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Sample records for dried blood specimens

  1. Field Study of Dried Blood Spot Specimens for HIV-1 Drug Resistance Genotyping

    PubMed Central

    Parry, C. M.; Diallo, K.; Mwebaza, S.; Batamwita, R.; DeVos, J.; Bbosa, N.; Lyagoba, F.; Magambo, B.; Jordan, M. R.; Downing, R.; Zhang, G.; Kaleebu, P.; Bertagnolio, S.

    2014-01-01

    Dried blood spots (DBS) are an alternative specimen type for HIV drug resistance genotyping in resource-limited settings. Data relating to the impact of DBS storage and shipment conditions on genotyping efficiency under field conditions are limited. We compared the genotyping efficiencies and resistance profiles of DBS stored and shipped at different temperatures to those of plasma specimens collected in parallel from patients receiving antiretroviral therapy in Uganda. Plasma and four DBS cards from anti-coagulated venous blood and a fifth card from finger-prick blood were prepared from 103 HIV patients with a median viral load (VL) of 57,062 copies/ml (range, 1,081 to 2,964,191). DBS were stored at ambient temperature for 2 or 4 weeks or frozen at −80°C and shipped from Uganda to the United States at ambient temperature or frozen on dry ice for genotyping using a broadly sensitive in-house method. Plasma (97.1%) and DBS (98.1%) stored and shipped frozen had similar genotyping efficiencies. DBS stored frozen (97.1%) or at ambient temperature for 2 weeks (93.2%) and shipped at ambient temperature also had similar genotyping efficiencies. Genotyping efficiency was reduced for DBS stored at ambient temperature for 4 weeks (89.3%, P = 0.03) or prepared from finger-prick blood and stored at ambient temperature for 2 weeks (77.7%, P < 0.001) compared to DBS prepared from venous blood and handled similarly. Resistance profiles were similar between plasma and DBS specimens. This report delineates the optimal DBS collection, storage, and shipping conditions and opens a new avenue for cost-saving ambient-temperature DBS specimen shipments for HIV drug resistance (HIVDR) surveillances in resource-limited settings. PMID:24871219

  2. Levels of Cotinine in Dried Blood Specimens from Newborns as a Biomarker of Maternal Smoking Close to the Time of Delivery

    PubMed Central

    Yang, Juan; Pearl, Michelle; Jacob, Peyton; DeLorenze, Gerald N.; Benowitz, Neal L.; Yu, Lisa; Havel, Christopher; Kharrazi, Martin

    2013-01-01

    The precise quantitation of smoking during pregnancy is difficult in retrospective studies. Routinely collected blood specimens from newborns, stored as dried blood spots, may provide a low-cost method to objectively measure maternal smoking close to the time of delivery. This article compares cotinine levels in dried blood spots to those in umbilical cord blood to assess cotinine in dried blood spots as a biomarker of maternal smoking close to the time of delivery. The California Genetic Disease Screening Program provided dried blood spots from 428 newborns delivered in 2001–2003 with known umbilical cord blood cotinine levels. Cotinine in dried blood spots was measured in 6.35­-mm punches by using liquid chromatography­–tandem mass spectrometry (quantitation limit, 3.1 ng/mL). Repeated measures of cotinine in dried blood spots were highly correlated (R2 = 0.99, P < 0.001) among 100 dried blood spots with cotinine quantitated in 2 separate punches. Linear regression revealed that cotinine levels in dried blood spots were slightly lower than those in umbilical cord blood and predicted umbilical cord blood cotinine levels well (β = 0.95, R2 = 0.80, and P < 0.001 for both cotinine levels in log10 scale). When defining active smoking as a cotinine level of 10 ng/mL or more and using umbilical cord blood cotinine as the criterion standard, we found that measurements of cotinine in dried blood spots had high sensitivity (92.3%) and specificity (99.7%) in the prediction of maternal active smoking. Cotinine levels in dried blood spots are an accurate biomarker of maternal smoking close to the time of delivery. PMID:24068198

  3. Committee report: Considerations and recommendations for national guidance regarding the retention and use of residual dried blood spot specimens after newborn screening.

    PubMed

    Therrell, Bradford L; Hannon, W Harry; Bailey, Donald B; Goldman, Edward B; Monaco, Jana; Norgaard-Pedersen, Bent; Terry, Sharon F; Johnson, Alissa; Howell, R Rodney

    2011-07-01

    Newborn screening programs are state based with variable policies. Guidance regarding the retention, storage, and use of portions of newborn screening dried blood spots that remain after screening (residual specimens) was first published in 1996. Since then, newborn screening programs have paid increased attention to specimen storage and usage issues. Standard residual specimen uses include quality assurance and program evaluation, treatment efficacy, test refinement, and result verification. In all cases, privacy and security are primary concerns. In general, two distinct state practices regarding the storage and use of residual newborn screening specimens exist: (1) short-term storage (<3 years), primarily for standard program uses and (2) long-term storage (>18 years), for standard program uses and possible important public health research uses. Recently, there have been concerns in some consumer communities regarding both the potential uses of residual specimens and patient (newborn and family) privacy. To assist in policy improvements that can protect the individual's privacy and allow for important public health uses of residual newborn screening specimens, the Secretary of Health and Human Services' Advisory Committee on Heritable Disorders in Newborns and Children has developed recommendations (with requested action by the Secretary where applicable). This report presents the Committee's recommendations and reviews the pertinent associated issues. PMID:21602691

  4. Single genome amplification of proviral HIV-1 DNA from dried blood spot specimens collected during early infant screening programs in Lusaka, Zambia

    PubMed Central

    Seu, Lillian; Mwape, Innocent; Guffey, M. Bradford

    2014-01-01

    The ability to evaluate individual HIV-1 virions from the quasispecies of vertically infected infants was evaluated in a field setting at the Centre for Infectious Disease Research in Zambia. Infant heel-prick blood specimens were spotted onto dried blood spot (DBS) filter paper cards at government health clinics. Nucleic acid was extracted and used as a template for HIV-1 proviral DNA detection by a commercial Amplicor HIV-1 PCR test (Roche, version 1.5). On samples that tested positive by commercial diagnostic assay, amplification of DNA was performed using an in-house assay of the 5′ and 3′ region of the HIV-1 genome. Additionally, fragments covering 1200 nucleotides within pol (full length protease and partial reverse transcriptase) and 1400 nucleotides within env (variable 1-variable 5 region) were further analyzed by single genome amplification (SGA). In summary, we have demonstrated an in-house assay for amplifying the 5′ and 3′ proviral HIV-1 DNA as well as pol and env proviral DNA fragments from DBS cards collected and analyzed entirely in Zambia. In conclusion, this study shows the feasibility of utilizing DBS cards to amplify the whole proviral HIV-1 genome as well as perform SGA on key HIV-1 genes. PMID:24667303

  5. Drying drops of blood

    NASA Astrophysics Data System (ADS)

    Brutin, David; Sobac, Benjamin; Loquet, Boris; Sampol, José.

    2010-11-01

    The drying of a drop of human blood is fascinating by the complexity of the physical mechanisms that occur as well as the beauty of the phenomenon which has never been previously evidenced in the literature. The final stage of full blood evaporation reveals for a healthy person the same regular pattern with a good reproducibility. Other tests on anemia and hyperlipidemic persons were performed and presented different patterns. By means of digital camera, the influence of the motion of red blood cells (RBCs) which represent about 50% of the blood volume, is revealed as well as its consequences on the final stages of drying. The mechanisms which lead to the final pattern of dried blood drops are presented and explained on the basis of fluid and solid mechanics in conjunction with the principles of hematology. Our group is the first to evidence that the specific regular patterns characteristic of a healthy individual do not appear in a dried drop of blood from a person with blood disease. Blood is a complex colloidal suspension for which the flow motion is clearly non-Newtonian. When drops of blood evaporate, all the colloids are carried by the flow motion inside the drop and interact.

  6. Simultaneous Detection of Major Drug Resistance Mutations of HIV-1 Subtype B Viruses from Dried Blood Spot Specimens by Multiplex Allele-Specific Assay.

    PubMed

    Zhang, Guoqing; Cai, Fangping; de Rivera, Ivette Lorenzana; Zhou, Zhiyong; Zhang, Jing; Nkengasong, John; Gao, Feng; Yang, Chunfu

    2016-01-01

    A multiplex allele-specific (MAS) assay has been developed for the detection of HIV-1 subtype C drug resistance mutations (DRMs). We have optimized the MAS assay to determine subtype B DRMs in dried blood spots (DBS) collected from patients on antiretroviral therapy. The new assay accurately detected DRMs, including low-abundance mutations that were often missed by Sanger sequencing. PMID:26560533

  7. Simultaneous Detection of Major Drug Resistance Mutations of HIV-1 Subtype B Viruses from Dried Blood Spot Specimens by Multiplex Allele-Specific Assay

    PubMed Central

    Zhang, Guoqing; Cai, Fangping; de Rivera, Ivette Lorenzana; Zhou, Zhiyong; Zhang, Jing; Nkengasong, John

    2015-01-01

    A multiplex allele-specific (MAS) assay has been developed for the detection of HIV-1 subtype C drug resistance mutations (DRMs). We have optimized the MAS assay to determine subtype B DRMs in dried blood spots (DBS) collected from patients on antiretroviral therapy. The new assay accurately detected DRMs, including low-abundance mutations that were often missed by Sanger sequencing. PMID:26560533

  8. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Blood specimen collection device. 862.1675 Section... Systems § 862.1675 Blood specimen collection device. (a) Identification. A blood specimen collection device is a device intended for medical purposes to collect and to handle blood specimens and to...

  9. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Blood specimen collection device. 862.1675 Section... Systems § 862.1675 Blood specimen collection device. (a) Identification. A blood specimen collection device is a device intended for medical purposes to collect and to handle blood specimens and to...

  10. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Blood specimen collection device. 862.1675 Section... Systems § 862.1675 Blood specimen collection device. (a) Identification. A blood specimen collection device is a device intended for medical purposes to collect and to handle blood specimens and to...

  11. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Blood specimen collection device. 862.1675 Section... Systems § 862.1675 Blood specimen collection device. (a) Identification. A blood specimen collection device is a device intended for medical purposes to collect and to handle blood specimens and to...

  12. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Blood specimen collection device. 862.1675 Section... Systems § 862.1675 Blood specimen collection device. (a) Identification. A blood specimen collection device is a device intended for medical purposes to collect and to handle blood specimens and to...

  13. Stabilized dried blood spot collection.

    PubMed

    McMorran, Darren; Chung, Dwayne Chung Kim; Toth, Monika; Liew, Oi Wah; Muradoglu, Murat; Ng, Tuck Wah

    2016-08-01

    During the collection phase of the dried blood spot method, practitioners need to ensure that there is no smearing of the blood sample on the filter paper or else readings from it will be invalid. This can be difficult to accomplish in the field if there is relative motion between the site of blood discharge on the finger and the filter paper. In this article, a gyroscope stabilization method is introduced and demonstrated to provide consistent and improved dried blood spot collection within a circular guide region notwithstanding the presence of rocking. PMID:27156813

  14. Microwave applications to rock specimen drying in laboratory

    NASA Astrophysics Data System (ADS)

    Park, Jihwan; Park, Hyeong-Dong

    2014-05-01

    Microwave heating is the process in which electromagnetic wave with 300 MHz - 300 GHz heats dielectric material. Although in the beginning microwave was mainly used in food industry to cook or heat the food, it soon became clear that microwave had a large potential for other applications. It was thus introduced in geological fields of investigation like mineral processing, oil sand and oil shale extraction, soil remediation, waste treatment. However, the drying techniques using microwave was rarely treated in geology field. According to the ISRM suggested methods, experimental rock specimens in laboratory test were dried in 105°C oven for a period of at least 24 hours. In this method, hot air transmits heats to material by means of thermal conduction, and the heat was transferred from the surface to the inside of the rock specimens. The thermal gradient and moisture gradient can deteriorate the specimens, and energy can be wasted in bulk heating the specimens. The aim of our study was to compare physical property, microstructural property, and energy efficiency between microwave drying method and conventional oven drying method, and to suggest new method for rock drying. Granite, basalt, and sandstone were selected as specimens and were made in cylinder shape with 54 mm diameter. To compare two different methods, one set of saturated specimens were dried in 105°C conventional oven and the other set of saturated specimens were dried in microwave oven. After dried, the specimens were cooled and saturated in 20°C water 48 hours. The saturation-drying were repeated 50 cycles, and the physical property and microstructural property were measured every 10 cycles. Absorption and elastic wave velocity were measured to investigate the change of physical property, and microscope image and X-ray computed tomography image were obtained to investigate the change of microstructural property of rock specimens. The electricity consumption of conventional oven and microwave oven

  15. Freeze-dried specimens for gross anatomy teaching

    PubMed Central

    O'SULLIVAN, E.; STEWART, I. J.

    1999-01-01

    As more and more emphasis is placed on the use of prosected specimens to support teaching and learning of gross anatomy, consideration must be given to developing new methods to preserve human cadaveric material, and in ways which will resist the wear and tear to which they are necessarily subjected. Taxidermists have developed techniques for freeze-drying whole small animals as a method of long term preservation (Metcalf, 1981). We have explored the use of this methodology to preserve small prosected specimens for use in the teaching of gross anatomy. The technique we report here was tested initially on larynges (Fig. 1) but has since been applied with equal success to other structures, including pieces of small intestine dissected to show the arterial arcades (Fig. 2). We have used material from cadavers which were preserved using our standard embalming procedure (O'Sullivan & Mitchell, 1993). PMID:10529067

  16. Comparison of blood specimens from plain and gel vacuum blood collection tubes.

    PubMed

    Wiwanitkit, V

    2001-05-01

    This study was set in the Division of Laboratory Medicine, Chulalongkorn Hospital. All 2,000 blood specimens were randomly collected using evacuated blood collection by plain or gel vacuum tubes. After collection, each specimen was considered and judged using criteria of specimen rejection to determine how proper the specimen presentations were. All data were reviewed, collected and interpreted. It revealed that there were only 20 (1%) improper specimens and all were improper in quality. There was no significant difference between the ratio of improper specimens in both groups (P > 0.30). From this study, it revealed that efficacy of both types of vacuum tubes was not different. The new gel vacuum tube seems to be an effective tool in the evacuated blood collection system due to its advantage in reduction of time in specimen processing. PMID:11560224

  17. Pattern formation in drying drops of blood

    NASA Astrophysics Data System (ADS)

    Brutin, D.; Sobac, B.; Loquet, B.; Sampol, J.

    2011-01-01

    The drying of a drop of human blood exhibits coupled physical mechanisms, such as Marangoni flow, evaporation and wettability. The final stage of a whole blood drop evaporation reveals regular patterns with a good reproducibility for a healthy person. Other experiments on anaemic and hyperlipidemic people were performed, and different patterns were revealed. The flow motion inside the blood drop is observed and analyzed with the use of a digital camera: the influence of the red blood cells (RBCs) motion is revealed at the drop periphery as well as its consequences on the final stage of drying. The mechanisms which lead to the final pattern of the dried blood drops are presented and explained on the basis of fluid mechanics in conjunction with the principles of haematology. The blood drop evaporation process is evidenced to be driven only by Marangoni flow. The same axisymetric pattern formation is observed, and can be forecast for different blood drop diameters. The evaporation mass flux can be predicted with a good agreement, assuming only the knowledge of the colloids mass concentration.

  18. Degradation and Stabilization of Peptide Hormones in Human Blood Specimens

    PubMed Central

    Yi, Jizu; Warunek, David; Craft, David

    2015-01-01

    Plasma hormone peptides, including GLP-1, GIP, Glucagon, and OXM, possess multiple physiological roles and potential therapeutic and diagnostic utility as biomarkers in the research of metabolic disorders. These peptides are subject to proteolytic degradation causing preanalytical variations. Stabilization for accurate quantitation of these active peptides in ex vivo blood specimens is essential for drug and biomarker development. We investigated the protease-driven instability of these peptides in conventional serum, plasma, anticoagulated whole blood, as well as whole blood and plasma stabilized with protease inhibitors. The peptide was monitored by both time-course Matrix-Assisted Laser Desorption Ionization Time-to-Flight Mass Spectrometry (MALDI –TOF MS) and Ab-based assay (ELISA or RIA). MS enabled the identification of proteolytic fragments. In non-stabilized blood samples, the results clearly indicated that dipeptidyl peptidase-IV (DPP-IV) removed the N-terminal two amino acid residues from GLP-1, GIP and OXM(1-37) and not-yet identified peptidase(s) cleave(s) the full-length OXM(1-37) and its fragments. DPP-IV also continued to remove two additional N-terminal residues of processed OXM(3–37) to yield OXM(5–37). Importantly, both DPP-IV and other peptidase(s) activities were inhibited efficiently by the protease inhibitors included in the BD P800* tube. There was preservation of GLP-1, GIP, OXM and glucagon in the P800 plasma samples with half-lives > 96, 96, 72, and 45 hours at room temperature (RT), respectively. In the BD P700* plasma samples, the stabilization of GLP-1 was also achieved with half-life > 96 hours at RT. The stabilization of these variable peptides increased their utility in drug and/or biomarker development. While stability results of GLP-1 obtained with Ab-based assay were consistent with those obtained by MS analysis, the Ab-based results of GIP, Glucagon, and OXM did not reflect the time-dependent degradations revealed by MS

  19. Effects of drying conditions, admixtures and specimen size on shrinkage strains

    SciTech Connect

    Al-Saleh, Saleh A. . E-mail: alsaleh@dr.com; Al-Zaid, Rajeh Z.

