiei Tabatabaei, Sedigheh; Esteghamati, Abdoul-Reza; Shiva, Farideh; Fallah, Fatemeh; Radmanesh, Raheleh; Abdinia, Babak; Shamshiri, Ahmad Reza; Khairkhah, Masoumeh; Shekari Ebrahimabad, Hamideh; Karimi, Abdollah
2013-01-01
In Iran, the measles, mumps and rubella vaccine (MMR) is administered in a two-dose protocol where the first dose is scheduled at 12 months of age. This study aims to determine the efficacy of the MMR vaccine by testing IgM and IgG antibody levels 4 - 7 weeks after primary vaccination. A single group cohort study was performed on healthy children, 12 - 15 months of age, who were vaccinated at health centers affiliated with Shahid Beheshti University of Medical Sciences in Tehran, from January to April 2009. Children with negative vaccination and/or clinical history for measles, mumps or rubella were administered the first dose of the MMR live attenuated vaccine. IgG and IgM antibodies were checked by enzyme linked immunoassay (ELISA) in serum samples 4 - 7 weeks after vaccination. A child was considered seropositive if antibody levels were higher than the assay cut-off level set by the ELISA kit. Samples from 240 children were checked for antibodies against measles and rubella. Measles serum IgM level was positive in 71.7% of samples and IgG in 75.8%. The rubella serum IgM level was positive in 71.7% of children and IgG in 73.8%. From 190 blood samples that were checked for mumps antibodies, serum IgM was positive in 68.9% and IgG in 95.3%. No significant relationship was found between seropositivity and age or gender. IgG and IgM antibody levels were below the assay cut-off levels against measles and rubella in approximately one-fourth of the children following primary MMR vaccination. A second dose was necessary to raise the level of protection against measles and rubella.
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Park, Jaehyun; Jeon, Byongtae; Kang, Sungki; Oh, Mirae; Kim, Myonghwa; Jang, Seyoung; Park, Pyojam; Kim, Sangwoo; Moon, Sangho
2015-09-01
This study was conducted to investigate changes in blood enzyme parameters and to evaluate the relationship between insulin-like growth factor-1 (IGF-1), antler growth and body weight during the antler growth of sika deer (Cervus nippon). Serum enzyme activity and IGF-1 concentrations were measured in blood samples collected from the jugular and femoral veins at regular intervals during the antler growth period. Blood samples were taken in the morning from fasted stags (n = 12) which were healthy and showed no clinical signs of disease. Alfalfa was available ad libitum and concentrates were given at 1% of body weight to all stags. The experimental diet was provided at 9 am with water available at all times. There were no significant differences in alkaline phosphatase, aspartate aminotransferase, and alanine aminotransferase during antler growth, but alkaline phosphatase concentrations increased with antler growth progression, and the highest alkaline phosphatase concentration was obtained 55 days after antler casting. Serum IGF-1 concentrations measured from blood samples taken from the jugular vein during antler growth, determined that levels of IGF-1 was associated with body weight and antler growth patterns. Serum IGF-1 concentrations were higher at the antler cutting date than other sampling dates. Antler length increased significantly during antler growth (p<0.001), and there was a similar trend to between right and left beams. Body weight increased with antler growth but was not significant. Consequently it appeared that serum alkaline phosphatase concentration was related to antler growth and both antler growth and body weight were associated positively with IGF-1 concentrations during antler growth.