    2006-10-15

    The paper presents the results of an experimental investigation on the effects of drying conditions, specimen size and presence of plasticizing admixture on the development of shrinkage strains. The measurements are taken in a harsh (50 deg. C and 5% R.H.) and a moderate environment (28 deg. C and 50% R.H.). The results include strain development at various levels of cross sections of concrete prisms. The drying conditions are found to be the dominant parameter affecting the shrinkage strain development particularly in specimens of smaller sizes. The effect of plasticizing admixture on shrinkage strains is negligible.

  20. [Counter-measures against patient misidentification and specimen mismanagement with blood collection].

    PubMed

    Ohgoe, Kazuhiko

    2013-08-01

    My theme for this symposium is counter-measures against patient misidentification and specimen mismanagement with blood collection due to the lack of using authentication systems. What is applicable to our laboratory is patient misidentification counter-measures for specimen management at the time of inpatient ward blood collection and specimen examination (mistakes in appending bar code labels and entering specimen numbers). During the period from January 2008 to July 2012 at our laboratory, there were 9 cases of patient misidentification for hospital ward blood collection and specimen management. There were 2 cases in 2008 (1 for blood collection, 1 for specimen management), no cases in 2009, 3 cases in 2010 involving blood collection, 1 case in 2011 involving specimen management, and 3 cases in 2012 (1 for blood collection, 2 for specimen management). All patient misidentifications involving hospital ward blood collection arose from bedside blood collection. As a counter-measure, training slides were created at a medical safety management review session, repeated training in attention and patient check procedures was conducted with staff members, and hands-on training in pointing and naming was carried out. With these training slides, the goal was the execution of verification duties by encouraging conversations that include the patient's name, such as "Mr./Ms. XXXX, today we'll be collecting 3 tubes of blood," as a link to patient verification duties. With specimen management, 3 of the 4 cases occurred during overtime for day-shift work. As counter measures: 1) adherence to 1 patient, 1 tray, and signing when matching; 2) as a counter-measure against mistaking specimens for blood gas hemolysis, confirmation as other specimens, separating approximately 3 drops of blood that cannot be used for sampling, confirming hemolysis, and preventing misidentification. PMID:24218774

  1. Panfungal PCR Assay for Detection of Fungal Infection in Human Blood Specimens

    PubMed Central

    Van Burik, Jo-Anne; Myerson, David; Schreckhise, Randall W.; Bowden, Raleigh A.

    1998-01-01

    A novel panfungal PCR assay which detects the small-subunit rRNA gene sequence of the two major fungal organism groups was used to test whole-blood specimens obtained from a series of blood or bone marrow transplant recipients. The 580-bp PCR product was identified after amplification by panfungal primers and hybridization to a 245-bp digoxigenin-labeled probe. The lower limit of detection of the assay was approximately four organisms per milliliter of blood. Multiple whole-blood specimens from five patients without fungal infection or colonization had negative PCR results. Specimens from 11 infected patients had positive PCR results. Blood from three patients with pulmonary aspergillosis had positive PCR results: one patient’s blood specimen obtained in the week prior to the diagnosis of infection by a positive bronchoalveolar lavage fluid culture result was positive by PCR, and blood specimens obtained from two patients 1 to 2 days after lung biopsy and which were sterile by culture were positive by PCR. The blood of four patients with candidemia, three patients with mixed fungal infections, and one patient with fusariosis also had positive PCR signals. The panfungal PCR assay can detect multiple fungal genera and may be used as an adjunct to conventional methods for the detection of fungal infection or for describing the natural history of fungal infection. Further studies are needed to define the sensitivity and specificity of this assay for the diagnosis of fungal infection prior to the existence of other clinical or laboratory indications of invasive fungal infection. PMID:9574670

  2. Effectiveness of saliva and fingerprints as alternative specimens to urine and blood in forensic drug testing.

    PubMed

    Kuwayama, Kenji; Miyaguchi, Hajime; Yamamuro, Tadashi; Tsujikawa, Kenji; Kanamori, Tatsuyuki; Iwata, Yuko T; Inoue, Hiroyuki

    2016-07-01

    In forensic drug testing, it is important to immediately take biological specimens from suspects and victims to prove their drug intake. We evaluated the effectiveness of saliva and fingerprints as alternative specimens to urine and blood in terms of ease of sampling, drug detection sensitivity, and drug detection periods for each specimen type. After four commercially available pharmaceutical products were administered to healthy subjects, each in a single dose, their urine, blood, saliva, and fingerprints were taken at predetermined sampling times over approximately four weeks. Fourteen analytes (the administered drugs and their main metabolites) were extracted from each specimen using simple pretreatments, such as dilution and deproteinization, and were analyzed using liquid chromatography/mass spectrometry (LC/MS). Most of the analytes were detected in saliva and fingerprints, as well as in urine and blood. The time-courses of drug concentrations were similar between urine and fingerprints, and between blood and saliva. Compared to the other compounds, the acidic compounds, for example ibuprofen, acetylsalicylic acid, were more difficult to detect in all specimens. Acetaminophen, dihydrocodeine, and methylephedrine were detected in fingerprints at later sampling times than in urine. However, a relationship between the drug structures and their detection periods in each specimen was not found. Saliva and fingerprints could be easily sampled on site without using special techniques or facilities. In addition, fingerprints could be immediately analyzed after simple and rapid treatment. In cases where it would be difficult to immediately obtain urine and blood, saliva and fingerprints could be effective alternative specimens for drug testing. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26074137

  3. Ancient pathogens in museal dry bone specimens: analysis of paleocytology and aDNA.

    PubMed

    Gaul, Johanna Sophia; Winter, Eduard; Grossschmidt, Karl

    2015-04-01

    Bone samples investigated in this study derive from the pathologic-anatomical collection of the Natural History Museum of Vienna. In order to explore the survival of treponemes and treponemal ancient DNA in museal dry bone specimens, we analyzed three individuals known to have been infected with Treponema pallidum pallidum. No reproducible evidence of surviving pathogen's ancient DNA (aDNA) was obtained, despite the highly sensitive extraction and amplification techniques (TPP15 and arp). Additionally, decalcification fluid of bone sections was smear stained with May-Gruenwald-Giemsa. The slides were examined using direct light microscope and dark field illumination. Remnants of spirochetal structures were detectable in every smear. Our results demonstrate that aDNA is unlikely to survive, but spirochetal remains are stainable and thus detectable. PMID:25994097

  4. The stability of hexacosanoyl lysophosphatidylcholine in dried-blood spot quality control materials for X-linked adrenoleukodystrophy newborn screening

    PubMed Central

    Haynes, Christopher A.; De Jesús, Víctor R.

    2016-01-01

    Objectives Newborn screening for X-linked adrenoleukodystrophy utilizes tandem mass spectrometry to analyze dried-blood spot specimens. Quality control materials (dried-blood spots enriched with hexacosanoyl lysophosphatidylcholine) were prepared and stored at different temperatures for up to 518 days to evaluate the stability of this biomarker for X-linked adrenoleukodystrophy. Design and methods Dried-blood spot storage included desiccant (45, 171, and 518 days) or omitted desiccant (53 days at >90% relative humidity). Specimens were stored for 171 and 518 days at −20 °C, 4 °C, ambient temperature, and 37 °C. Each weekday for 45 days, a bag of specimens stored at 4 °C was warmed to ambient temperature and one specimen was removed for storage at −80 °C. Specimens were analyzed by high-performance liquid-chromatography electrospray ionization tandem mass spectrometry and data was plotted as concentration (micromoles per liter) vs. time. Linear regression provided slope and y-intercept values for each storage condition. Results Small slope values (0.01 or less) and y-intercept values close to the enrichment indicated less than 11% loss of hexacosanoyl lysophosphatidylcholine under all storage conditions tested. Conclusions Quality control materials for X-linked adrenoleukodystrophy are stable for at least 1 year when stored with desiccant. PMID:25307302

  5. A digital microfluidic method for dried blood spot analysis.

    PubMed

    Jebrail, Mais J; Yang, Hao; Mudrik, Jared M; Lafrenière, Nelson M; McRoberts, Christine; Al-Dirbashi, Osama Y; Fisher, Lawrence; Chakraborty, Pranesh; Wheeler, Aaron R

    2011-10-01

    Blood samples stored as dried blood spots (DBSs) are emerging as a useful sampling and storage vehicle for a wide range of applications. Unfortunately, the surging popularity of DBS samples has not yet been accompanied by an improvement in automated techniques for extraction and analysis. As a first step towards overcoming this challenge, we have developed a prototype microfluidic system for quantification of amino acids in dried blood spots, in which analytes are extracted, mixed with internal standards, derivatized, and reconstituted for analysis by (off-line and in-line) tandem mass spectrometry. The new method is fast, robust, precise, and most importantly, compatible with automation. We propose that the new method can potentially contribute to a new generation of analytical techniques for quantifying analytes in DBS samples for a wide range of applications. PMID:21869989

  6. Process for measuring low cadmium levels in blood and other biological specimens

    DOEpatents

    Peterson, David P.; Huff, Edmund A.; Bhattacharyya, Maryka H.

    1994-05-03

    A process for measuring low levels of cadmium in blood and other biological specimens is provided without interference from high levels of alkali metal contaminants by forming an aqueous solution and without contamination by environmental cadmium absent the proteins from the specimen, selectively removing cadmium from the aqueous solution on an anion exchange resin, thereby removing the alkali metal contaminants, resolubilizing cadmium from the resin to form a second solution and analyzing the second solution for cadmium, the process being carried out in a cadmium-free environment.

  7. Process for measuring low cadmium levels in blood and other biological specimens

    DOEpatents

    Peterson, David P.; Huff, Edmund A.; Bhattacharyya, Maryka H.

    1994-01-01

    A process for measuring low levels of cadmium in blood and other biological specimens is provided without interference from high levels of alkali metal contaminants by forming an aqueous solution and without contamination by environmental cadmium absent the proteins from the specimen, selectively removing cadmium from the aqueous solution on an anion exchange resin, thereby removing the alkali metal contaminants, resolubilizing cadmium from the resin to form a second solution and analyzing the second solution for cadmium, the process being carried out in a cadmium-free environment.

  8. Quantitation of HIV-1 RNA in dried blood and plasma spots.

    PubMed

    Reigadas, S; Schrive, M H; Aurillac-Lavignolle, V; Fleury, H J

    2009-10-01

    Dried blood spots (DBSs) and dried plasma spots (DPSs) are an attractive method for serological and molecular diagnosis of HIV infection. Recently, Youngpairoj et al. [Youngpairoj, A.S., Masciotra, S., Garrido, C., Zahonero, N., de Mendoza, C., Garcia-Lerma, J.G., 2008. HIV-1 drug resistance genotyping from dried blood spots stored for 1 year at 4 degrees C. J. Antimicrob. Chemother. 61, 1217-1220] showed that HIV-1 can be genotyped efficiently from DBS specimens stored at 4 degrees C for 1 year. The viral load obtained from DBS and DPS samples was compared with that obtained from plasma samples. A total of 86 samples was prepared from people infected with HIV subtype B or non-B and spotted on 903 filter papers stored with desiccant at 4 degrees C. RNA was extracted using the QIAamp Viral RNA mini kit (Qiagen, Courtaboeuf, France). RNA from DBS or DPS samples was quantified in accordance with the Agence Nationale de Recherche sur le SIDA (ANRS, AC11, Paris, France) assay for HIV-1 quantitation. When the mean viral load of plasma samples and DPS samples or plasma samples and DBS samples were compared, there were no significant differences. The overall data showed that although the sensitivity threshold of the assays was different, there was a correlation between the three specimen types and that DBS and DPS samples can be routinely used for viral load quantification particularly in resource-limited settings. PMID:19523984

  9. Hi-Plex targeted sequencing is effective using DNA derived from archival dried blood spots.

    PubMed

    Nguyen-Dumont, T; Mahmoodi, M; Hammet, F; Tran, T; Tsimiklis, H; Giles, G G; Hopper, J L; Southey, M C; Park, D J

    2015-02-01

    Many genetic epidemiology resources have collected dried blood spots (predominantly as Guthrie Cards) as an economical and efficient means of archiving sources of DNA, conferring great value to genetic screening methods that are compatible with this medium. We applied Hi-Plex to screen the breast cancer predisposition gene PALB2 in 93 Guthrie Card-derived DNA specimens previously characterized for PALB2 genetic variants via DNA derived from lymphoblastoid cell lines, whole blood, and buffy coat. Of the 93 archival Guthrie Card-derived DNAs, 92 (99%) were processed successfully and sequenced using approximately half of a MiSeq run. From these 92 DNAs, all 59 known variants were detected and no false-positive variant calls were yielded. Fully 98.13% of amplicons (5417/5520) were represented within 15-fold of the median coverage (2786 reads), and 99.98% of amplicons (5519/5520) were represented at a depth of 10 read-pairs or greater. With Hi-Plex, we show for the first time that a High-Plex amplicon-based massively parallel sequencing (MPS) system can be applied effectively to DNA prepared from dried blood spot archival specimens and, as such, can dramatically increase the scopes of both method and resource. PMID:25447460

  10. Hi-Plex targeted sequencing is effective using DNA derived from archival dried blood spots

    PubMed Central

    Nguyen-Dumont, T; Mahmoodi, M; Hammet, F; Tran, T; Tsimiklis, H; Giles, GG; Hopper, JL; Southey, MC; Park, DJ

    2014-01-01

    Many genetic epidemiology resources have collected dried blood spots (predominantly as Guthrie Cards) as an economical and efficient means of archiving sources of DNA, conferring great value to genetic screening methods that are compatible with this medium. We applied Hi-Plex to screen the breast cancer predisposition gene PALB2 in 93 Guthrie Card-derived DNA specimens previously characterised for PALB2 genetic variants via DNA derived from lymphoblastoid cell lines, whole blood and buffy coat. 92 of the 93 archival Guthrie Card-derived DNAs (99%) were processed successfully and sequenced using approximately half of a MiSeq run. From these 92, all 59 known variants were detected and no false-positive variant calls were yielded. 98.13% of amplicons (5417/5520) were represented within 15-fold of the median coverage (2786 reads) and 99.98% of amplicons (5519/5520) were represented at a depth of 10 read-pairs or greater. With Hi-Plex, we show for the first time that a high-plex amplicon based MPS system can be applied effectively to DNA prepared from dried blood spot archival specimens and, as such, dramatically increase the scopes of both method and resource. PMID:25447460

  11. Hemoglobin variant analysis of whole blood and dried blood spots by MS.

    PubMed

    Edwards, Rebecca L; Martin, Nicholas J; Cooper, Helen J

    2013-08-01

    MS allows for the unequivocal diagnosis of hemoglobin variants, or hemoglobinopathies. Hemoglobinopathies are the most common inherited disorder and there is a need for rapid detection of clinically significant variants, such as sickle hemoglobin, which is responsible for sickle cell disease. In this review, we describe the development of MS approaches for the determination of hemoglobin variants from both whole blood samples and dried blood spots. MS approaches that are suitable for population screening are discussed, as are recent advances in direct surface analysis of dried blood spots. PMID:23937138

  12. Evaluation of PNA FISH® Yeast Traffic Light in identification of Candida species from blood and non-blood culture specimens.

    PubMed

    Radic, Marina; Goic-Barisic, Ivana; Novak, Anita; Rubic, Zana; Tonkic, Marija

    2016-08-01

    PNA FISH(®) (peptide nucleic acid fluorescent in situ hybridization) Yeast Traffic Light (PNA FISH(®) YTL) assay is a commercially avaliable method for rapid identification of Candida spp. directly from positive blood cultures. This report provides a one-year experience in identification of yeasts from 25 specimens (15 positive blood cultures and 10 other clinically significant specimens) using PNA FISH(®) YTL and comparing it to VITEK 2 System. Overall, assay identification compatibility with VITEK 2 System was found among 21/25 (84%) isolates tested. Only 3/25 (12%) of the isolates were not identified, and one isolate was misidentified by the PNA FISH(®) YTL assay. Our results show that the assay is a reliable method in identification of Candida spp. not only from blood cultures, but even from other clinically significant specimens (urine cultures, catheter tip cultures, peritoneal fluid cultures) when compared to automated method like VITEK 2 System. This novel application of the PNA FISH(®) YTL assay could therefore contribute to cost savings and significant benefit to patients, as rapid information about isolated yeast species is provided. PMID:27067303

  13. [Diagnosis of congenital cytomegalovirus infection in newborn dried blood spots on Guthrie cards. A promissory technique].

    PubMed

    Distéfano, Angélica L; González, Cecilia A; Pardón, Fabián; Sarubi, María A; Canero Velazco, Cristina

    2008-04-01

    Laboratories play a crucial role in the diagnosis of congenital and perinatal cytomegalovirus infection, considering that other viral infections in newborn infants have similar clinical characteristics. The objectives of this work are to compare the results of the polymerase reaction in blood spots and urine as well as point out the relevance of the result in the Guthrie cards to differentiate congenital from perinatal infection. A total of 148 patients suspicious of CMVH infections were studied in the Congenital Perinatal Infections and Sexual Transmission Laboratory, at the National Institute "Carlos G. Malbrán". The dry blood samples (Guthrie cards) and urine of all patients were studied through the polymerase chain reaction. From the 148 patients, 3 presented other infections, 95 tested negative and 50 positive for cytomegalovirus: 35 had congenital infection and 15 perinatal. In the congenital cases, the polymerase reaction in dry blood was positive (sensitivity 100%, specificity 98.9%, VPP 98% and VPN 100%). Four of them with tardive symptoms were studied retrospectively. The urine specimens from the remaining 15 patients that were taken 15 days after birth were analyzed through the same methods, showing a sensitivity of 100%, the retrospective analysis of this dry blood group yielded negative results, so the infection was considered perinatal. Thus, the dry blood polymerase reaction of the newborn infants makes it a reliable assay for diagnosing congenital cytomegalovirus infection and could be used as an alternative method to urine polymerase reaction. In addition, this test is able to reveal whether the infection is congenital or perinatal in those cases of late symptom or other cases of controversial origin. PMID:18661038

  14. Molecular Detection of Streptococcus pneumoniae on Dried Blood Spots from Febrile Nigerian Children Compared to Culture

    PubMed Central

    Iroh Tam, Pui-Ying; Hernandez-Alvarado, Nelmary; Schleiss, Mark R.; Hassan-Hanga, Fatimah; Onuchukwu, Chuma; Umoru, Dominic; Obaro, Stephen K.

    2016-01-01

    Background Nigeria has one of the highest burdens of pneumococcal disease in the world, but accurate surveillance is lacking. Molecular detection of infectious pathogens in dried blood spots (DBS) is an ideal method for surveillance of infections in resource-limited settings because of its low cost, minimal blood volumes involved, and ease of storage at ambient temperature. Our study aim was to evaluate a Streptococcus pneumoniae real-time polymerase chain reaction (rt-PCR) assay on DBS from febrile Nigerian children on Whatman 903 and FTA filter papers, compared to the gold standard of culture. Methods Between September 2011 to May 2015, blood was collected from children 5 years of age or under who presented to six hospital study sites throughout northern and central Nigeria with febrile illness, and inoculated into blood culture bottles or spotted onto Whatman 903 or FTA filter paper. Culture and rt-PCR were performed on all samples. Results A total of 537 DBS specimens from 535 children were included in the study, of which 15 were culture-positive for S. pneumoniae. The rt-PCR assay detected S. pneumoniae in 12 DBS specimens (2.2%). One positive rt-PCR result was identified in a culture-negative specimen from a high-risk subject, and two positive rt-PCR results were negative on repeat testing. Six culture-confirmed cases of S. pneumoniae bacteremia were missed. Compared to culture, the overall sensitivities of Whatman 903 and FTA DBS for detection of S. pneumoniae were 57.1% (95% CI 18.4–90.1%) and 62.5% (95% CI 24.5–91.5%), respectively. Nonspecific amplification was noted in an additional 22 DBS (4.1%). Among these, six were positive for a non-S. pneumoniae pathogen on culture. Conclusions Rt-PCR was able to detect S. pneumoniae from clinical DBS specimens, including from a culture-negative specimen. Our findings show promise of this approach as a surveillance diagnostic, but also raise important cautionary questions. Several DBS specimens were detected as

  15. [Blood sampling using "dried blood spot": a clinical biology revolution underway?].

    PubMed

    Hirtz, Christophe; Lehmann, Sylvain

    2015-01-01

    Blood testing using the dried blood spot (DBS) is used since the 1960s in clinical analysis, mainly within the framework of the neonatal screening (Guthrie test). Since then numerous analytes such as nucleic acids, small molecules or lipids, were successfully measured on the DBS. While this pre-analytical method represents an interesting alternative to classic blood sampling, its use in routine is still limited. We review here the different clinical applications of the blood sampling on DBS and estimate its future place, supported by the new methods of analysis as the LC-MS mass spectrometry. PMID:25582720

  16. 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.

  17. EFFECT OF SPECIMEN SIZE, SHAPE, AND ORIENTATION ON THE DRY DEPOSITION TO GALVANIZED STEEL SURFACES

    EPA Science Inventory

    An analysis is presented of the variability in chemical composition of soluble corrosion products on galvanized steel specimens exposed at Research Triangle Park, NC, in the absence of natural precipitation. he specimens varied in size, shape, orientation angle, and previous expo...

  18. Rapid determination of rifaximin on dried blood spots by LC-ESI-MS.

    PubMed

    Rao, R Nageswara; Vali, R Mastan; Ramachandra, Bondigalla; Maurya, Pawan K

    2011-11-01

    The use of blood spot collection cards is a simple way to obtain specimens for therapeutic drug monitoring, assessing adherence to medications and preventing toxicity in a clinical setting. A high-throughput liquid chromatography-electrospray ionization mass spectrometric (LC-ESI-MS) method for determination of rifaximin on dried blood spots (DBS) was developed and validated. It involves solvent extraction of a punch of DBS followed by reversed-phase LC on a monolithic column consisting of a silica rod with bimodal pore structure and detection by ESI-MS. Rifampicin was used as an internal standard (IS). The run time was within 5.0 min with a very low back-pressure at a flow rate of 0.5 mL/min. The assay was linear from 0.1 to 10 ng/mL. The mean recovery was 98.42%. The developed method is very simple, rapid and useful for clinical applications. PMID:21287584

  19. Airway blood flow response to dry air hyperventilation in sheep

    SciTech Connect

    Parsons, G.H.; Baile, E.M.; Pare, P.D.

    1986-03-01

    Airway blood flow (Qaw) may be important in conditioning inspired air. To determine the effect of eucapneic dry air hyperventilation (hv) on Qaw in sheep the authors studied 7 anesthetized open-chest sheep after 25 min. of warm dry air hv. During each period of hv the authors have recorded vascular pressures, cardiac output (CO), and tracheal mucosal and inspired air temperature. Using a modification of the reference flow technique radiolabelled microspheres were injected into the left atrium to make separate measurements after humid air and dry air hv. In 4 animals a snare around the left main pulmonary artery was used following microsphere injection to prevent recirculation (entry into L lung of microspheres from the pulmonary artery). Qaw to the trachea and L lung as measured and Qaw for the R lung was estimated. After the final injection the sheep were killed and bronchi (Br) and lungs removed. Qaw (trachea plus L lung plus R lung) in 4 sheep increased from a mean of 30.8 to 67.0 ml/min. Airway mucosal temp. decreased from 39/sup 0/ to 33/sup 0/C. The authors conclude that dry air hv cools airway mucosa and increases Qaw in sheep.

  20. Quality impact on diagnostic blood specimen collection using a new device to relieve venipuncture pain.

    PubMed

    Lima-Oliveira, Gabriel; Lippi, Giuseppe; Salvagno, Gian Luca; Montagnana, Martina; Picheth, Geraldo; Guidi, Gian Cesare

    2013-07-01

    A new device called Buzzy(®) has been recently presented that combines a cooling ice pack and a vibrating motor in order to relieve the venipuncture pain. The aim of this study was to evaluate the impact of Buzzy(®) use during diagnostic blood specimen collection by venipuncture for routine immunochemistry tests. Blood was collected from 100 volunteers by a single, expert phlebotomist. A vein was located on the left forearm without applying tourniquet, in order to prevent any interference from venous stasis, and blood samples were collected using a 20-G straight needle directly into 5 mL vacuum tubes with clot activator and gel separator. In sequence, external cold and vibration by Buzzy(®) was applied on the right forearm-5 cm above the chosen puncture site-for 1 min before venipuncture and continued until the end of the same procedure already done in the left forearm. The panel of tests included the following: glucose, total cholesterol, HDL-cholesterol, triglycerides, total protein, albumin, c-reactive protein, urea, creatinine, uric acid, alkaline phosphatase, amylase, AST, ALT, g-glutamyltransferase, lactate dehydrogenase, creatine kinase, total bilirubin, phosphorus, calcium, magnesium, iron, sodium, potassium, chloride, lipase, cortisol, insulin, thyroid-stimulating hormone, total triiodothyronine, free triiodothyronine, total thyroxine, free thyroxine and haemolysis index. Clinically significant differences between samples were found only for: total protein, albumin and transferrin. The Buzzy(®) can be used during diagnostic blood specimens collection by venipuncture for the majority of the routine immunochemistry tests. We only suggest avoiding this device during blood collection when protein, albumin and transferrin determinations should be performed. PMID:24426217

  1. Incorporation of dried blood alpha fetoprotein into traditional first trimester Down syndrome screening service

    PubMed Central

    Carmichael, Jonathan; Krantz, David; Liu, Hsiao-Pin; Janik, David; Hallahan, Terrence

    2015-01-01

    Abstract Objective The aim of this study was to determine whether incorporation of dried blood alpha fetoprotein (AFP) into first trimester screening using the biochemical markers free Beta human chorionic gonadotropin (hCG) and pregnancy-associated plasma protein A (PAPP-A) can improve screening performance. Methods A retrospective study of 34 Down syndrome and 1185 unaffected dried blood specimens. First trimester dried blood AFP was performed using in-house immunofluorometric time-resolved assay. False positive and detection rates were determined from modeling. Results The multiple of the median in Down syndrome cases was 0.73. At a fixed 5% false positive rate, incorporating AFP into a free Beta hCG, PAPP-A, and nuchal translucency protocol adds 2% detection resulting in detection rates of 92% to 94% depending on the gestational age of the blood draw. At a fixed 90% detection rate, AFP reduced the false positive rate by 1.0 to 1.6 percentage points depending on gestational age. Using a cutoff of 1/1000, the combination of free beta hCG, PAPP-A, AFP, and nuchal translucency achieved a detection rate of 96% with a false positive rate of 8.4% to 9.9%. Adding in nasal bone increased detection to 98% while reducing false positive rates to 4.1% to 4.7%. Conclusion Inclusion of dried blood AFP into traditional first trimester screening improves detection while optimizing contingent protocols so that cell-free fetal DNA testing may be offered in a more cost effective manner. © 2015 The Authors. Prenatal Diagnosis published by John Wiley & Sons Ltd. What’s already known about this topic? Traditional first trimester Down syndrome screening programs can achieve detection rates of 90% for a 5% false positive rate. Cell-free fetal DNA screening can achieve high detection rates and low false positive rates but due to its cost may be ill-suited to population-wide screening. What does this study add? Incorporation of AFP into a dried blood first trimester screening program

  2. Experimental basis of standardized specimen collection: effect of posture on blood picture.

    PubMed

    Leppänen, E A; Gräsbeck, R

    1988-03-01

    22 subjectively healthy females were supine, sat in an armchair and stood while specimens of peripheral venous blood were collected after at least 15 min in each position without using a tourniquet. The albumin, haemoglobin, erythrocyte concentration and the haematocrit increased significantly when the subjects assumed a more erect position, probably as a result of increased hydrostatic pressure. The leucocyte count did not rise, and there was a statistically significant drop in the lymphocyte concentration when changing from supine to sitting. However, the leucocyte concentration rose significantly from supine to sitting or standing. When interpreting laboratory data, the difference in the behaviour of different cell species should be kept in mind. However, on the whole this study supports the stipulation contained in international recommendations that the posture of the subject should be standardized before collection of peripheral blood for haematological tests. PMID:3356238

  3. Detection of immunoglobulin isotypes from dried blood spots.

    PubMed

    Andersen, Nancy J; Mondal, Tapan Kumar; Preissler, Mark T; Freed, Brian M; Stockinger, Sabine; Bell, Erin; Druschel, Charlotte; Louis, Germaine M Buck; Lawrence, David A

    2014-02-01

    The study was designed to determine the sensitivity and reproducibility of recovering immunoglobulin (Ig) isotypes (IgG subclasses, IgA, IgE and IgM classes) from dried blood spots (DBS), a methodologic subcomponent of the Upstate KIDS Study. A multiplexed Luminex assay was used for IgG1/2/3/4, IgA and IgM analysis; an ELISA was used for IgE. Plasma samples from de-identified patients were used to compare the Luminex assay with nephelometry, which is routinely used to quantify IgA, IgG and IgM in clinical samples. The IgE ELISA was compared to an immunofluorescence assay. Prior to evaluation of punches from newborn dried blood spots (NDBSs), recoveries of Ig from punches of cord blood DBSs (CBDBSs) vs. plasma from the same cord bloods were compared. Although the recoveries of Ig from plasma and DBSs were not comparable, which could be due to cell lysates in the DBS samples, the analyses were reproducible. Additionally, the levels of IgA, IgG2, IgG4, and IgM recovered from CBDBSs positively correlated with those in plasma. The DBS data is a relative value since it is not equivalent to the plasma concentration. The majority of Ig concentrations recovered from 108 newborns of the Upstate KIDs Study were within the range of newborn plasma Ig levels with the exception of IgG3. The IgG4 values displayed the greatest variance with a wide range (0.01-319 mg/dl), whereas, IgG1 values had the narrowest range (85.2-960.4 mg/dl). PMID:24333851

  4. Improved Diagnosis of Prosthetic Joint Infection by Culturing Periprosthetic Tissue Specimens in Blood Culture Bottles

    PubMed Central

    Peel, Trisha N.; Dylla, Brenda L.; Hughes, John G.; Lynch, David T.; Greenwood-Quaintance, Kerryl E.; Cheng, Allen C.; Mandrekar, Jayawant N.

    2016-01-01

    ABSTRACT Despite known low sensitivity, culture of periprosthetic tissue specimens on agars and in broths is routine. Culture of periprosthetic tissue samples in blood culture bottles (BCBs) is potentially more convenient, but it has been evaluated in a limited way and has not been widely adopted. The aim of this study was to compare the sensitivity and specificity of inoculation of periprosthetic tissue specimens into blood culture bottles with standard agar and thioglycolate broth culture, applying Bayesian latent class modeling (LCM) in addition to applying the Infectious Diseases Society of America (IDSA) criteria for prosthetic joint infection. This prospective cohort study was conducted over a 9-month period (August 2013 to April 2014) at the Mayo Clinic, Rochester, MN, and included all consecutive patients undergoing revision arthroplasty. Overall, 369 subjects were studied; 117 (32%) met IDSA criteria for prosthetic joint infection, and 82% had late chronic infection. Applying LCM, inoculation of tissues into BCBs was associated with a 47% improvement in sensitivity compared to the sensitivity of conventional agar and broth cultures (92.1 versus 62.6%, respectively); this magnitude of change was similar when IDSA criteria were applied (60.7 versus 44.4%, respectively; P = 0.003). The time to microorganism detection was shorter with BCBs than with standard media (P < 0.0001), with aerobic and anaerobic BCBs yielding positive results within a median of 21 and 23 h, respectively. Results of our study demonstrate that the semiautomated method of periprosthetic tissue culture in blood culture bottles is more sensitive than and as specific as agar and thioglycolate broth cultures and yields results faster. PMID:26733067

  5. The use of mass spectrometry to analyze dried blood spots.

    PubMed

    Wagner, Michel; Tonoli, David; Varesio, Emmanuel; Hopfgartner, Gérard

    2016-01-01

    Dried blood spots (DBS) typically consist in the deposition of small volumes of capillary blood onto dedicated paper cards. Comparatively to whole blood or plasma samples, their benefits rely in the fact that sample collection is easier and that logistic aspects related to sample storage and shipment can be relatively limited, respectively, without the need of a refrigerator or dry ice. Originally, this approach has been developed in the sixties to support the analysis of phenylalanine for the detection of phenylketonuria in newborns using bacterial inhibition test. In the nineties tandem mass spectrometry was established as the detection technique for phenylalanine and tyrosine. DBS became rapidly recognized for their clinical value: they were widely implemented in pediatric settings with mass spectrometric detection, and were closely associated to the debut of newborn screening (NBS) programs, as a part of public health policies. Since then, sample collection on paper cards has been explored with various analytical techniques in other areas more or less successfully regarding large-scale applications. Moreover, in the last 5 years a regain of interest for DBS was observed and originated from the bioanalytical community to support drug development (e.g., PK studies) or therapeutic drug monitoring mainly. Those recent applications were essentially driven by improved sensitivity of triple quadrupole mass spectrometers. This review presents an overall view of all instrumental and methodological developments for DBS analysis with mass spectrometric detection, with and without separation techniques. A general introduction to DBS will describe their advantages and historical aspects of their emergence. A second section will focus on blood collection, with a strong emphasis on specific parameters that can impact quantitative analysis, including chromatographic effects, hematocrit effects, blood effects, and analyte stability. A third part of the review is dedicated to

  6. Application of dried blood spot cards to determine olive oil phenols (hydroxytyrosol metabolites) in human blood.

    PubMed

    de Las Hazas, María Carmen López; Motilva, Maria José; Piñol, Carme; Macià, Alba

    2016-10-01

    In this study, a fast and simple blood sampling and sample pre-treatment method based on the use of the dried blood spot (DBS) cards and ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) for the quantification of olive oil phenolic metabolites in human blood was developed and validated. After validation, the method was applied to determine hydroxytyrosol metabolites in human blood samples after the acute intake of an olive oil phenolic extract. Using the FTA DMPK-A DBS card under optimum conditions, with 20µL as the blood solution volume, 100µL of methanol/Milli-Q water (50/50, v/v) as the extraction solvent and 7 disks punched out from the card, the main hydroxytyrosol metabolites (hydroxytyrosol-3-O-sulphate and hydroxytyrosol acetate sulphate) were identified and quantified. The developed methodology allowed detecting and quantifying the generated metabolites at low μM levels. The proposed method is a significant improvement over existing methods to determine phenolic metabolites circulating in blood and plasma samples, thus making blood sampling possible with the volunteer pricking their own finger, and the subsequent storage of the blood in the DBS cards prior to chromatographic analysis. PMID:27474297

  7. Measurement and Comparison of Organic Compound Concentrations in Plasma, Whole Blood, and Dried Blood Spot Samples

    PubMed Central

    Batterman, Stuart A.; Chernyak, Sergey; Su, Feng-Chiao

    2016-01-01

    The preferred sampling medium for measuring human exposures of persistent organic compounds (POPs) is blood, and relevant sample types include whole blood, plasma, and dried blood spots (DBS). Because information regarding the performance and comparability of measurements across these sample types is limited, it is difficult to compare across studies. This study evaluates the performance of POP measurements in plasma, whole blood and DBS, and presents the distribution coefficients needed to convert concentrations among the three sample types. Blood samples were collected from adult volunteers, along with demographic and smoking information, and analyzed by GC/MS for organochlorine pesticides (OCPs), chlorinated hydrocarbons (CHCs), polychlorinated biphenyls (PCBs), and brominated diphenyl ethers (PBDEs). Regression models were used to evaluate the relationships between the sample types and possible effects of personal covariates. Distribution coefficients also were calculated using physically-based models. Across all compounds, concentrations in plasma were consistently the highest; concentrations in whole blood and DBS samples were comparable. Distribution coefficients for plasma to whole blood concentrations ranged from 1.74 to 2.26 for pesticides/CHCs, averaged 1.69 ± 0.06 for the PCBs, and averaged 1.65 ± 0.03 for the PBDEs. Regression models closely fit most chemicals (R2 > 0.80), and whole blood and DBS samples generally showed very good agreement. Distribution coefficients estimated using biologically-based models were near one and did not explain the observed distribution. Among the study population, median concentrations of several pesticides/CHCs and PBDEs exceeded levels reported in the 2007–2008 National Health and Nutrition Examination Survey, while levels of other OCPs and PBDEs were comparable or lower. Race and smoking status appeared to slightly affect plasma/blood concentration ratios for several POPs. The experimentally

  8. X-ray fluorescence analysis of Mexican varieties of dried chili peppers II: Commercial and home-grown specimens

    NASA Astrophysics Data System (ADS)

    Romero-Dávila, E.; Miranda, J.; Pineda, J. C.