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Diesel, Alison; DeBoer, Douglas J
2011-02-01
Feline and canine atopic dermatitis are thought to have a similar immunopathogenesis. As with dogs, detection of allergen-specific IgE in cat serum merely supports a diagnosis of feline atopy based on compatible history, clinical signs and elimination of other pruritic dermatoses. In this study, a rapid screening immunoassay (Allercept(®) E-Screen 2nd Generation; Heska AG, Fribourg, Switzerland; ES2G) was compared with a complete-panel serum allergen-specific IgE assay (Allercept(®); Heska AG; CP) in healthy cats with no history of skin disease and in atopic cats. The latter had no diagnosis of external parasitism, infection, food hypersensitivity or other skin disease explaining their pruritus, and expressed cutaneous reaction patterns typically associated with feline allergic skin disease (head, neck or pinnal pruritus, miliary dermatitis, self-induced alopecia, eosinophilic granuloma complex). The proportion of cats positive on either the ES2G or the CP assays was not significantly different between the atopic and healthy cat groups. There was, however, strong agreement between the results of the ES2G and CP assay; overall, the two tests were in agreement for 43 of 49 (88%) serum samples. There was also strong agreement when individual allergen groups were evaluated (agreement noted: indoor, 41 of 49 samples; grasses/weeds, 37 of 49 samples; and trees, 41 of 49 samples). These results indicate that although neither test is diagnostic for feline atopic dermatitis, the screening assay is beneficial for predicting the results of a complete-panel serum allergen-specific IgE assay in cats. © 2010 The Authors. Journal compilation © 2010 ESVD and ACVD.
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Borte, Stephan; Janzi, Magdalena; Pan-Hammarström, Qiang; von Döbeln, Ulrika; Nordvall, Lennart; Winiarski, Jacek; Fasth, Anders; Hammarström, Lennart
2012-01-01
There is a need for neonatal screening tools to improve the long-term clinical outcome of patients with primary immunodeficiency diseases (PID). Recently, a PCR-based screening method for both TRECs and KRECs using Guthrie card samples has been developed. However, the applicability of these excision circle assays is limited to patients with severe T or B cell lymphopenia (SCID, XLA and A-T), whereas the most common forms of PID are not detected. Absence of serum IgA is seen in a major fraction of patients with immunological defects. As serum IgA in newborns is considered to be of fetal origin, eluates from routinely collected dried blood spot samples might thus be suitable for identification of children with PID. To assess the applicability of such screening assays, stored Guthrie card samples were obtained from 47 patients with various forms of primary immunodeficiency diseases (SCID, XLA, A-T, HIGM and IgAD), 20 individuals with normal serum IgA levels born to IgA-deficient mothers and 51 matched healthy newborns. Surprisingly, normal serum IgA levels were found in all SCID, XLA, A-T and HIGM patients and, additionally, in all those IgAD patients born to IgA-sufficient mothers. Conversely, no serum IgA was found in any of the 16 IgAD patients born by IgA-deficient mothers. Moreover, half of the IgA-sufficient individuals born by IgA-deficient mothers also lacked IgA at birth whereas no IgA-deficient individuals were found among the controls. IgA in neonatal dried blood samples thus appears to be of both maternal and fetal origin and precludes its use as a reliable marker for neonatal screening of primary immunodeficiency diseases. PMID:22916257
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Borte, Stephan; Janzi, Magdalena; Pan-Hammarström, Qiang; von Döbeln, Ulrika; Nordvall, Lennart; Winiarski, Jacek; Fasth, Anders; Hammarström, Lennart
2012-01-01
There is a need for neonatal screening tools to improve the long-term clinical outcome of patients with primary immunodeficiency diseases (PID). Recently, a PCR-based screening method for both TRECs and KRECs using Guthrie card samples has been developed. However, the applicability of these excision circle assays is limited to patients with severe T or B cell lymphopenia (SCID, XLA and A-T), whereas the most common forms of PID are not detected. Absence of serum IgA is seen in a major fraction of patients with immunological defects. As serum IgA in newborns is considered to be of fetal origin, eluates from routinely collected dried blood spot samples might thus be suitable for identification of children with PID. To assess the applicability of such screening assays, stored Guthrie card samples were obtained from 47 patients with various forms of primary immunodeficiency diseases (SCID, XLA, A-T, HIGM and IgAD), 20 individuals with normal serum IgA levels born to IgA-deficient mothers and 51 matched healthy newborns. Surprisingly, normal serum IgA levels were found in all SCID, XLA, A-T and HIGM patients and, additionally, in all those IgAD patients born to IgA-sufficient mothers. Conversely, no serum IgA was found in any of the 16 IgAD patients born by IgA-deficient mothers. Moreover, half of the IgA-sufficient individuals born by IgA-deficient mothers also lacked IgA at birth whereas no IgA-deficient individuals were found among the controls. IgA in neonatal dried blood samples thus appears to be of both maternal and fetal origin and precludes its use as a reliable marker for neonatal screening of primary immunodeficiency diseases.
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Evaluation of microRNA Stability in Plasma and Serum from Healthy Dogs.
Enelund, Lars; Nielsen, Lise N; Cirera, Susanna
2017-01-01
Early and specific detection of cancer is of great importance for successful treatment of the disease. New biomarkers, such as microRNAs, could improve treatment efficiency and survival ratio. In human medicine, deregulation of microRNA profiles in circulation has shown great potential as a new type of biomarker for cancer diagnostics. There are, however, few studies of circulating microRNAs in dogs. Extracellular circulating microRNAs have shown a high level of stability in human blood and other body fluids. Nevertheless, there are still important issues to be solved before microRNAs can be applied routinely as a clinical tool, one of them being their stability over time in media commonly used for blood sampling. Evaluation of the stability of microRNA levels in plasma and serum from healthy dogs after storage at room temperature for different time points before being processed. The levels of four microRNAs (cfa-let-7a, cfa-miR-16, cfa-miR-23a and cfa-miR-26a) known to be stably expressed from other canine studies, have been measured by quantitative real-time PCR (qPCR). MicroRNA levels were found sufficiently stable for gene profiling in serum- and plasma stored at room temperature for 1 hour but not for samples stored at room temperature for 24 hours. Storage at room temperature of serum and plasma samples intended for microRNA profiling should be kept for a minimum period of time before proceeding with RNA isolation. For the four microRNAs investigated in the present study, we did not find significant differences in microRNA levels between serum and plasma samples from the same time point. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.
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Zhu, Xiao-Dong; Su, Fang; Liang, Zhong-Guo; Li, Ling; Qu, Song; Liang, Xia; Wang, Qi; Liang, Shi-Xiong; Chen, Long
2014-08-01
As diagnosis of nasopharyngeal carcinoma at an early disease stage is important, we attempted to distinguish between patients with nasopharyngeal carcinoma and noncancer controls by using serum protein profiles. Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry and CM10 protein chip were used to detect the serum proteomic patterns of 65 patients with nasopharyngeal carcinoma before radiotherapy and 93 noncancer controls. Proteomic spectra of serum samples from 50 nasopharyngeal carcinoma patients and 60 noncancer controls were used as a training set. The validity of the classification tree was then challenged with a blind test set which included another 15 patients with nasopharyngeal carcinoma and 33 noncancer controls. Biomarker Wizard 3.01 and Biomarker Pattern 5.01 were used in combination to analyze the data and to develop diagnostic models. 21 protein peaks were significantly different between nasopharyngeal carcinoma and controls. 4 mass peaks (M4182, M5343, M5913 and M8702 mass/charge ratio) were chosen automatically to construct a classification tree. The classification tree correctly determined 93.8 % (45/48) of the test samples with 93.3 % (14/15) of the nasopharyngeal carcinoma samples and 93.9 % (31/33) of the noncancer samples. Using a combination of serum protein profiles and Epstein-Barr viral capsid antigen immunoglobulin A antibody tests, the diagnostic sensitivity and specificity were increased to 100 and 97 %, respectively. Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry could correctly distinguish nasopharyngeal carcinoma from noncancer individuals and showed great potential for the development of a screening test for the detection of nasopharyngeal carcinoma.
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Kucherenko, I S; Soldatkin, O O; Lagarde, F; Jaffrezic-Renault, N; Dzyadevych, S V; Soldatkin, A P
2015-11-01
Creatine kinase (CK: adenosine-5-triphosphate-creatine phosphotransferase) is an important enzyme of muscle cells; the presence of a large amount of the enzyme in blood serum is a biomarker of muscular injuries, such as acute myocardial infarction. This work describes a bi-enzyme (glucose oxidase and hexokinase based) biosensor for rapid and convenient determination of CK activity by measuring the rate of ATP production by this enzyme. Simultaneously the biosensor determines glucose concentration in the sample. Platinum disk electrodes were used as amperometric transducers. Glucose oxidase and hexokinase were co-immobilized via cross-linking with BSA by glutaraldehyde and served as a biorecognition element of the biosensor. The biosensor work at different concentrations of CK substrates (ADP and creatine phosphate) was investigated; optimal concentration of ADP was 1mM, and creatine phosphate - 10 mM. The reproducibility of the biosensor responses to glucose, ATP and CK during a day was tested (relative standard deviation of 15 responses to glucose was 2%, to ATP - 6%, to CK - 7-18% depending on concentration of the CK). Total time of CK analysis was 10 min. The measurements of creatine kinase in blood serum samples were carried out (at 20-fold sample dilution). Twentyfold dilution of serum samples was chosen as optimal for CK determination. The biosensor could distinguish healthy and ill people and evaluate the level of CK increase. Thus, the biosensor can be used as a test-system for CK analysis in blood serum or serve as a component of multibiosensors for determination of important blood substances. Determination of activity of other kinases by the developed biosensor is also possible for research purposes. Copyright © 2015 Elsevier B.V. All rights reserved.