    2015-07-01

    Elemental analyses of samples of Mexican varieties of dried chili peppers were carried out using X-ray Fluorescence (XRF). Several specimens of Capsicum annuum L., Capsicum chinense, and Capsicum pubescens were analyzed and the results compared to previous studies of elemental contents in other varieties of Capsicum annuum (ancho, morita, chilpotle, guajillo, pasilla, and árbol). The first set of samples was bought packaged in markets. In the present work, the study focuses on home-grown samples of the árbol and chilpotle varieties, commercial habanero (Capsicum chinense), as well as commercial and home-grown specimens of manzano (Capsicum pubescencs). Samples were freeze dried and pelletized. XRF analyses were carried out using a spectrometer based on an Rh X-ray tube, using a Si-PIN detector. The system detection calibration was performed through the analysis of the NIST certified reference materials 1547 (peach leaves) and 1574 (tomato leaves), while accuracy was checked with the reference material 1571 (orchard leaves). Elemental contents of all elements in the new set of samples were similar to those of the first group. Nevertheless, it was found that commercial samples contain high amounts of Br, while home-grown varieties do not.

  9. High Quality Genome-Wide Genotyping from Archived Dried Blood Spots without DNA Amplification

    PubMed Central

    St. Julien, Krystal R.; Jelliffe-Pawlowski, Laura L.; Shaw, Gary M.; Stevenson, David K.; O’Brodovich, Hugh M.; Krasnow, Mark A.

    2013-01-01

    Spots of blood are routinely collected from newborn babies onto filter paper called Guthrie cards and used to screen for metabolic and genetic disorders. The archived dried blood spots are an important and precious resource for genomic research. Whole genome amplification of dried blood spot DNA has been used to provide DNA for genome-wide SNP genotyping. Here we describe a 96 well format procedure to extract DNA from a portion of a dried blood spot that provides sufficient unamplified genomic DNA for genome-wide single nucleotide polymorphism (SNP) genotyping. We show that SNP genotyping of the unamplified DNA is more robust than genotyping amplified dried blood spot DNA, is comparable in cost, and can be done with thousands of samples. This procedure can be used for genome-wide association studies and other large-scale genomic analyses that require robust, high-accuracy genotyping of dried blood spot DNA. PMID:23737996

  10. Validation and Application of a Dried Blood Spot Ceftriaxone Assay

    PubMed Central

    Page-Sharp, Madhu; Nunn, Troy; Salman, Sam; Moore, Brioni R.; Batty, Kevin T.; Davis, Timothy M. E.

    2015-01-01

    Dried blood spot (DBS) antibiotic assays can facilitate pharmacokinetic/pharmacodynamic (PK/PD) studies in situations where venous blood sampling is logistically and/or ethically problematic. In this study, we aimed to develop, validate, and apply a DBS ceftriaxone assay. A liquid chromatography-tandem mass spectroscopy (LC-MS/MS) DBS ceftriaxone assay was assessed for matrix effects, process efficiency, recovery, variability, and limits of quantification (LOQ) and detection (LOD). The effects of hematocrit, protein binding, red cell partitioning, and chad positioning were evaluated, and thermal stability was assessed. Plasma, DBS, and cell pellet ceftriaxone concentrations in 10 healthy adults were compared, and plasma concentration-time profiles of DBS and plasma ceftriaxone were incorporated into population PK models. The LOQ and LOD for ceftriaxone in DBS were 0.14 mg/liter and 0.05 mg/liter, respectively. Adjusting for hematocrit, red cell partitioning, and relative recovery, DBS-predicted plasma concentrations were comparable to measured plasma concentrations (r > 0.95, P < 0.0001), and Bland-Altman plots showed no significant bias. The final population PK estimates of clearance, volume of distribution, and time above threshold MICs for measured and DBS-predicted plasma concentrations were similar. At 35°C, 21°C, 4°C, −20°C, and −80°C, ceftriaxone retained >95% initial concentrations in DBS for 14 h, 35 h, 30 days, 21 weeks, and >11 months, respectively. The present DBS ceftriaxone assay is robust and can be used as a surrogate for plasma concentrations to provide valid PK and PK/PD data in a variety of clinical situations, including in studies of young children and of those in remote or resource-poor settings. PMID:26438505

  11. Enhanced Diagnostic Yields of Bacteremia and Candidemia in Blood Specimens by PCR-Electrospray Ionization Mass Spectrometry

    PubMed Central

    Laffler, Thomas G.; Cummins, Lendell L.; McClain, Colt M.; Quinn, Criziel D.; Toro, Michelle A.; Carolan, Heather E.; Toleno, Donna M.; Rounds, Megan A.; Eshoo, Mark W.; Stratton, Charles W.; Sampath, Rangarajan; Blyn, Lawrence B.; Ecker, David J.

    2013-01-01

    A prospective study was performed to determine the value of direct molecular testing of whole blood for detecting the presence of culturable and unculturable bacteria and yeasts in patients with suspected bloodstream infections. A total of 464 adult and pediatric patients with positive blood cultures matched with 442 patients with negative blood cultures collected during the same period were recruited during a 10-month study. PCR amplification coupled with electrospray ionization mass spectrometry (PCR-ESI-MS) plus blood culture reached an overall agreement of 78.6% in the detection and species-level identification of bacterial and candidal pathogens. Of 33 culture-negative/PCR-ESI-MS-positive specimens, 31 (93.9%) were judged to be truly bacteremic and/or candidemic based on a medical chart review and analytical metrics. Among the 15 culture-positive specimens in which PCR-ESI-MS detected additional bacterial or yeast species, 66.7% and 20.0% of the additional positive specimens by PCR-ESI-MS were judged to be truly or possibly bacteremic and/or candidemic, respectively. Direct analysis of blood samples by PCR-ESI-MS rapidly detects bacterial and yeast pathogens in patients with bloodstream infections. When used in conjunction with blood culture, PCR-ESI-MS enhances the diagnostics of septicemia by shortening test turnaround time and improving yields. PMID:23966503

  12. Simultaneous determination of cocaine and opiates in dried blood spots by electrospray ionization tandem mass spectrometry.

    PubMed

    Antelo-Domínguez, Ángel; Cocho, José Ángel; Tabernero, María Jesús; Bermejo, Ana María; Bermejo-Barrera, Pilar; Moreda-Piñeiro, Antonio

    2013-12-15

    A sample pre-treatment method based on blood spot collection filter cards was optimized as a means of using small volume samples for the screening and confirmation of cocaine and opiates abuse. Dried blood spots (DBSs) were prepared by dispersing 20 µL of whole blood specimens previously mixed with the internal standards (deuterated analogs of each target), and subjecting the whole DBS to extraction with 5 mL of methanol under orbital-horizontal shaking (180 rpm) for 10 min. Determinations were based on direct electrospray ionization tandem mass spectrometry (ESI-MS/MS) by injecting the re-dissolved methanol extract with the delivery solution (acetonitrile-water-formic acid, 80:19.875:0.125) at a flow rate of 60 µL min(-1), and using multiple reaction monitoring (MRM) mode with the m/z (precursor ion)→m/z (product ion) transitions for acquisition. Matrix effect has been found to be statistically significant (Multiple Range Test) when assessing cocaine, BZE, codeine and morphine, and the use of the standard addition method (dispersion of whole blood previously mixed with standards onto the filter papers) was needed for accurate determinations. The developed DBS-ESI-MS/MS procedure offered good intra-day and inter-day precisions (lower than 10% and 12%, respectively), as well as good intra-day and inter-day accuracies (inter-day absolute recoveries, expressed as the mean analytical recovery over three target concentration levels, of 103%, 100%, 101%, 98% and 100% for cocaine, BZE, codeine, morphine and 6-MAM, respectively). The high sensitivity inherent to MS/MS determinations combined with the minimal dilution of sample allowed low limits of quantification for all targets, and the developed method results therefore adequate for cocaine and opiates screening and confirmation purposes. The procedure was finally applied to DBSs prepared from whole blood from polydrug abusers, and results were compared with those obtained after a conventional sample pretreatment

  13. Clinical Evaluation of a Dried Blood Spot Assay for Atazanavir▿

    PubMed Central

    Van Schooneveld, Trevor; Swindells, Susan; Nelson, Sarah R.; Robbins, Brian L.; Moore, Ryan; Fletcher, Courtney V.

    2010-01-01

    Current procedures for obtaining and measuring plasma concentrations of HIV protease inhibitors (PIs) are technically challenging. Dried blood spot (DBS) assays offer a way to overcome many of the obstacles. We sought to develop a DBS assay for quantitation of the PI atazanavir (ATV) and to compare this method with a previously validated plasma assay. We prospectively enrolled 48 patients with well-controlled HIV disease who had been on ATV for at least 7 days. ATV was quantified from plasma by use of high-performance liquid chromatography (HPLC). A reversed-phase ultrahigh-performance liquid chromatography (UPLC) assay was utilized for DBS samples. The concentrations of ATV quantified in a DBS matrix showed very strong agreement with those measured in plasma (r2 = 0.988). The mean difference in ATV concentration between the two methods was −10.8% (95% confidence interval [95% CI], −7.65% to −13.95%), indicating that the DBS method has a slight negative bias. A majority (97.8%) of the differences in concentration between the two assays fell within ±2 standard deviations. ATV concentrations were lower in subjects who had detectable HIV RNA in plasma (mean, 543 ng/ml) than in those with HIV RNA of <50 copies/ml (mean, 1,582 ng/ml) (P = 0.03, Wilcoxon rank-sum test). In conclusion, our study demonstrated that ATV quantitation in a DBS matrix is feasible and accurate. DBS use offers a convenient alternative for measuring plasma concentrations of ATV and may have utility in monitoring of drug concentrations in clinical practice and in future studies. PMID:20660680

  14. High-Throughput Immunoassay for the Biochemical Diagnosis of Friedreich Ataxia in Dried Blood Spots and Whole Blood

    PubMed Central

    Oglesbee, Devin; Kroll, Charles; Gakh, Oleksandr; Deutsch, Eric C.; Lynch, David R.; Gavrilova, Ralitza; Tortorelli, Silvia; Raymond, Kimiyo; Gavrilov, Dimitar; Rinaldo, Piero; Matern, Dietrich; Isaya, Grazia

    2014-01-01

    BACKGROUND Friedreich ataxia (FRDA) is caused by reduced frataxin (FXN) concentrations. A clinical diagnosis is typically confirmed by DNA-based assays for GAA-repeat expansions or mutations in the FXN (frataxin) gene; however, these assays are not applicable to therapeutic monitoring and population screening. To facilitate the diagnosis and monitoring of FRDA patients, we developed an immunoassay for measuring FXN. METHODS Antibody pairs were used to capture FXN and an internal control protein, ceruloplasmin (CP), in 15 μL of whole blood (WB) or one 3-mm punch of a dried blood spot (DBS). Samples were assayed on a Luminex LX200 analyzer and validated according to standard criteria. RESULTS The mean recovery of FXN from WB and DBS samples was 99%. Intraassay and interassay imprecision (CV) values were 4.9%–13% and 9.8%–16%, respectively. The FXN limit of detection was 0.07 ng/mL, and the reportable range of concentrations was 2–200 ng/mL. Reference adult and pediatric FXN concentrations ranged from 15 to 82 ng/mL (median, 33 ng/mL) for DBS and WB. The FXN concentration range was 12–22 ng/mL (median, 15 ng/mL) for FRDA carriers and 1–26 ng/mL (median 5 ng/mL) for FRDA patients. Measurement of the FXN/CP ratio increased the ability to distinguish between patients, carriers, and the reference population. CONCLUSIONS This assay is applicable to the diagnosis and therapeutic monitoring of FRDA. This assay can measure FXN and the control protein CP in both WB and DBS specimens with minimal sample requirements, creating the potential for high-throughput population screening of FRDA. PMID:23838345

  15. Mass spectrometry in cancer biomarker research: a case for immunodepletion of abundant blood-derived proteins from clinical tissue specimens

    PubMed Central

    Prieto, DaRue A; Johann, Donald J; Wei, Bih-Rong; Ye, Xiaoying; Chan, King C; Nissley, Dwight V; Simpson, R Mark; Citrin, Deborah E; Mackall, Crystal L; Linehan, W Marston; Blonder, Josip

    2014-01-01

    The discovery of clinically relevant cancer biomarkers using mass spectrometry (MS)-based proteomics has proven difficult, primarily because of the enormous dynamic range of blood-derived protein concentrations and the fact that the 22 most abundant blood-derived proteins constitute approximately 99% of the total plasma protein mass. Immunodepletion of clinical body fluid specimens (e.g., serum/plasma) for the removal of highly abundant proteins is a reasonable and reproducible solution. Often overlooked, clinical tissue specimens also contain a formidable amount of highly abundant blood-derived proteins present in tissue-embedded networks of blood/lymph capillaries and interstitial fluid. Hence, the dynamic range impediment to biomarker discovery remains a formidable obstacle, regardless of clinical sample type (solid tissue and/or body fluid). Thus, we optimized and applied simultaneous immunodepletion of blood-derived proteins from solid tissue and peripheral blood, using clear cell renal cell carcinoma as a model disease. Integrative analysis of data from this approach and genomic data obtained from the same type of tumor revealed concordant key pathways and protein targets germane to clear cell renal cell carcinoma. This includes the activation of the lipogenic pathway characterized by increased expression of adipophilin (PLIN2) along with 'cadherin switching', a phenomenon indicative of transcriptional reprogramming linked to renal epithelial dedifferentiation. We also applied immunodepletion of abundant blood-derived proteins to various tissue types (e.g., adipose tissue and breast tissue) showing unambiguously that the removal of abundant blood-derived proteins represents a powerful tool for the reproducible profiling of tissue proteomes. Herein, we show that the removal of abundant blood-derived proteins from solid tissue specimens is of equal importance to depletion of body fluids and recommend its routine use in the context of biological discovery and

  16. Enhanced Stability of Blood Matrices Using a Dried Sample Spot Assay to Measure Human Butyrylcholinesterase Activity and Nerve Agent Adducts

    PubMed Central

    Perez, Jonas W.; Pantazides, Brooke G.; Watson, Caroline M.; Thomas, Jerry D.; Blake, Thomas A.; Johnson, Rudolph C.