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Khaksari, Maryam; Mazzoleni, Lynn R; Ruan, Chunhai; Kennedy, Robert T; Minerick, Adrienne R
2017-04-01
Two separate liquid chromatography (LC)-mass spectrometry (MS) methods were developed for determination and quantification of water-soluble and fat-soluble vitamins in human tear and blood serum samples. The water-soluble vitamin method was originally developed to detect vitamins B 1 , B 2 , B 3 (nicotinamide), B 5 , B 6 (pyridoxine), B 7 , B 9 and B 12 while the fat-soluble vitamin method detected vitamins A, D 3 , 25(OH)D 3, E and K 1 . These methods were then validated with tear and blood serum samples. In this data in brief article, we provide details on the two LC-MS methods development, methods sensitivity, as well as precision and accuracy for determination of vitamins in human tears and blood serum. These methods were then used to determine the vitamin concentrations in infant and parent samples under a clinical study which were reported in "Determination of Water-Soluble and Fat-Soluble Vitamins in Tears and Blood Serum of Infants and Parents by Liquid Chromatography/Mass Spectrometry DOI:10.1016/j.exer.2016.12.007 [1]". This article provides more details on comparison of vitamin concentrations in the samples with the ranges reported in the literature along with the medically accepted normal ranges. The details on concentrations below the limits of detection (LOD) and limits of quantification (LOQ) are also discussed. Vitamin concentrations were also compared and cross-correlated with clinical data and nutritional information. Significant differences and strongly correlated data were reported in [1]. This article provides comprehensive details on the data with slight differences or slight correlations.
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Greenacre, C B; Young, D W; Behrend, E N; Wilson, G H
2001-11-01
To validate a novel high-sensitivity radioimmunoassay (RIA) procedure developed to accurately measure the relatively low serum total thyroxine (T4) concentrations of birds and reptiles and to establish initial reference ranges forT4 concentration in selected species of psittacine birds and snakes. 56 healthy nonmolting adult psittacine birds representing 6 species and 42 captive snakes representing 4 species. A solid-phase RIA designed to measure free T4 concentrations in dialysates of human serum samples was used without dialysis to evaluate total T4 concentration in treated samples obtained from birds and reptiles. Serum T4 binding components were removed to allow assay of undialyzed samples. Assay validation was assessed by determining recovery of expected amounts of T4 in treated samples that were serially diluted or to which T4 was added. Intra- and interassay coefficient of variation (CV) was determined. Mean recovery of T4 added at 4 concentrations ranged from 84.9 to 115.0% and 95.8 to 119.4% in snakes and birds, respectively. Intra- and interassay CV was 3.8 and 11.3%, respectively. Serum total T4 concentrations for 5 species of birds ranged from 2.02 to 768 nmol/L but ranged from 3.17 to 142 nmol/L for blue-fronted Amazon parrots; concentrations ranged from 0.21 to 6.06 nmol/L for the 4 species of snakes. This new RIA method provides a commercially available, accurate, and sensitive method for measurement of the relatively low serum T4 concentrations of birds and snakes. Initial ranges for the species evaluated were established.
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Investigation of Crimean-Congo Hemorrhagic Fever and Hemorrhagic Fever with Renal Syndrome in Greece
1988-10-12
patients with rheumatoid arthritis (20 samples), with macroglobulinemia (3 samples), with pyrexia of unknown origin (41 samples) and from healthy...precomatose or comatose and 10 had renal failure with oliguria or anuria, requiring hemodialysis . The indication for dialysis in these patients was serum...against Hantaan vi- us and Crimean-Congo Hemorrhagic Fever respectively. Two hundred and two blood samples were aken from patients with disease resembling
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DNA Repair and Ethnic Differences in Prostate Cancer Risk
2006-03-01
Georgetown University for processing . Each sample is centrifuged and the blood components are separated into serum, clot, buffy coat, and plasma...within 4 hours of reception. The processed , aliquoted, and bar-coded samples are stored in a repository at GUH at -80oC. The slow growth of prostate...completeness. Daily backups are performed to protect data against accidental destruction or corruption. Blood samples are processed within 24 hours of sample
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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.
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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.
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Individualized correction of insulin measurement in hemolyzed serum samples.
Wu, Zhi-Qi; Lu, Ju; Chen, Huanhuan; Chen, Wensen; Xu, Hua-Guo
2017-06-01
Insulin measurement plays a key role in the investigation of patients with hypoglycemia, subtype classification of diabetes mellitus, insulin resistance, and impaired beta cell function. However, even slight hemolysis can negatively affect insulin measurement due to RBC insulin-degrading enzyme (IDE). Here, we derived and validated an individualized correction equation in an attempt to eliminate the effects of hemolysis on insulin measurement. The effects of hemolysis on insulin measurement were studied by adding lysed self-RBCs to serum. A correction equation was derived, accounting for both percentage and exposure time of hemolysis. The performance of this individualized correction was evaluated in intentionally hemolyzed samples. Insulin concentration decreased with increasing percentage and exposure time of hemolysis. Based on the effects of hemolysis on insulin measurement of 17 donors (baseline insulin concentrations ranged from 156 to 2119 pmol/L), the individualized hemolysis correction equation was derived: INS corr = INS meas /(0.705lgHb plasma /Hb serum - 0.001Time - 0.612). This equation can revert insulin concentrations of the intentionally hemolyzed samples to values that were statistically not different from the corresponding insulin baseline concentrations (p = 0.1564). Hemolysis could lead to a negative interference on insulin measurement; by individualized hemolysis correction equation for insulin measurement, we can correct and report reliable serum insulin results for a wide range of degrees of sample hemolysis. This correction would increase diagnostic accuracy, reduce inappropriate therapeutic decisions, and improve patient satisfaction with care.
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Vaidya, Sunil R; Dvivedi, Garima M; Jadhav, Santoshkumar M
2016-01-01
The reports from the countries where mumps vaccine is given as routine immunization suggest differences in mumps virus neutralizing antibody titres when tested with vaccine and wild type viruses. Such reports are unavailable from countries like India where mumps vaccine is not included in routine immunization. We, therefore, undertook this study to understand the cross-neutralization activity of Indian mumps viruses. By using commercial mumps IgG enzyme immunoassay (EIA) and a rapid focus reduction neutralization test (FRNT), a panel of serum samples was tested. The panel consisted of 14 acute and 14 convalescent serum samples collected during a mumps outbreak and 18 archived serum samples. Two wild types (genotypes C and G) and Leningrad-Zagreb vaccine strain (genotype N) were used for the challenge experiments and FRNT titres were determined and further compared. The HN protein sequence of three mumps viruses was analyzed for the presence of key epitopes. All serum samples effectively neutralized mumps virus wild types and a vaccine strain. However, significantly lower FRNT titres were noted to wild types than to vaccine strain (P<0.05). The comparison between EIA and FRNT results revealed 95.6 per cent agreement. No amino acid changes were seen in the epitopes in the Indian wild type strains. All potential N-linked glycosylation sites were observed in Indian strains. Good cross-neutralization activity was observed for three mumps virus strains, however, higher level of FRNT titres was detected for mumps virus vaccine strain compared to Indian wild type isolates.