    2015-01-01

    Dried matrix spots are safer to handle and easier to store than wet blood products, but factors such as intra-spot variability and unknown sample volumes have limited their appeal as a sampling format for quantitative analyses. In this work, we introduce a dried spot activity assay for quantifying butyrylcholinesterase (BChE) specific activity which is BChE activity normalized to the total protein content in a sample spot. The method was demonstrated with blood, serum, and plasma spotted on specimen collection devices (cards) which were extracted to measure total protein and BChE activity using a modified Ellman assay. Activity recovered from dried spots was ∼80% of the initial spotted activity for blood and >90% for plasma and serum. Measuring total protein in the sample and calculating specific activity substantially improved quantification and reduced intra-spot variability. Analyte stability of nerve agent adducts was also evaluated, and the results obtained via BChE-specific activity measurements were confirmed by quantification of BChE adducts using a previously established LC-MS/MS method. The spotted samples were up to 10-times more resistant to degradation compared to unspotted control samples when measuring BChE inhibition by the nerve agents sarin and VX. Using this method, both BChE activity and adducts can be accurately measured from a dried sample spot. This use of a dried sample spot with normalization to total protein is robust, demonstrates decreased intra-spot variability without the need to control for initial sample volume, and enhances analyte stability. PMID:25955132

  17. Impact of a large-scale educational intervention program on venous blood specimen collection practices

    PubMed Central

    2013-01-01

    Background Phlebotomy performed with poor adherence to venous blood specimen collection (VBSC) guidelines jeopardizes patient safety and may lead to patient suffering and adverse events. A first questionnaire study demonstrated low compliance to VBSC guidelines, motivating an educational intervention of all phlebotomists within a county council. The aim was to evaluate the impact of a large-scale educational intervention program (EIP) on primary health care phlebotomists’ adherence to VBSC guidelines. We hypothesised that the EIP would improve phlebotomists’ VBSC practical performance. Methods The present study comprise primary health care centres (n = 61) from two county councils in northern Sweden. The final selected study group consisted of phlebotomists divided into an intervention group (n = 84) and a corresponding control group (n = 79). Both groups responded to a validated self-reported VBSC questionnaire twice. The EIP included three parts: guideline studies, an oral presentation, and an examination. Non-parametric statistics were used for comparison within and between the groups. Results Evaluating the EIP, we found significant improvements in the intervention group compared to the control group on self-reported questionnaire responses regarding information search (ES = 0.23-0.33, p < 0.001-0.003), and patient rest prior to phlebotomy (ES = 0.27, p = 0.004). Test request management, patient identity control, release of venous stasis, and test tube labelling had significantly improved in the intervention group but did not significantly differ from the control group (ES = 0.22- 0.49, p = < 0.001- 0.006). The control group showed no significant improvements at all (ES = 0–0.39, p = 0.016-0.961). Conclusions The present study demonstrated several significant improvements on phlebotomists’ adherence to VBSC practices. Still, guideline adherence improvement to several crucial phlebotomy practices is needed. We

  18. Evaluation of Amount of Blood in Dry Blood Spots: Ring-Disk Electrode Conductometry.

    PubMed

    Kadjo, Akinde F; Stamos, Brian N; Shelor, C Phillip; Berg, Jordan M; Blount, Benjamin C; Dasgupta, Purnendu K

    2016-06-21

    A fixed area punch in dried blood spot (DBS) analysis is assumed to contain a fixed amount of blood, but the amount actually depends on a number of factors. The presently preferred approach is to normalize the measurement with respect to the sodium level, measured by atomic spectrometry. Instead of sodium levels, we propose electrical conductivity of the extract as an equivalent nondestructive measure. A dip-type small diameter ring-disk electrode (RDE) is ideal for very small volumes. However, the conductance (G) measured by an RDE depends on the depth (D) of the liquid below the probe. There is no established way of computing the specific conductance (σ) of the solution from G. Using a COMSOL Multiphysics model, we were able to obtain excellent agreement between the measured and the model predicted conductance as a function of D. Using simulations over a large range of dimensions, we provide a spreadsheet-based calculator where the RDE dimensions are the input parameters and the procedure determines the 99% of the infinite depth conductance (G99) and the depth D99 at which this is reached. For typical small diameter probes (outer electrode diameter ∼ <2 mm), D99 is small enough for dip-type measurements in extract volumes of ∼100 μL. We demonstrate the use of such probes with DBS extracts. In a small group of 12 volunteers (age 20-66), the specific conductance of 100 μL aqueous extracts of 2 μL of spotted blood showed a variance of 17.9%. For a given subject, methanol extracts of DBS spots nominally containing 8 and 4 μL of blood differed by a factor of 1.8-1.9 in the chromatographically determined values of sulfate and chloride (a minor and major constituent, respectively). The values normalized with respect to the conductance of the extracts differed by ∼1%. For serum associated analytes, normalization of the analyte value by the extract conductance can thus greatly reduce errors from variations in the spotted blood volume/unit area. PMID:27226021

  19. Quantitation of 5-Methyltetrahydrofolic Acid in Dried Blood Spots and Dried Plasma Spots by Stable Isotope Dilution Assays

    PubMed Central

    Kopp, Markus; Rychlik, Michael

    2015-01-01

    Because of minimal data available on folate analysis in dried matrix spots (DMSs), we combined the advantages of stable isotope dilution assays followed by LC-MS/MS analysis with DMS sampling to develop a reliable method for the quantitation of plasma 5-methyltetrahydrofolic acid in dried blood spots (DBSs) and dried plasma spots (DPSs) as well as for the quantitation of whole blood 5-methyltetrahydrofolic acid in DBSs. We focused on two diagnostically conclusive parameters exhibited by the plasma and whole blood 5-methyltetrahydrofolic acid levels that reflect both temporary and long-term folate status. The method is performed using the [2H4]-labeled isotopologue of the vitamin as the internal standard, and three steps are required for the extraction procedure. Elution of the punched out matrix spots was performed using stabilization buffer including Triton X-100 in a standardized ultrasonication treatment followed by enzymatic digestion (whole blood only) and solid-phase extraction with SAX cartridges. This method is sensitive enough to quantify 27 nmol/L whole blood 5-methyltetrahydrofolic acid in DBSs and 6.3 and 4.4 nmol/L plasma 5-methyltetrahydrofolic acid in DBSs and DPSs, respectively. The unprecedented accurate quantification of plasma 5-methyltetrahydrofolic acid in DBSs was achieved by thermal treatment prior to ultrasonication, inhibiting plasma conjugase activity. Mass screenings are more feasible and easier to facilitate for this method in terms of sample collection and storage compared with conventional clinical sampling for the assessment of folate status. PMID:26605791

  20. Potential of Dried Blood Self-Sampling for Cyclosporine C2 Monitoring in Transplant Outpatients

    PubMed Central

    Leichtle, Alexander Benedikt; Ceglarek, Uta; Witzigmann, Helmut; Gäbel, Gábor; Thiery, Joachim; Fiedler, Georg Martin

    2010-01-01

    Background. Close therapeutic drug monitoring of Cyclosporine (CsA) in transplant outpatients is a favourable procedure to maintain the long-term blood drug levels within their respective narrow therapeutic ranges. Compared to basal levels (C0), CsA peak levels (C2) are more predictive for transplant rejection. However, the application of C2 levels is hampered by the precise time of blood sampling and the need of qualified personnel. Therefore, we evaluated a new C2 self-obtained blood sampling in transplant outpatients using dried capillary and venous blood samples and compared the CsA levels, stability, and clinical practicability of the different procedures. Methods. 55 solid organ transplant recipients were instructed to use single-handed sampling of each 50 μL capillary blood and dried blood spots by finger prick using standard finger prick devices. We used standardized EDTA-coated capillary blood collection systems and standardized filter paper WS 903. CsA was determined by LC-MS/MS. The patients and technicians also answered a questionnaire on the procedure and sample quality. Results. The C0 and C2 levels from capillary blood collection systems (C0 [ng/mL]: 114.5 ± 44.5; C2: 578.2 ± 222.2) and capillary dried blood (C0 [ng/mL]: 175.4 ± 137.7; C2: 743.1 ± 368.1) significantly (P < .01) correlated with the drug levels of the venous blood samples (C0 [ng/mL]: 97.8 ± 37.4; C2: 511.2 ± 201.5). The correlation at C0 was ρcap.-ven. = 0.749, and ρdried blood-ven = 0.432; at C2: ρcap.-ven. = 0.861 and ρdried blood-ven = 0.711. The patients preferred the dried blood sampling because of the more simple and less painful procedure. Additionally, the sample quality of self-obtained dried blood spots for LC-MS/MS analytics was superior to the respective capillary blood samples. Conclusions. C2 self-obtained dried blood sampling can easily be performed by transplant outpatients and is therefore suitable and cost-effective for close therapeutic drug monitoring

  1. Automatic disease screening method using image processing for dried blood microfluidic drop stain pattern recognition.

    PubMed

    Sikarwar, Basant S; Roy, Mukesh; Ranjan, Priya; Goyal, Ayush

    2016-07-01

    This paper examines programmed automatic recognition of infection from samples of dried stains of micro-scale drops of patient blood. This technique has the upside of being low-cost and less-intrusive and not requiring puncturing the patient with a needle for drawing blood, which is especially critical for infants and the matured. It also does not require expensive pathological blood test laboratory equipment. The method is shown in this work to be successful for ailment identification in patients suffering from tuberculosis and anaemia. Illness affects the physical properties of blood, which thus influence the samples of dried micro-scale blood drop stains. For instance, if a patient has a severe drop in platelet count, which is often the case of dengue or malaria patients, the blood's physical property of viscosity drops substantially, i.e. the blood is thinner. Thus, the blood micro-scale drop stain samples can be utilised for diagnosing maladies. This paper presents programmed automatic examination of the dried micro-scale drop blood stain designs utilising an algorithm based on pattern recognition. The samples of micro-scale blood drop stains of ordinary non-infected people are clearly recognisable as well as the samples of micro-scale blood drop stains of sick people, due to key distinguishing features. As a contextual analysis, the micro-scale blood drop stains of patients infected with tuberculosis have been contrasted with the micro-scale blood drop stains of typical normal healthy people. The paper dives into the fundamental flow mechanics behind how the samples of the dried micro-scale blood drop stain is shaped. What has been found is a thick ring like feature in the dried micro-scale blood drop stains of non-ailing people and thin shape like lines in the dried micro-scale blood drop stains of patients with anaemia or tuberculosis disease. The ring like feature at the periphery is caused by an outward stream conveying suspended particles to the edge

  2. [The use of dried blood spots for HIV-antibody testing in Sahel].

    PubMed

    Ouwe-Missi-Oukem-Boyer, O N; Hamidou, A A; Sidikou, F; Garba, A; Louboutin-Croc, J P

    2005-12-01

    Undertaking a HIV seroepidemiological survey in Sahel is logistically problematic, since countries like Niger or Mali are very large with scattered populations and harsh climatic conditions. Therefore, the replacement of serum samples by whole blood dried on filter papers has been studied for HIV-antibody testing with commercial kits that are commonly used. In Niger, two tests ELISA (Genscreen HIV1/2 version 2, Vironostika HIV Uni-Form II Ag/Ab) and two rapid tests (Determine HIV1/2 et Immunocomb II HIV1&2 Bispot) were used to compare the dried blood spots and serum samples from 43 control individuals. Both ELISAs gave an excellent correlation (r = 0.99 et r = 0.98) between the dried blood spots and serum absorbance values. Using the rapid tests, the HIV status was found 100% concordant with dried blood spots and serum samples. An algorithm using three out of the four mentioned tests was defined then validated on the dried blood spots of 163 control individuals (100% concordant). In conclusion, dried blood spots may accurately and profitably replace serum samples for the serodiagnosis of HIV infection and for mass serosurveys in Sahel. PMID:16425709

  3. Active tracking of rejected dried blood samples in a large program in Nigeria

    PubMed Central

    Inalegwu, Auchi; Phillips, Sunny; Datir, Rawlings; Chime, Christopher; Ozumba, Petronilla; Peters, Samuel; Ogbanufe, Obinna; Mensah, Charles; Abimiku, Alash’Le; Dakum, Patrick; Ndembi, Nicaise

    2016-01-01

    AIM: To study the impact of rejection at different levels of health care by retrospectively reviewing records of dried blood spot samples received at the molecular laboratory for human immunodeficiency virus (HIV) early infant diagnosis (EID) between January 2008 and December 2012. METHODS: The specimen rejection rate, reasons for rejection and the impact of rejection at different levels of health care was examined. The extracted data were cleaned and checked for consistency and then de-duplicated using the unique patient and clinic identifiers. The cleaned data were ciphered and exported to SPSS version 19 (SPSS 2010 IBM Corp, New York, United States) for statistical analyses. RESULTS: Sample rejection rate of 2.4% (n = 786/32552) and repeat rate of 8.8% (n = 69/786) were established. The mean age of infants presenting for first HIV molecular test among accepted valid samples was 17.83 wk (95%CI: 17.65-18.01) vs 20.30 wk (95%CI: 16.53-24.06) for repeated samples. HIV infection rate was 9.8% vs 15.9% for accepted and repeated samples. Compared to tertiary healthcare clinics, secondary and primary clinics had two-fold and three-fold higher likelihood of sample rejection, respectively (P < 0.05). We observed a significant increase in sample rejection rate with increasing number of EID clinics (r = 0.893, P = 0.041). The major reasons for rejection were improper sample collection (26.3%), improper labeling (16.4%) and insufficient blood (14.8%). CONCLUSION: Programs should monitor pre-analytical variables and incorporate continuous quality improvement interventions to reduce errors associated with sample rejection and improve patient retention. PMID:27175352

  4. Occurrence of ethanol and other drugs in blood and urine specimens from female victims of alleged sexual assault.

    PubMed

    Jones, Alan Wayne; Kugelberg, Fredrik C; Holmgren, Anita; Ahlner, Johan

    2008-10-25

    Results of toxicological analysis of blood and urine specimens from 1806 female victims of alleged non-consensual sexual activity are reported. After making contact with the police authorities, the victims were examined by a physician for injuries and biological specimens were taken for forensic toxicology and other purposes (e.g. DNA). Urine if available or otherwise on an aliquot of blood after protein precipitation was screened for the presence of drugs by enzyme immunoassay methods (EMIT/CEDIA). All positive results from screening were verified by more specific methods, involving isotope dilution gas chromatography-mass spectrometry (GC-MS) for illicit drugs. A large number of prescription drugs were analyzed in blood by capillary column gas chromatography with a nitrogen-phosphorous (N-P) detector. Ethanol was determined in blood and urine by headspace gas chromatography and concentrations less than 0.1g/L were reported as negative. The number of reported cases of alleged sexual assault was highest during the warmer summer months and the mean age of victims was 24 years (median 20 years), with approximately 60% being between 15 and 25 years. In 559 cases (31%) ethanol and drugs were negative. In 772 cases (43% of total) ethanol was the only drug identified in blood or urine. In 215 cases (12%) ethanol occurred together with at least one other drug. The mean, median and highest concentrations of ethanol in blood (N=806) were 1.24 g/L, 1.19 g/L and 3.7 g/L, respectively. The age of victims and their blood-alcohol concentration (BAC) were positively correlated (r=0.365, p<0.001). Because BAC decreases at a rate of 0.10-0.25 g/(Lh), owing to metabolism the concentration in blood at time of sampling is often appreciably less than when the crime was committed several hours earlier. Licit or illicit drugs were identified in blood or urine in N=262 cases (15%). Amphetamine and tetrahydrocannabinol were the most common illicit drugs at mean (median) concentrations in

  5. Buprenorphine and major metabolites in blood specimens collected for drug analysis in law enforcement purposes.

    PubMed

    Oechsler, Stephanie; Skopp, Gisela

    2010-02-25

    A liquid chromatographic/electrospray ionization tandem mass spectrometric method for the quantification of buprenorphine (BUP), norbuprenorphine (NBUP), buprenorphine-3-beta-D-glucuronide (BUPG) and norbuprenorphine-3-beta-D-glucuronide (NBUPG) in serum samples was developed and validated. Pre-treatment of BUP and NBUP was by liquid-liquid extraction, while glucuronides were favourably isolated by solid phase extraction. Separation in 2 separate runs (2 x 5 min) was achieved using isocratic elution. The method was applied to 20 authentic serum specimens collected for law enforcement purposes where BUP intake had been indicated. The parent drug was not detectable in half of the specimens at a lower limit of detection of 0.2 ng/mL, whereas NBUP could be determined from any sample but one. NBUPG is the major metabolite present, which could be identified along with BUPG in all samples under investigation. In authentic specimens it could be advisable to monitor BUP metabolites along with the parent drug. PMID:20006453

  6. Dried-Blood Spots: A Cost-Effective Field Method for the Detection of Chikungunya Virus Circulation in Remote Areas

    PubMed Central

    Randrianasolo, Laurence; Rafisandratantsoa, Jean Théophile; Andriamamonjy, Seta; Richard, Vincent

    2013-01-01

    Background In 2005, there were outbreaks of febrile polyarthritis due to Chikungunya virus (CHIKV) in the Comoros Islands. CHIKV then spread to other islands in the Indian Ocean: La Réunion, Mauritius, Seychelles and Madagascar. These outbreaks revealed the lack of surveillance and preparedness of Madagascar and other countries. Thus, it was decided in 2007 to establish a syndrome-based surveillance network to monitor dengue-like illness. Objective This study aims to evaluate the use of capillary blood samples blotted on filter papers for molecular diagnosis of CHIKV infection. Venous blood samples can be difficult to obtain and the shipment of serum in appropriate temperature conditions is too costly for most developing countries. Methodology and principal findings Venous blood and dried-blood blotted on filter paper (DBFP) were collected during the last CHIKV outbreak in Madagascar (2010) and as part of our routine surveillance of dengue-like illness. All samples were tested by real-time RT-PCR and results with serum and DBFP samples were compared for each patient. The sensitivity and specificity of tests performed with DBFP, relative to those with venous samples (defined as 100%) were 93.1% (95% CI:[84.7–97.7]) and 94.4% (95% CI:[88.3–97.7]), respectively. The Kappa coefficient 0.87 (95% CI:[0.80–0.94]) was excellent. Conclusion This study shows that DBFP specimens can be used as a cost-effective alternative sampling method for the surveillance and monitoring of CHIKV circulation and emergence in developing countries, and probably also for other arboviruses. The loss of sensitivity is insignificant and involved a very small number of patients, all with low viral loads. Whether viruses can be isolated from dried blood spots remains to be determined. PMID:23936570

  7. Analysis of the use of dried blood spot measurements in disease screening.

    PubMed

    Lakshmy, Ramakrishnan

    2008-03-01

    The collection of dried blood spots on filter paper offers a powerful tool in screening programs and in large population-based surveys. The method has the advantage of being less invasive and relatively painless and is particularly suitable for collection in neonates and the elderly. Blood can be collected and transported economically without requiring a cold chain. The use of blood spots for the measurement of insulin, high sensitivity C-reactive protein, and triglycerides is reported. A good correlation between measurement of these analytes in dried blood and sera suggests that the method is valid and has the potential to be used for the screening of cardiometabolic risk factors. This method of blood collection is particularly suited for developing countries where cost cutting is important. PMID:19885349

  8. Hairy Cell Leukemia with Systemic Lymphadenopathy: Detection of BRAF Mutations in Both Lymph Node and Peripheral Blood Specimens.