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TOISHI, Yuko; TSUNODA, Nobuo; NAGATA, Shun-ichi; KIRISAWA, Rikio; NAGAOKA, Kentaro; WATANABE, Gen; YANAGAWA, Yojiro; KATAGIRI, Seiji; TAYA, Kazuyoshi
2017-01-01
Testosterone (T) concentration is a useful indicator of reproductive function in male animals. However, T concentration is not usually measured in veterinary clinics, partly due to the unavailability of reliable and rapid assays for animal samples. In this study, a rapid chemiluminescent enzyme immunoassay system (CLEIA system) that was developed for the measurement of T concentration in humans use was validated for stallion blood samples. First, serum T concentrations were measured using the CLEIA system and compared with those measured by a fluoroimmunoassay that has been validated for use in stallions. The serum T concentrations measured by the two methods were highly correlated (r = 0.9865, n = 56). Second, to validate the use of whole blood as assay samples, T concentrations in whole blood and in the serum were measured by the CLEIA system. T concentrations in both samples were highly correlated (r = 0.9665, n = 64). Finally, to evaluate the practical value of the CLEIA system in clinical settings, T concentrations were measured in three stallions with reproductive abnormalities after the administration of human chorionic gonadotropin (hCG). Two stallions with small or absent testes in the scrotum showed an increase in T production in response to hCG administration and one stallion with seminoma did not. In conclusion, the CLEIA system was found to be a rapid and reliable tool for measuring T concentrations in stallions and may improve reproductive management in clinical settings and in breeding studs. PMID:29129877
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Avian influenza H9N2 seroprevalence among pig population and pig farm staff in Shandong, China.
Li, Song; Zhou, Yufa; Zhao, Yuxin; Li, Wenbo; Song, Wengang; Miao, Zengmin
2015-03-01
Shandong province of China has a large number of pig farms with the semi-enclosed houses, allowing crowds of wild birds to seek food in the pig houses. As the carriers of avian influenza virus (AIV), these wild birds can easily pass the viruses to the pigs and even the occupational swine-exposed workers. However, thus far, serological investigation concerning H9N2 AIV in pig population and pig farm staff in Shandong is sparse. To better understand the prevalence of H9N2 AIV in pig population and pig farm staff in Shandong, the serum samples of pigs and occupational pig-exposed workers were collected and tested for the antibodies for H9N2 AIV by both hemagglutination inhibition (HI) and micro-neutralization (MN) assays. When using the antibody titers ≥40 as cut-off value, 106 (HI: 106/2176, 4.87%) and 84 (MN: 84/2176, 3.86%) serum samples of pigs were tested positive, respectively; 6 (HI: 6/287, 2.09%) and 4 (MN: 4/287, 1.39%) serum samples of the pig farm staff were positive, respectively; however, serum samples from the control humans were tested negative in both HI and MN assays. These findings revealed that there were H9N2 AIV infections in pig population and pig farm staff in Shandong, China. Therefore, it is of utmost importance to conduct the long-term surveillance of AIV in pig population and the pig farm staff.
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Biological monitoring of welders' exposure to chromium, molybdenum, tungsten and vanadium.
Ellingsen, Dag G; Chashchin, Maxim; Berlinger, Balazs; Fedorov, Vladimir; Chashchin, Valery; Thomassen, Yngvar
2017-05-01
Welders are exposed to a number of metallic elements during work. Bioaccessability, that is important for element uptake, has been little studied. This study addresses bioaccessability and uptake of chromium (Cr), molybdenum (Mo), tungsten (W) and vanadium (V) among welders. Bioaccessability of Cr, Mo, V and W was studied in airborne particulate matter collected by personal sampling of the workroom air among shipyard welders by using the lung lining fluid simulant Hatch solution. Associations between concentrations of Hatch soluble and non-soluble elements (Hatch sol and Hatch non-sol ) and concentrations of the four elements in whole blood, serum, blood cells and urine were studied. Air concentrations of the four elements were low. Only a small fraction of Cr, V and W was Hatch sol , while similar amounts of Mo were Hatch sol and Hatch non-sol . Welders (N=70) had statistically significantly higher concentrations of all four elements in urine and serum when compared to referents (N=74). Highly statistically significant associations were observed between urinary W and Hatch sol W (p<0.001) and serum V and Hatch sol V (p<0.001), in particular when air samples collected the day before collection of biological samples were considered. Associations between Hatch sol elements in air and their biological concentrations were higher than when Hatch non-sol concentrations were considered. Associations were generally higher when air samples collected the day before biological sampling were considered as compared to air samples collected two days before. Copyright © 2017 Elsevier GmbH. All rights reserved.