    PubMed

    Okada, Kazuya; Kunitomi, Akane; Sakai, Kazuya; Muranushi, Hiroyuki; Okamoto, Yusuke; Tsukamoto, Taku; Sugiura, Hiroyuki; Matsui, Hiroyuki; Jo, Tomoyasu; Ueda, Tomoaki; Onishi, Tatsuhito; Ide, Masaru; Kimura, Shinya; Notohara, Kenji; Ueda, Yasunori

    2015-01-01

    A 47-year-old woman with pancytopenia, excessive systemic lymphadenopathy and splenomegaly was referred to our hospital. The peripheral blood (PB) smear findings indicated neutropenia with lymphoid cells exhibiting hairy projections, while the histological findings of the cervical lymph node (LN) suggested hairy cell leukemia (HCL). In addition, the BRAF V600E mutation was detected, and the immunoglobulin gene rearrangement patterns were identical in both the cervical LN and PB specimens. Based on these findings, we diagnosed the patient with systemic lymphadenopathy due to HCL. This is the first report of a BRAF mutation detected in both the PB and LN at the onset of HCL. PMID:26027995

  9. Quantitation of pregabalin in dried blood spots and dried plasma spots by validated LC-MS/MS methods.

    PubMed

    Kostić, Nađa; Dotsikas, Yannis; Jović, Nebojša; Stevanović, Galina; Malenović, Anđelija; Medenica, Mirjana

    2015-05-10

    In this paper, novel LC-MS/MS methods for the determination of antiepileptic drug pregabalin in dried matrix spots (DMS) are presented. This attractive technique of sample collection in micro amount was utilized in the form of dried blood spots (DBS) and dried plasma spots (DPS). Following a pre-column derivatization procedure, using n-propyl chloroformate in the presence of n-propanol, and consecutive liquid-liquid extraction, derivatized pregabalin and its internal standard, 4-aminocyclohexanecarboxylic acid, were detected in positive ion mode by applying two SRM transitions per analyte. A YMC-Pack Octyl column (50mm×4.0mm, 3μm particle size) maintained at 30°C, was utilized with running mobile phase composed of acetonitrile: 0.15% formic acid (85:15, v/v). Flow rate was 550μL/min and total run time 2min. Established methods were fully validated over the concentration range of 0.200-20.0μg/mL for DBS and 0.400-40.0μg/mL for DPS, respectively, while specificity, accuracy, precision, recovery, matrix-effect, stability, dilution integrity and spot homogeneity were found within acceptance criteria. Validated methods were applied for the determination of pregabalin levels in dried blood and plasma samples obtained from patients with epilepsy, after per os administration of commercial capsules. Comparison of drug level in blood and plasma, as well as correction steps undertaken in order to overcome hematocrit issue, when analyzing DBS, are also given. PMID:25767905

  10. RESIDUES AND METABOLITES OF SELECTED PERSISTENT HALOGENATED HYDROCARBONS IN BLOOD SPECIMENS FROM A GENERAL POPULATION SURVEY

    EPA Science Inventory

    The National Center for Health Statistics collaborated with the National Human Monitoring Program of the U.S. Environmental Protection Agency (EPA) in a four-year study to assess the exposure of the general population to selected pesticides through analysis of blood serum and uri...

  11. The feasibility and cost of neonatal screening for prenatal alcohol exposure by measuring phosphatidylethanol in dried blood spots

    PubMed Central

    Bakhireva, Ludmila N.; Savich, Renate D.; Raisch, Dennis W.; Cano, Sandra; Annett, Robert D.; Leeman, Lawrence; Garg, Mahek; Goff, Chelsea; Savage, Daniel D.

    2012-01-01

    Background Accurate confirmation of prenatal alcohol exposure (PAE) is required as a diagnostic criterion for the majority of children adversely affected by PAE who do not manifest the physical features associated with Fetal Alcohol Syndrome. A number of ethanol biomarkers have been used to assess PAE, often with suboptimal results. The purpose of this study was to evaluate the feasibility and cost of PAE screening in newborns by measuring phosphatidylethanol (PEth) in dried blood spot (DBS) cards. Methods The feasibility of collecting an additional DBS card during routine newborn screening and the background prevalence of PAE were evaluated in a de-identified sample of newborn children delivered at the University of New Mexico Hospital. Electronic orders to collect DBS cards from newborns who continue to bleed after the routine newborn screen, glucose or hematocrit testing were initiated for all infants delivered during a 4-week timeframe. Specimens were sent to a contract laboratory for PEth analysis by liquid chromatography-tandem mass spectrometry. A cost analysis was conducted to compare the cost of PAE screening by PEth in DBS vs. PEth in conventional blood specimens and by meconium fatty acid ethyl esters (FAEE). Results From 230 collected cards, 201 (87.4%) had at least one full blood spot (amount sufficient for PEth analysis), and 6.5% had PEth >20ng/mL indicative of potential PAE in late pregnancy. PAE screening by PEth in DBS is logistically simpler and less expensive compared to two other screening approaches. Conclusions These results indicate that screening for PAE in DBS cards is a feasible procedure and that a majority of infants have enough blood after the routine heel prick to fill an additional card. Moreover, screening by PEth analysis from DBS cards is cost-efficient. The acceptability of such screening by parents and corresponding ethical issues remain to be investigated. PMID:23421919

  12. Forward and inverse dielectric modeling of oven-dried cement paste specimens in the frequency range of 1.02 GHz to 4.50 GHz

    NASA Astrophysics Data System (ADS)

    Owusu Twumasi, Jones; Yu, Tzuyang

    2015-04-01

    The use of radar non-destructive evaluation (NDE) technique for condition assessment of deteriorated civil infrastructure systems is an effective approach for preserving the sustainability of these systems. Radar NDE utilizes the interaction between radar signals (electromagnetic waves) and construction materials for surface and subsurface sensing based on dielectric properties and geometry. In the success of radar inspection, it is imperative to develop models capable of predicting the dielectric properties of the materials under investigation. The dielectric properties (dielectric constant and loss factor) of oven-dried cement paste specimens with water-to-cement (w/c) ratios (0.35, 0.40, 0.45, 0.50, 0.55) in the frequency range of 1.02 GHz to 4.50 GHz were studied and modeled using modified Debye's models. An open-ended coaxial probe and a network analyzer were used to measure dielectric properties. Forward models are proposed and inversed for predicting the w/c ratio of a given oven-dried cement paste specimen. Modeling results agreed with the experimental data. The proposed models can be used for predicting the dielectric properties of oven-dried cement paste specimens. Also, the modeling approach can be applied to other cementitious materials (e.g., concrete) with additional modification.

  13. A Pyrosequencing-Based Assay for the Rapid Detection of the 22q11.2 Deletion in DNA from Buccal and Dried Blood Spot Samples

    PubMed Central

    Koontz, Deborah; Baecher, Kirsten; Kobrynski, Lisa; Nikolova, Stanimila; Gallagher, Margaret

    2015-01-01

    The 22q11.2 deletion syndrome is one of the most common deletion syndromes in newborns. Some affected newborns may be diagnosed shortly after birth because of the presence of heart defects, palatal defects, or severe immune deficiencies. However, diagnosis is often delayed in patients presenting with other associated conditions that would benefit from early recognition and treatment, such as speech delays, learning difficulties, and schizophrenia. Fluorescence in situ hybridization (FISH) is the gold standard for deletion detection, but it is costly and time consuming and requires a whole blood specimen. Our goal was to develop a suitable assay for population-based screening of easily collectible specimens, such as buccal swabs and dried blood spots (DBS). We designed a pyrosequencing assay and validated it using DNA from FISH–confirmed 22q11 deletion syndrome patients and normal controls. We tested DBS from nine patients and paired buccal cell and venous blood specimens from 20 patients. Results were 100% concordant with FISH assay results. DNA samples from normal controls (n = 180 cell lines, n = 15 DBS, and n = 88 buccal specimens) were negative for the deletion. Limiting dilution experiments demonstrated that accurate results could be obtained from as little as 1 ng of DNA. This method represents a reliable and low-cost alternative for detection of the common 22q11.2 microdeletions and can be adapted to high-throughput population screening. PMID:24973633

  14. A method for drying red blood cells for solid-phase immunoassay.

    PubMed

    Tamai, T; Mazda, T

    1999-12-01

    To develop a simpler and quicker alloantibody screening method, red cell stroma were bound and dried to microplate wells for use in magnetic-mixed passive haemagglutination (M-MPHA) tests. In the procedure of drying stroma, the Triton X-100-based haemolysing method gave lowest denaturation of red blood cells, and this method gave increased reactivity to Kidd and Rh antigens and clinically significant antibodies were detected as well as with the M-MPHA test. But long incubation with Triton X-100 and using high concentrations of Triton X-100 gave rise to some reduction in antigenicity, so the precise conditions for haemolysis are critical. This dried stroma-coated microplate can be stored for longer and more easily at room temperature than nondried intact red blood cells. The new system gave good sensitivity and the overall test time was shortened and should give a particular advantage for mass screening and for automation of this test. PMID:10583889

  15. A novel method for quantification of human hemoglobin from dried blood spots by use of tandem mass spectrometry.

    PubMed

    Yu, Chaowen; Zhang, Juan; Yuan, Zhaojian; Liu, Hao; Wang, Xingbin; Wang, Ming; Zou, Lin

    2015-10-01

    Quantification of human hemoglobin (Hb) is essential for diagnosis of anemia, especially for screening for thalassemia and sickle cell disease. The main methods currently used for quantification of Hb, including spectrophotometry, fluorimetry, and electrochemical assays, are all based on the structural integrity of Hb, which could be affected by hemolysis and degradation. When used for disease screening, whole blood specimens cannot meet requirements for sample collecting, transport, and storage. Here, we report a novel MS-MS method for quantification of Hb from dried blood spots (DBS) by use of a triple-quadrupole mass spectrometer. Proteospecific peptides from α-globin chains were selected after tryptic digestion. The precursor → product ion transitions of representative peptides were studied to identify the best choice with regard to sensitivity and chromatographic properties. For quantification, stable isotope-labeled peptides were used as internal standards. The concentration of Hb in each sample was obtained by calculation on the basis of established equations. The precision of the method was within 15 % and accuracy was in the range -7 to 13.0 %. Compared with routine clinical results obtained by use of the automated hematology analyzer (AHA) assay, the correlation, r (2), was >0.993. When used for determination of anemia levels the sensitivity of the assay was 95.7 % and specificity 96.5 %. Our new approach for quantification of the concentration of Hb from DBS is feasible, and precision is acceptable. The method could be used for determination of anemia levels when screening for hemoglobin disorders. Graphical Abstract Quantification of human hemoglobin from digested dried blood spot samples using tandem mass spectrometry. PMID:26345440

  16. Freeze-Drying of Mononuclear Cells Derived from Umbilical Cord Blood Followed by Colony Formation

    PubMed Central

    Natan, Dity; Nagler, Arnon; Arav, Amir

    2009-01-01

    Background We recently showed that freeze-dried cells stored for 3 years at room temperature can direct embryonic development following cloning. However, viability, as evaluated by membrane integrity of the cells after freeze-drying, was very low; and it was mainly the DNA integrity that was preserved. In the present study, we improved the cells' viability and functionality after freeze-drying. Methodology/Principal Findings We optimized the conditions of directional freezing, i.e. interface velocity and cell concentration, and we added the antioxidant EGCG to the freezing solution. The study was performed on mononuclear cells (MNCs) derived from human umbilical cord blood. After freeze-drying, we tested the viability, number of CD34+-presenting cells and ability of the rehydrated hematopoietic stem cells to differentiate into different blood cells in culture. The viability of the MNCs after freeze-drying and rehydration with pure water was 88%–91%. The total number of CD34+-presenting cells and the number of colonies did not change significantly when evaluated before freezing, after freeze-thawing, and after freeze-drying (5.4×104±4.7, 3.49×104±6 and 6.31×104±12.27 cells, respectively, and 31±25.15, 47±45.8 and 23.44±13.3 colonies, respectively). Conclusions This is the first report of nucleated cells which have been dried and then rehydrated with double-distilled water remaining viable, and of hematopoietic stem cells retaining their ability to differentiate into different blood cells. PMID:19381290

  17. Experimental microbial contamination and disinfection of dry (vapour) shipper dewars designed for short-term storage and transportation of cryopreserved germplasm and other biological specimens.

    PubMed

    Bielanski, A

    2005-04-15

    Cryopreservation, storage and transport of cryopreserved germplasm without the risk of disease transmission is of great concern to animal and human health authorities. Here we report on the efficacy of microbial decontamination of the liquid nitrogen (LN) dry (vapour) shippers used for short-term storage and transportation of germplasm and other biological specimens. Dry shippers containing either a hydrophobic or a non-hydrophobic LN absorbent were experimentally contaminated with high titers of cultures of Pseudomonas aeruginosa, Escherichia coli, Staphylococus aureus, bovine viral diarrhea virus (BVDV) and bovine herpesvirus-1 (BHV-1). Biocidals with broad spectrum antimicrobial activity and gas vapours of formalin and ethylene oxide were used for disinfection of the dewars. Among the biocidals used, treatment with sodium hypochlorite solution, the quaternary ammonium-based disinfectants and peracetic acid were the most effective and useful for dry shippers with a hydrophobic LN absorbent. None of the bacterial or viral microorganisms were detected in samples of semen and embryos stored in dry shippers following their disinfection with these biocides. An application of some other disinfectants, due to their foaming properties or to the permeability of the absorbent hydrophobic membrane (HM) was not effective or may have caused irreversible damage to the LN absorbent. Gas sterilization by ethylene oxide in contrast to formalin was fully effective for both types of dry shippers. PMID:15823351

  18. Urine as a biological specimen for forensic analysis of alcohol and variability in the urine-to-blood relationship.

    PubMed

    Jones, Alan W

    2006-01-01

    This article concerns the use of urine as a biological specimen for determination of alcohol in clinical and forensic toxicology and discusses factors that might influence variability in the urine/blood concentration ratio of alcohol. A large number of human drinking experiments were conducted to determine the time course of urine-alcohol concentrations (UAC) in relation to blood-alcohol concentrations (BAC). The UAC and BAC curves were shifted in time and the BAC curve always began to decrease before the UAC started to decline. During the early absorption phase the UAC/BAC ratio was less than unity, whereas in the late absorption/distribution period the ratio was between 1.0-1.2. On reaching the post-absorptive phase, the UAC always exceeded BAC and UAC/BAC ratios averaged 1.3-1.4, increasing appreciably as BAC decreased towards zero. Alcohol-induced diuresis was most pronounced during the rising portion of the BAC curve and near to the peak value. After about 2 hours post-drinking, the production rate of urine diminished to the pre-drinking rate of about 0.5-1 mL/min. Drinking water during the post-absorptive phase of the alcohol curve produced dilute urine, as reflected in lower creatinine content and osmolality, although the concentration of ethanol remained unchanged. After subjects drank a moderate dose of ethanol (0.54-0.85 g/kg) about 2% of the dose was recoverable in the urine after 7 hours. Ethyl glucuronide, a minor metabolite of ethanol, was measured in urine samples from drunk drivers. The UAC/BAC ratio of ethanol in drunk drivers did not depend on the creatinine content of the urine and therefore the relative dilution of the specimens. When alcohol-free urine was spiked with glucose and infected with the yeast species Candida albicans, ethanol was produced by fermentation after approximately 24 hours storage at room temperature. This post-sampling synthesis of ethanol was prevented by sodium fluoride (1% weight by volume) in the urine tubes or by

  19. Development of an assay to simultaneously measure orotic acid, amino acids, and acylcarnitines in dried blood spots

    PubMed Central

    Held, Patrice K.; Haynes, Christopher A.; De Jesús, Víctor R.; Baker, Mei W.