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Thiazide increases serum calcium in anuric patients: the role of parathyroid hormone.
Vasco, Raquel F V; Reis, Eduardo T; Moyses, Rosa M A; Elias, Rosilene M
2017-12-01
We evaluated the effect of hydrochlorothiazide in a sample of anuric patients on hemodialysis and found an increase in serum calcium, which occurred only in those with parathyroid hormone >300 pg/ml. This finding highlights the extra-renal effect of this diuretic and a possible role of parathyroid hormone in the mechanism. Thiazide diuretics are commonly used in patients with chronic kidney disease to treat hypertension. Their effects on calcium and bone metabolism are not well established, once calciuria may not fully explain levels of calcium and parathyroid hormone (PTH) in this population. A previous study has suggested that thiazides require the presence of PTH as a permissive condition for its renal action. In anuric patients, however, the role of PTH, if any, in the thiazide effect is unknown. To assess thiazide extra renal effect on serum calcium and whether such an effect is reliant on PTH, hydrochlorothiazide (HCTZ) 100 mg was given orally once a day to a sample of 19 anuric patients on hemodialysis for 2 weeks. Laboratories' analyses were obtained in three phases: baseline, after diuretic use, and after a 2-week washout phase. We demonstrated that serum calcium (Ca) increased in ten patients (52.6%) after HCTZ use, returning to previous levels after the washout period. Out of the 19 patients, ten presented PTH ≥ 300 pg/ml, and Ca has increased in eight of them, whereas in the other nine patients with PTH < 300 pg/ml, serum Ca has increased only in two individuals (RR risk of increase Ca 3.9; p = 0.012). HCTZ was capable of increasing serum Ca in a sample of anuric patients on hemodialysis and seems this effect is highly dependent on PTH levels. Caution is required while interpreting this result, as the small sample size might implicate in a finding caused by chance.
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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
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Targeted quantification of low ng/mL level proteins in human serum without immunoaffinity depletion
DOE Office of Scientific and Technical Information (OSTI.GOV)
Shi, Tujin; Sun, Xuefei; Gao, Yuqian
2013-07-05
We recently reported an antibody-free targeted protein quantification strategy, termed high-pressure, high-resolution separations with intelligent selection and multiplexing (PRISM) for achieving significantly enhanced sensitivity using selected reaction monitoring (SRM) mass spectrometry. Integrating PRISM with front-end IgY14 immunoaffinity depletion, sensitive detection of targeted proteins at 50-100 pg/mL levels in human blood plasma/serum was demonstrated. However, immunoaffinity depletion is often associated with undesired losses of target proteins of interest. Herein we report further evaluation of PRISM-SRM quantification of low-abundance serum proteins without immunoaffinity depletion and the multiplexing potential of this technique. Limits of quantification (LOQs) at low ng/mL levels with a medianmore » CV of ~12% were achieved for proteins spiked into human female serum using as little as 2 µL serum. PRISM-SRM provided up to ~1000-fold improvement in the LOQ when compared to conventional SRM measurements. Multiplexing capability of PRISM-SRM was also evaluated by two sets of serum samples with 6 and 21 target peptides spiked at the low attomole/µL levels. The results from SRM measurements for pooled or post-concatenated samples were comparable to those obtained from individual peptide fractions in terms of signal-to-noise ratios and SRM peak area ratios of light to heavy peptides. PRISM-SRM was applied to measure several ng/mL-level endogenous plasma proteins, including prostate-specific antigen, in clinical patient sera where correlation coefficients > 0.99 were observed between the results from PRISM-SRM and ELISA assays. Our results demonstrate that PRISM-SRM can be successfully used for quantification of low-abundance endogenous proteins in highly complex samples. Moderate throughput (50 samples/week) can be achieved by applying the post-concatenation or fraction multiplexing strategies. We anticipate broad applications for targeted PRISM-SRM quantification of low-abundance cellular proteins in systems biology studies as well as candidate biomarkers in biofluids.« less
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Identification of a Hemolysis Threshold That Increases Plasma and Serum Zinc Concentration.