    2016-01-01

    Background Orotic aciduria in the presence of hyperammonemia is a key indicator for a defect in the urea cycle, specifically ornithine transcarbamylase (OTC) deficiency. Current newborn screening (NBS) protocols can detect several defects of the urea cycle, but screening for OTC deficiency remains a challenge due to the lack of a suitable assay. The purpose of this study was to develop a high-throughput assay to measure orotic acid in dried blood spot (DBS) specimens as an indicator for urea cycle dysfunction, which can be readily incorporated into routine NBS. Methods Orotic acid was extracted from DBS punches and analyzed using flow-injection analysis tandem mass spectrometry (FIA–MS/MS) with negative-mode ionization, requiring <2 min/sample run time. This method was then multiplexed into a conventional newborn screening assay for analysis of amino acids, acylcarnitines, and orotic acid. Results We describe 2 assays which can quantify orotic acid in DBS: a stand-alone method and a combined method for analysis of orotic acid, amino acids, and acylcarnitines. Both methods demonstrated orotic acid recovery of 75–85% at multiple levels of enrichment. Precision was also comparable to traditional FIA–MS/MS methods. Analysis of residual presumptively normal NBS specimens demonstrated a 5:1 signal to noise ratio and the average concentration of orotic acid was approximately 1.2 μmol/l. The concentration of amino acids and acylcarnitines as measured by the combined method showed no significant differences when compared to the conventional newborn screening assay. In addition, retrospective analysis of confirmed patients and presumptively normal newborn screening specimens suggests potential for the methods to identify patients with OTC deficiency, as well as other urea cycle defects. Conclusion The assays described here quantify orotic acid in DBS using a simple extraction and FIA–MS/MS analysis procedures that can be implemented into current NBS protocols. PMID

  20. Supercritical fluid extraction as a preparation method for mass spectrometry of dried blood spots.

    PubMed

    Matsubara, Atsuki; Izumi, Yoshihiro; Nishiumi, Shin; Suzuki, Makoto; Azuma, Takeshi; Fukusaki, Eiichiro; Bamba, Takeshi; Yoshida, Masaru

    2014-10-15

    The potential of supercritical fluid extraction (SFE) as a preparation method for mass spectrometry of dried blood spots (DBS) was examined. SFE is generally used for the extraction of hydrophobic compounds, but hydrophilic metabolites such as amino acids, amines, and nucleic-acid-related metabolites could be extracted by adding a low level of methanol as a modifier. Under the optimized conditions, over 200 metabolites were detected from a dried serum spot, of which over 160 metabolites could be analyzed stably (RSD <20%). These results show that SFE is an effective extraction method of metabolites with a wide range of polarity in DBS. PMID:25178194

  1. Validation and Modification of Dried Blood Spot-Based Glycosylated Hemoglobin Assay for the Longitudinal Aging Study in India

    PubMed Central

    Hu, Peifeng; Edenfield, Michael; Potter, Alan; Kale, Varsha; Risbud, Arun; Williams, Sharon; Lee, Jinkook; Bloom, David E.; Crimmins, Eileen; Seeman, Teresa

    2014-01-01

    Objectives This study aims to validate a modified dried blood spot (DBS)-based glycosylated hemoglobin (HbA1c) assay protocol, after a pretest in India showed poor correlation between the original DBS-based protocol and venous results. Methods The original protocol was tested on different chemistry analyzers and then simplified at the University of Washington (UW). A second pretest was conducted in India to validate the modified assay protocol, using 44 quality control specimens. Results Data from UW indicated that, using the original protocol, the correlation coefficients between DBS and venous results were above 0.98 on both Bio-Rad and Olympus chemistry analyzers. The protocol worked equally well on filter paper, with or without pre-treatment, and when the recommended amount of blood spot material, or less, was used. A second pretest of the modified protocol confirmed that DBS-based levels from both Olympus and Roche chemistry analyzers were well correlated with DBS results from UW (correlation coefficients were above 0.96), as well as with venous values (correlation coefficients were above 0.94). Conclusions The DBS-based HbA1c values are highly correlated with venous results. The pre-treatment of filter paper does not appear to be necessary. The poor results from the first pretest are probably due to factors unrelated to the protocol, such as problems with the chemistry analyzer or assay reagents. PMID:25472916

  2. Emerging liquid chromatography-mass spectrometry technologies improving dried blood spot analysis.

    PubMed

    Rao, Ramisetti Nageswara

    2014-08-01

    Dried blood spots (DBS), a micro blood sampling technique, has recently gained interest in drug discovery and development due to its inherent advantages over the conventional whole blood, plasma or serum sample collection. Since the regulatory authorities have agreed to the use of blood as an acceptable biological matrix for drug exposure measurements, its applications have been extended not only to therapeutic drug monitoring but also to toxicokinetic and pharmacokinetic studies. The pharmaceutical industry is keen to promote DBS as a prominent tool in bioanalytical applications due to the financial, ethical and organizational issues involved in clinical trials. This could be accomplished due to the latest advances in modern analytical technology, particularly liquid chromatography-mass spectrometry. The present review discusses some of the emerging liquid chromatography-mass spectrometry technologies in improving DBS analysis for its innovative applications in the development of new drugs. PMID:24697571

  3. Effects of blood sample handling procedures on measurable inflammatory markers in plasma, serum and dried blood spot samples.

    PubMed

    Skogstrand, Kristin; Ekelund, Charlotte K; Thorsen, Poul; Vogel, Ida; Jacobsson, Bo; Nørgaard-Pedersen, Bent; Hougaard, David M

    2008-07-20

    The interests in monitoring inflammation by immunoassay determination of blood inflammatory markers call for information on the stability of these markers in relation to the handling of blood samples. The increasing use of stored biobank samples for such ventures that may have been collected and stored for other purposes, justifies the study hereof. Blood samples were stored for 0, 4, 24, and 48 h at 4 degrees C, room temperature (RT), and at 35 degrees C, respectively, before they were separated into serum or plasma and frozen. Dried blood spot samples (DBSS) were stored for 0, 1, 2, 3, 7, and 30 days at the same temperatures. 27 inflammatory markers in serum and plasma and 25 markers in DBSS were measured by a previously validated multiplex sandwich immunoassay using Luminex xMAP technology. The measurable concentrations of several cytokines in serum and plasma were significantly increased when blood samples were stored for a period of time before the centrifugation, for certain cytokines more than 1000 fold compared to serum and plasma isolated and frozen immediately after venepuncture. The concentrations in serum generally increased more than in plasma. The measurable concentrations of inflammatory markers also changed in DBSS stored under various conditions compared to controls frozen immediately after preparation, but to a much lesser degree than in plasma or serum. The study demonstrates that trustworthy measurement of several inflammatory markers relies on handling of whole blood samples at low temperatures and rapid isolation of plasma and serum. Effects of different handling procedures for all markers studied are given. DBSS proved to be a robust and convenient way to handle samples for immunoassay analysis of inflammatory markers in whole blood. PMID:18495149

  4. Dried Blood Spots - Preparing and Processing for Use in Immunoassays and in Molecular Techniques

    PubMed Central

    Grüner, Nico; Stambouli, Oumaima; Ross, R. Stefan

    2015-01-01

    The idea of collecting blood on a paper card and subsequently using the dried blood spots (DBS) for diagnostic purposes originated a century ago. Since then, DBS testing for decades has remained predominantly focused on the diagnosis of infectious diseases especially in resource-limited settings or the systematic screening of newborns for inherited metabolic disorders and only recently have a variety of new and innovative DBS applications begun to emerge. For many years, pre-analytical variables were only inappropriately considered in the field of DBS testing and even today, with the exception of newborn screening, the entire pre-analytical phase, which comprises the preparation and processing of DBS for their final analysis has not been standardized. Given this background, a comprehensive step-by-step protocol, which covers al the essential phases, is proposed, i.e., collection of blood; preparation of blood spots; drying of blood spots; storage and transportation of DBS; elution of DBS, and finally analyses of DBS eluates. The effectiveness of this protocol was first evaluated with 1,762 coupled serum/DBS pairs for detecting markers of hepatitis B virus, hepatitis C virus, and human immunodeficiency virus infections on an automated analytical platform. In a second step, the protocol was utilized during a pilot study, which was conducted on active drug users in the German cities of Berlin and Essen. PMID:25867233

  5. Elevation of guanidinoacetate in newborn dried blood spots and impact of early treatment in GAMT deficiency.

    PubMed

    El-Gharbawy, Areeg H; Goldstein, Jennifer L; Millington, David S; Vaisnins, Amie E; Schlune, Andrea; Barshop, Bruce A; Schulze, Andreas; Koeberl, Dwight D; Young, Sarah P

    2013-06-01

    Guanidinoacetate methyltransferase (GAMT) deficiency is a good candidate disorder for newborn screening because early treatment appears to improve outcomes. We report elevation of guanidinoacetate in archived newborn dried blood spots for 3 cases (2 families) of GAMT deficiency compared with an unaffected carrier and controls. We also report a new case of a patient treated from birth with normal developmental outcome at the age of 42 months. PMID:23583224

  6. Guidelines for the qualitative detection of viral genomes in dried blood spots.

    PubMed

    Gibellini, Davide; De Crignis, Elisa; Re, Maria Carla

    2012-01-01

    Dried blood spots (DBSs) are a useful alternative to blood sampling especially in children or for screening high-risk populations in developing countries. DBS blood collection can be employed in the diagnosis of viral infections by PCR or RT-PCR and also in viral genome sequencing. In addition, the advent of multiplex PCR approaches has led to further diagnostic and methodological improvements allowing simultaneous detection of two or more different viral genomes in the same sample and amplification reaction. This chapter describes general guidelines for the qualitative viral genome amplification and detection in DBS providing an example application of a qualitative real-time SYBR Green-based multiplex RT-PCR assay targeting two major viral pathogens, HIV-1 and HCV. PMID:22782809

  7. Determination of Total Glutathione in Dried Blood Spot Samples Using a High-Performance Liquid Chromatography.

    PubMed

    Kanďár, Roman; Štramová, Xenie; Drábková, Petra; Brandtnerová, Martina

    2015-07-01

    A method is described for the determination of total glutathione (TGSH) in dried blood spot (DBS) samples using high-performance liquid chromatography with fluorescence detection. Whole blood and DBS samples were obtained from a group of blood donors. After GSH reduction with dithiothreitol and protein precipitation with ethanol, the samples were derivatized with naphthalene-2,3-dicarboxaldehyde to form a very stable, highly fluorescent derivative. For the separation, a reversed phase HPLC method was used. The mixture of ethanol and deionized water (8 : 92, v/v) was used as a mobile phase. The analytical performance of this method was satisfactory: the intra- and interassay coefficients of variation were below 10%. Quantitative recoveries from spiked DBS samples were between 98.3 and 103.6%. The presented method is inexpensive and suitable for clinical testing purposes. PMID:25344838

  8. Quantification of the Immunosuppressant Tacrolimus on Dried Blood Spots Using LC-MS/MS.

    PubMed

    Shokati, Touraj; Bodenberger, Nicholas; Gadpaille, Holly; Schniedewind, Björn; Vinks, Alexander A; Jiang, Wenlei; Alloway, Rita R; Christians, Uwe

    2015-01-01

    The calcineurin inhibitor tacrolimus is the cornerstone of most immunosuppressive treatment protocols after solid organ transplantation in the United States. Tacrolimus is a narrow therapeutic index drug and as such requires therapeutic drug monitoring and dose adjustment based on its whole blood trough concentrations. To facilitate home therapeutic drug and adherence monitoring, the collection of dried blood spots is an attractive concept. After a finger stick, the patient collects a blood drop on filter paper at home. After the blood is dried, it is mailed to the analytical laboratory where tacrolimus is quantified using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) in combination with a simple manual protein precipitation step and online column extraction. For tacrolimus analysis, a 6-mm disc is punched from the saturated center of the blood spot. The blood spot is homogenized using a bullet blender and then proteins are precipitated with methanol/0.2 M ZnSO4 containing the internal standard D2,(13)C-tacrolimus. After vortexing and centrifugation, 100 µl of supernatant is injected into an online extraction column and washed with 5 ml/min of 0.1 formic acid/acetonitrile (7:3, v:v) for 1 min. Hereafter, the switching valve is activated and the analytes are back-flushed onto the analytical column (and separated using a 0.1% formic acid/acetonitrile gradient). Tacrolimus is quantified in the positive multi reaction mode (MRM) using a tandem mass spectrometer. The assay is linear from 1 to 50 ng/ml. Inter-assay variability (3.6%-6.1%) and accuracy (91.7%-101.6%) as assessed over 20 days meet acceptance criteria. Average extraction recovery is 95.5%. There are no relevant carry-over, matrix interferences and matrix effects. Tacrolimus is stable in dried blood spots at RT and at +4 °C for 1 week. Extracted samples in the autosampler are stable at +4 °C for at least 72 hr. PMID:26575262

  9. Protein expression profiling by antibody array analysis with use of dried blood spot samples on filter paper.

    PubMed

    Jiang, Weidong; Mao, Ying Qing; Huang, Ruochun; Duan, Chaohui; Xi, Yun; Yang, Kai; Huang, Ruo-Pan

    2014-01-31

    Dried blood spot samples (DBSS) on filter paper offer several advantages compared to conventional serum/plasma samples: they do not require any phlebotomy or separation of blood by centrifugation; they are less invasive; they allow sample stability and shipment at room temperature; and they pose a negligible risk of infection with blood-borne viruses, such as HIV, HBV and HCV, to those who handle them. Therefore dried blood spot samples (DBSS) on filter paper can be a quick, convenient and inexpensive means of obtaining blood samples for biomarker discovery, disease screening, diagnosis and treatment monitoring in non-hospitalized, public health settings. In this study, we investigated for the first time the potential application of dried blood spot samples (DBSS) in protein expression profiling using antibody array technology. First, optimal conditions for array assay performance using dried blood spot samples (DBSS) was established, including sample elution buffer, elution time, elution temperature and assay blocking buffer. Second, we analyzed dried blood spot samples (DBSS) using three distinct antibody array platforms, including sandwich-based antibody arrays, quantitative antibody arrays and biotin-label-based antibody arrays. In comparison with paired serum samples, detection of circulating proteins in dried blood spot samples (DBSS) correlated well for both low- and high-abundance proteins on all three antibody array platforms. In conclusion, our study strongly indicates the novel application of multiplex antibody array platforms to analyze dried blood spot samples (DBSS) on filter paper represents a viable, cost-effective method for protein profiling, biomarker discovery and disease screening in a large, population-based survey. PMID:24287424

  10. Quantitation of Tenofovir and Emtricitabine in Dried Blood Spots (DBS) with LC-MS/MS

    PubMed Central

    Zheng, Jia-Hua; Guida, Louis A; Rower, Caitlin; Castillo-Mancilla, Jose; Meditz, Amie; Klein, Brandon; Kerr, Becky Jo; Langness, Jacob; Bushman, Lane; Kiser, Jennifer; Anderson, Peter L.

    2013-01-01

    A reversed-phase high performance liquid chromatographic (LC), tandem mass spectrometry (MS/MS) assay for the determination of tenofovir (TFV) and emtricitabine (FTC) in dried blood spots (DBS) from human whole blood was developed and validated. Whole blood samples were spotted, dried, and a 3mm punch was extracted with methanol for analysis by LC-MS/MS utilizing stable isotope labeled internal standards. The assay was validated over the range of 2.5ng/mL to 1,000ng/mL for TFV and 2.5ng/mL to 5,000ng/mL for FTC. The method was accurate (within ± 15% of control) and precise (coefficient of variation ≤ 15%) for hematocrit concentrations ranging from 25% to 76%; using edge punches versus center punches; and spot volumes of 10µL to 50µL. Analytes were stable for five freeze/thaw cycles and up to 6 days at room temperature, whereas long-term storage required −20°C or −80°C. Comparison of TFV and FTC in DBS versus plasma yielded r2 ≥ 0.96, indicating that DBS can be used as a plasma alternative for pharmacokinetic analyses in vivo. PMID:24055850

  11. Tacrolimus and sirolimus in capillary dried blood spots allows for remote monitoring.

    PubMed

    Dickerson, Jane A; Sinkey, Marian; Jacot, Kathleen; Stack, Jennifer; Sadilkova, Katerina; Law, Yuk M; Jack, Rhona M

    2015-02-01

    Therapeutic drug monitoring of tacrolimus and sirolimus plays a significant role in the clinical follow-up of transplant patients receiving IMS therapy. Success of transplant and favorable patient outcome relies on maintaining adequate therapeutic drug levels. The purpose of this research is to assess the clinical utility of remote collection of DBS for immunosuppressant monitoring and compare the IMS level in paired collections of venous whole blood and DBS. Sirolimus and tacrolimus levels were clinically correlated in capillary blood collected from a finger poke with venous whole blood from pediatric, post-transplant patients. The participants took the dried blood spot card home with them with a pre-addressed, postage-paid envelope and mailed it back to the laboratory. Overall, a small but statistically significant negative bias was observed (-0.6 ng/mL, p = 0.0011). A chart review was performed to assess whether clinical management would have changed, and none of the cases revealed a clinically significant change. Sirolimus in DBS also correlated with venous levels. Overall, a small but statistically negative bias was observed (-0.8 ng/mL, p = 0.029). In summary, analysis of IMS levels in DBS is possible, and the difference noted between capillary and venous blood is within the clinically acceptable limits. PMID:25414084

  12. Cryopreservation of glucose-6-phosphate dehydrogenase activity inside red blood cells: developing a specimen repository in support of development and evaluation of glucose-6-phosphate dehydrogenase deficiency tests

    PubMed Central

    2013-01-01

    Background Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a common human enzyme deficiency. It is characterized by abnormally low levels of G6PD activity. Individuals with G6PD deficiency are at risk of undergoing acute haemolysis when exposed to 8‒aminoquinoline-based drugs, such as primaquine. For this reason it is imperative to identify individuals with G6PD deficiency prior to administering these anti-malarial drugs. There is a need for the development and evaluation of point-of-care G6PD deficiency screening tests suitable for areas of the developing world where malarial treatments are frequently administered. The development and evaluation of new G6PD tests will be greatly assisted with the availability of specimen repositories. Methods Cryopreservation of erythrocytes was evaluated as a means to preserve G6PD activity. Blood specimens from 31 patients including ten specimens with normal G6PD activity, three with intermediate activity, and 18 with deficient activity were cryopreserved for up to six months. Results Good correlation in G6PD activity between fresh and cryopreserved specimens (R2 = 0.95). The cryopreserved specimens show an overall small drop in mean G6PD activity of 0.23 U/g Hb (P=0.23). Cytochemical staining showed that intracellular G6PD activity distribution within the red blood cell populations is preserved during cryopreservation. Furthermore, the mosaic composition of red blood cells in heterozygous women is also preserved for six months or more. The fluorescent spot and the BinaxNOW qualitative tests for G6PD deficiency also showed high concordance in G6PD status determination between cryopreserved specimens and fresh specimens. Conclusions A methodology for establishing a specimen panel for evaluation of G6PD tests is described. The approach is similar to that used in several malaria research facilities for the cryopreservation of parasites in clinical specimens and axenic cultures. Specimens stored in this manner will aid

  13. Dried Blood Spot Proteomics: Surface Extraction of Endogenous Proteins Coupled with Automated Sample Preparation and Mass Spectrometry Analysis

    NASA Astrophysics Data System (ADS)

    Martin, Nicholas J.; Bunch, Josephine; Cooper, Helen J.

    2013-08-01

    Dried blood spots offer many advantages as a sample format including ease and safety of transport and handling. To date, the majority of mass spectrometry analyses of dried blood spots have focused on small molecules or hemoglobin. However, dried blood spots are a potentially rich source of protein biomarkers, an area that has been overlooked. To address this issue, we have applied an untargeted bottom-up proteomics approach to the analysis of dried blood spots. We present an automated and integrated method for extraction of endogenous proteins from the surface of dried blood spots and sample preparation via trypsin digestion by use of the Advion Biosciences Triversa Nanomate robotic platform. Liquid chromatography tandem mass spectrometry of the resulting digests enabled identification of 120 proteins from a single dried blood spot. The proteins identified cross a concentration range of four orders of magnitude. The method is evaluated and the results discussed in terms of the proteins identified and their potential use as biomarkers in screening programs.