Killilea, David W; Rohner, Fabian; Ghosh, Shibani; Otoo, Gloria E; Smith, Lauren; Siekmann, Jonathan H; King, Janet C
2017-06-01
Background: Plasma or serum zinc concentration (PZC or SZC) is the primary measure of zinc status, but accurate sampling requires controlling for hemolysis to prevent leakage of zinc from erythrocytes. It is not established how much hemolysis can occur without changing PZC/SZC concentrations. Objective: This study determines a guideline for the level of hemolysis that can significantly elevate PZC/SZC. Methods: The effect of hemolysis on PZC/SZC was estimated by using standard hematologic variables and mineral content. The calculated hemolysis threshold was then compared with results from an in vitro study and a population survey. Hemolysis was assessed by hemoglobin and iron concentrations, direct spectrophotometry, and visual assessment of the plasma or serum. Zinc and iron concentrations were determined by inductively coupled plasma spectrometry. Results: A 5% increase in PZC/SZC was calculated to result from the lysis of 1.15% of the erythrocytes in whole blood, corresponding to ∼1 g hemoglobin/L added into the plasma or serum. Similarly, the addition of simulated hemolysate to control plasma in vitro caused a 5% increase in PZC when hemoglobin concentrations reached 1.18 ± 0.10 g/L. In addition, serum samples from a population nutritional survey were scored for hemolysis and analyzed for changes in SZC; samples with hemolysis in the range of 1-2.5 g hemoglobin/L showed an estimated increase in SZC of 6% compared with nonhemolyzed samples. Each approach indicated that a 5% increase in PZC/SZC occurs at ∼1 g hemoglobin/L in plasma or serum. This concentration of hemoglobin can be readily identified directly by chemical hemoglobin assays or indirectly by direct spectrophotometry or matching to a color scale. Conclusions: A threshold of 1 g hemoglobin/L is recommended for PZC/SZC measurements to avoid increases in zinc caused by hemolysis. The use of this threshold may improve zinc assessment for monitoring zinc status and nutritional interventions.
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Identification of a Hemolysis Threshold That Increases Plasma and Serum Zinc Concentration123
Otoo, Gloria E; Smith, Lauren; Siekmann, Jonathan H
2017-01-01
Background: Plasma or serum zinc concentration (PZC or SZC) is the primary measure of zinc status, but accurate sampling requires controlling for hemolysis to prevent leakage of zinc from erythrocytes. It is not established how much hemolysis can occur without changing PZC/SZC concentrations. Objective: This study determines a guideline for the level of hemolysis that can significantly elevate PZC/SZC. Methods: The effect of hemolysis on PZC/SZC was estimated by using standard hematologic variables and mineral content. The calculated hemolysis threshold was then compared with results from an in vitro study and a population survey. Hemolysis was assessed by hemoglobin and iron concentrations, direct spectrophotometry, and visual assessment of the plasma or serum. Zinc and iron concentrations were determined by inductively coupled plasma spectrometry. Results: A 5% increase in PZC/SZC was calculated to result from the lysis of 1.15% of the erythrocytes in whole blood, corresponding to ∼1 g hemoglobin/L added into the plasma or serum. Similarly, the addition of simulated hemolysate to control plasma in vitro caused a 5% increase in PZC when hemoglobin concentrations reached 1.18 ± 0.10 g/L. In addition, serum samples from a population nutritional survey were scored for hemolysis and analyzed for changes in SZC; samples with hemolysis in the range of 1–2.5 g hemoglobin/L showed an estimated increase in SZC of 6% compared with nonhemolyzed samples. Each approach indicated that a 5% increase in PZC/SZC occurs at ∼1 g hemoglobin/L in plasma or serum. This concentration of hemoglobin can be readily identified directly by chemical hemoglobin assays or indirectly by direct spectrophotometry or matching to a color scale. Conclusions: A threshold of 1 g hemoglobin/L is recommended for PZC/SZC measurements to avoid increases in zinc caused by hemolysis. The use of this threshold may improve zinc assessment for monitoring zinc status and nutritional interventions. PMID:28490675