  14. Genome-Wide Profiling of RNA from Dried Blood Spots: Convergence with Bioinformatic Results Derived from Whole Venous Blood and Peripheral Blood Mononuclear Cells.

    PubMed

    McDade, Thomas W; M Ross, Kharah; L Fried, Ruby; Arevalo, Jesusa M G; Ma, Jeffrey; Miller, Gregory E; Cole, Steve W

    2016-01-01

    Genome-wide transcriptional profiling has emerged as a powerful tool for analyzing biological mechanisms underlying social gradients in health, but utilization in population-based studies has been hampered by logistical constraints and costs associated with venipuncture blood sampling. Dried blood spots (DBS) provide a minimally invasive, low-cost alternative to venipuncture, and in this article we evaluate how closely the substantive results from DBS transcriptional profiling correspond to those derived from parallel analyses of gold-standard venous blood samples (PAXgene whole blood and peripheral blood mononuclear cells [PBMC]). Analyses focused on differences in gene expression between African-Americans and Caucasians in a community sample of 82 healthy adults (age 18-70 years; mean 35). Across 19,679 named gene transcripts, DBS-derived values correlated r = .85 with both PAXgene and PBMC values. Results from bioinformatics analyses of gene expression derived from DBS samples were concordant with PAXgene and PBMC samples in identifying increased Type I interferon signaling and up-regulated activity of monocytes and natural killer (NK) cells in African-Americans compared to Caucasian participants. These findings demonstrate the feasibility of DBS in field-based studies of gene expression and encourage future studies of human transcriptome dynamics in larger, more representative samples than are possible with clinic- or lab-based research designs. PMID:27337553

  15. Lack of Detection of Human Retrovirus-5 Proviral DNA in Synovial Tissue and Blood Specimens From Individuals With Rheumatoid Arthritis or Osteoarthritis

    PubMed Central

    PIPER, KERRYL E.; HANSSEN, ARLEN D.; LEWALLEN, DAVID G.; MATTESON, ERIC L.; OSMON, DOUGLAS R.; DUFFY, MARY C.; HAGAN, ROCHELLE A.; STECKELBERG, JAMES M.; PATEL, ROBIN

    2006-01-01

    Objective Prior studies have suggested an association of human retrovirus 5 with rheumatoid arthritis. The purpose of this study was to determine if human retrovirus-5 proviral DNA is present in synovial tissue and blood specimens from patients with rheumatoid arthritis or osteoarthritis, or those without joint disease. Methods Synovial tissue and whole blood from 75 patients with rheumatoid arthritis, 75 patients with osteoarthritis, and 50 patients without a primary arthritis diagnosis were assayed by real-time quantitative polymerase chain reaction (PCR) using primers that amplify a 186-bp fragment of human retrovirus-5 proviral DNA. Results A total of 200 tissue specimens, 200 mononuclear cells, and 196 of 200 granulocyte specimens tested negative for human retrovirus-5 proviral DNA. No association between human retrovirus 5 and rheumatoid arthritis or osteoarthritis (P = 0.516) was identified. Granulocyte specimens from 4 patients, 2 with rheumatoid arthritis and 2 with osteoarthritis, yielded a low positive human retrovirus-5 proviral DNA signal (83–1,365 copies of human retrovirus-5 proviral DNA/ml blood). Conclusion Contrary to prior reports, we did not find an association between human retrovirus 5 and rheumatoid arthritis or osteoarthritis using a real-time PCR assay. Our findings are consistent with the recent finding that human retrovirus 5 is actually rabbit endogenous retrovirus H. PMID:16463423

  16. Rapid assays for Gaucher and Hurler diseases in dried blood spots using digital microfluidics

    PubMed Central

    Sista, Ramakrishna S.; Wang, Tong; Wu, Ning; Graham, Carrie; Eckhardt, Allen; Bali, Deeksha; Millington, David S.; Pamula, Vamsee K.

    2013-01-01

    Objective Easy tool for newborn screening of Gaucher and Hurler diseases. Methods Method comparison between fluorometric enzymatic activity assay on a digital microfluidic platform and micro-titer plate bench assay was performed on normal (n=100), Gaucher (n=10) and Hurler (n=7) dried blood spot samples. Results Enzymatic activity analysis of glucocerebrosidase (Gaucher) and α-L-iduronidase (Hurler) revealed similar discrimination between normal and affected samples on both platforms. Conclusions Digital microfluidics is suitable for Gaucher and Hurler newborn screening. PMID:23578771

  17. Dried blood spots analysis with mass spectrometry: Potentials and pitfalls in therapeutic drug monitoring.

    PubMed

    Antunes, Marina Venzon; Charão, Mariele Feiffer; Linden, Rafael

    2016-09-01

    Therapeutic drug monitoring (TDM) relays in the availability of specialized laboratory assays, usually available in reference centers that are not accessible to all patients. In this context, there is a growing interest in the use of dried blood spot (DBS) sampling, usually obtained from finger pricks, which allows simple and cost-effective logistics in many settings, particularly in Developing Countries. The use of DBS assays to estimate plasma concentrations is highly dependent on the hematocrit of the blood, as well as the particular characteristics of the measured analyte. DBS assays require specific validation assays, most of them are related to hematocrit effects. In the present manuscript, the application of mass spectrometric assays for determination of drugs for TDM purposes in the last ten years is reviewed, as well as the particular validation assays for new DBS methods. PMID:27179588

  18. What do hemolyzed whole-blood specimens look like? Analysis with a CellaVision DM96 automated image analysis system.

    PubMed

    Lippi, Giuseppe; Pavesi, Fernanda; Benegiamo, Anna; Pipitone, Silvia

    2015-02-01

    We planned an original study to investigate the morphological changes caused by spurious hemolysis of whole-blood samples, analyzed using an automated image analysis system. Seven whole-blood specimens anticoagulated with EDTA were pooled and divided in two aliquots. The former was left untreated, whereas the latter was subjected to mechanical hemolysis by forced aspiration with an insulin syringe. The complete blood cell count was performed on a Sysmex XE-2100, and the aliquots were then processed with CellaVision DM96. In spuriously hemolyzed samples, the main findings included a rarefaction of erythrocytes, the presence of a remarkable number of cellular debris, a greater degree of microcytosis and anisocytosis, the appearance of band neutrophils, a shift of values between lymphocytes and monocytes, and an increase in smudge cells, artifacts, and large platelets. The results of this study demonstrate for the first time that blood cell morphology may be consistently biased in spuriously hemolyzed whole blood and that the use of an automated image analysis system such as the CellaVision DM96 may be a suitable approach to identify spurious hemolysis in whole-blood specimens. PMID:25395293

  19. A dried blood spot mass spectrometry metabolomic approach for rapid breast cancer detection

    PubMed Central

    Wang, Qingjun; Sun, Tao; Cao, Yunfeng; Gao, Peng; Dong, Jun; Fang, Yanhua; Fang, Zhongze; Sun, Xiaoyu; Zhu, Zhitu

    2016-01-01

    Objective Breast cancer (BC) is still a lethal threat to women worldwide. An accurate screening and diagnosis strategy performed in an easy-to-operate manner is highly warranted in clinical perspective. Besides the routinely focused protein markers, blood is full of small molecular metabolites with diverse structures and properties. This study aimed to screen metabolite markers with BC diagnosis potentials. Methods A dried blood spot-based direct infusion mass spectrometry (MS) metabolomic analysis was conducted for BC and non-BC differentiation. The targeted analytes included 23 amino acids and 26 acylcarnitines. Results Multivariate analysis screened out 21 BC-related metabolites in the blood. Regression analysis generated a diagnosis model consisting of parameters Pip, Asn, Pro, C14:1/C16, Phe/Tyr, and Gly/Ala. Tested with another set of BC and non-BC samples, this model showed a sensitivity of 92.2% and a specificity of 84.4%. Compared to the routinely used protein markers, this model exhibited distinct advantage with its higher sensitivity. Conclusion Blood metabolites screening is a more plausible approach for BC detection. Furthermore, this direct MS analysis could be finished within few minutes, which means that its throughput is higher than the currently used imaging techniques. PMID:27042107

  20. Do capillary dried blood spot concentrations of gamma-hydroxybutyric acid mirror those in venous blood? A comparative study.

    PubMed

    Sadones, Nele; Archer, John R H; Ingels, Ann-Sofie M E; Dargan, Paul I; Wood, David M; Wood, Michelle; Neels, Hugo; Lambert, Willy E; Stove, Christophe P

    2015-04-01

    Gamma-hydroxybutyric acid (GHB) is a well-known illicit club and date-rape drug. Dried blood spot (DBS) sampling is a promising alternative for classical venous sampling in cases of (suspected) GHB intoxication since it allows rapid sampling, which is of interest for the extensively metabolized GHB. However, there is limited data if -and how- capillary DBS concentrations correlate with venous concentrations. We conducted a comparative study in 50 patients with suspected GHB intoxication, to determine and to correlate GHB concentrations in venous DBS (vDBS) and capillary DBS (cDBS). This is the first study that evaluates in a large cohort the correlation between capillary and venous concentrations of an illicit drug in real-life samples. Of the 50 paired samples, 7 were excluded: the vDBS concentration was below the LLOQ of 2 µg/mL in 3 cases and 4 samples were excluded after visual inspection of the DBS. Bland-Altman analysis revealed a mean % difference of -2.8% between cDBS and vDBS concentrations, with the zero value included in the 95% confidence interval of the mean difference in GHB concentration. A paired sample t-test confirmed this observation (p = 0.17). Also the requirement for incurred sample reproducibility was fulfilled: for more than two-thirds of the samples the concentrations obtained in cDBS and those in vDBS were within 20% of their mean. Since equivalent concentrations were observed in cDBS and vDBS, blood obtained by fingerprick can be considered a valid alternative for venous blood for GHB determination. PMID:25565078

  1. Application of automated serial blood sampling and dried blood spot technique with liquid chromatography-tandem mass spectrometry for pharmacokinetic studies in mice.

    PubMed

    Wong, Philip; Pham, Roger; Whitely, Carl; Soto, Marcus; Salyers, Kevin; James, Christopher; Bruenner, Bernd A

    2011-11-01

    The goal of this work was to obtain full pharmacokinetic profiles from individual mice with the use of an automated blood sampling system and dried blood spot (DBS) technique. AMG 517, a potent and selective vanilloid receptor (VR1) antagonist, was dosed to mice (n=3) intravenously and blood samples were collected using the automated blood sampling system with the "no blood waste" method. The collected blood samples were a mixture of 25 μL blood and 50 μL of heparinized saline solution. Two 15 μL aliquots were manually spotted onto a DBS card and dried at room temperature for at least 2h before being stored in zip bags with desiccant. The remaining samples (45 μL) were stored at -70°C until analysis. Both the DBS and the whole blood samples (diluted with saline (1:2, v/v)) were extracted and analyzed by liquid chromatography-tandem mass spectrometry. The overall extraction recovery of the analyte from the dried blood spots was determined to be about 90%. The pharmacokinetic parameters calculated using the whole blood or the DBS concentration data were comparable, and were obtained from only 3 mice, whereas conventional sampling and analysis would have required up to 27 mice to achieve the same result. The analyte was shown to be stable in the diluted whole blood (blood:saline 1:2) at room temperature for at least 4h and in the DBS for at least 34 days when stored at room temperature. These results indicated that the automated blood sampling system and DBS collection are promising techniques to obtain full pharmacokinetic profiles from individual mice and reduce the use of animals. PMID:21784595

  2. Dried Blood Spot Collection of Health Biomarkers to Maximize Participation in Population Studies

    PubMed Central

    Ostler, Michael W.; Porter, James H.; Buxton, Orfeu M.

    2014-01-01

    Biomarkers are directly-measured biological indicators of disease, health, exposures, or other biological information. In population and social sciences, biomarkers need to be easy to obtain, transport, and analyze. Dried Blood Spots meet this need, and can be collected in the field with high response rates. These elements are particularly important in longitudinal study designs including interventions where attrition is critical to avoid, and high response rates improve the interpretation of results. Dried Blood Spot sample collection is simple, quick, relatively painless, less invasive then venipuncture, and requires minimal field storage requirements (i.e. samples do not need to be immediately frozen and can be stored for a long period of time in a stable freezer environment before assay). The samples can be analyzed for a variety of different analytes, including cholesterol, C-reactive protein, glycosylated hemoglobin, numerous cytokines, and other analytes, as well as provide genetic material. DBS collection is depicted as employed in several recent studies. PMID:24513728

  3. Dried blood spot in the genotyping, quantification and storage of HCV RNA: a systematic literature review

    PubMed Central

    Greenman, Jamie; Roberts, Teri; Cohn, Jennifer; Messac, Luke

    2015-01-01

    The entry of new all-oral direct acting antiviral therapy for hepatitis C provides an opportunity to scale up HCV care in low- and middle-income countries. In HIV, use of dried blood spots (DBS) has facilitated the diagnosis and management of HIV in resource-poor settings. DBS may be used in a similar way to facilitate diagnosis and management of HCV. Here, we present a systematic review of the literature of DBS for HCV RNA detection and genotyping. Using an a priori review protocol, three databases were searched for studies published up to August 2013 that reported the use of dried blood and serum spots in genotyping, detection and measurement of HCV RNA, as well as the rate of degradation of HCV RNA when stored in DBS at room temperature. Nine papers were eligible for inclusion; eight studied DBS and one dried serum. Two studies measured concordance between genotype and subtype determined by DBS and whole plasma and both found 100% concordance. Four studies measured endpoint detection limits of HCV RNA-positive samples by DBS and found positive predictive values of 100% down to 250, 334, 2500 and 24160 IU/mL. Two studies found deterioration of HCV RNA in DBS samples stored at room temperature, while two others failed to detect such deterioration. These results support the potential use of DBS for genotyping and HCV RNA detection. Studies of the use of DBS for HCV RNA viral load measurement and of the rate of degradation of HCV RNA when stored in DBS at ambient temperatures remain inconclusive. PMID:25367722

  4. Limited Utility of Dried-Blood- and Plasma Spot-Based Screening for Antiretroviral Treatment Failure with Cobas Ampliprep/TaqMan HIV-1 Version 2.0

    PubMed Central

    Shiningavamwe, Andreas; Chang, Joy; Maher, Andrew D.; Zhang, Guoqing; Yang, Chunfu; Gaeb, Esegiel; Kaura, Harold; Ellenberger, Dennis; Lowrance, David W.

    2014-01-01

    The 2013 WHO antiretroviral therapy (ART) guidelines recommend dried blood spots (DBS) as an alternative specimen type for viral load (VL) monitoring. We assessed the programmatic utility of screening for antiretroviral (ARV) treatment failure (TF) at 5,000 and 1,000 copies/ml using DBS and dried plasma spots (DPS) with a commonly used VL assay, the Roche Cobas Ampliprep/Cobas TaqMan V.2.0 (CAP/CTM). Plasma, DBS, and DPS were prepared from 839 whole-blood specimens collected from patients on ART for ≥6 months at three public facilities in Namibia. Using the CAP/CTM test, VL were measured in plasma, DBS, and DPS, and the results were compared using the plasma VL as the reference standard. The clinical sensitivities, specificities, and positive (PPV) and negative predictive values (NPV) of DBS at ARV TF diagnostic thresholds of 5,000 copies/ml and 1,000 copies/ml were 0.99, 0.55, 0.33, and 0.99 and 0.99, 0.26, 0.29, and 0.99, respectively, and for DPS at TF diagnostic thresholds of 5,000 copies/ml and 1,000 copies/ml, they were 0.88, 0.98, 0.92, and 0.97 and 0.91, 0.96, 0.89, and 0.97, respectively. The prevalences of TF were overestimated in DBS by 33% and 57% at these two thresholds, respectively. A high rate of false-positive results would occur if the CAP/CTM with DBS were to be used to screen for ARV TF. WHO recommendations for DBS-based VL monitoring should be specific to the VL assay version and type. Despite the better performance of DPS, the programmatic utility for TF screening may be limited by requirements for processing the whole blood at the collection site. PMID:25143579

  5. Diagnosis of Morquio Syndrome in Dried Blood Spots Based on a New MRM-MS Assay

    PubMed Central

    Cozma, Claudia; Eichler, Sabrina; Wittmann, Gyula; Flores Bonet, Alba; Kramp, Guido Johannes; Giese, Anne-Katrin; Rolfs, Arndt

    2015-01-01

    Background Mucopolysaccharidosis IVA (MPS IVA; Morquio A disease) is an autosomal recessive disease caused and characterized by a decreased activity of N-acetylgalactosamine-6-sulfate sulfatase (GALNS), resulting in accumulation of keratan sulfate and chondroitin-6-sulfate in tissues and secondary organ damage. Recently approved enzyme replacement therapy renders the easy and early identification of MPS IVA of out-most importance. Methodology We propose a completely new assay for the stable and reproducible detection of GALNS deficiency in dry blood spots (DBS). For the validation blood samples were taken from 59 healthy individuals and 24 randomly selected genetically confirmed MPS IVA patients. The material extracted from DBS was incubated with a 4-methylumbelliferyl-β-D-galactopyranoside-6-sulfate as a specific substrate. Final enzymatic product, 4-methylumbelliferone, obtained after adding exogenous beta-galactosidase, was quantified by LC/MRM-MS (liquid-chromatography/multiple-reaction-monitoring mass-spectrometry). 4-propyl-5-hydroxy-7-methyl-2h-chromen-2-one was used as internal standard, a compound with a similar molecular structure and fragmentation pattern in negative ion mode as 4-methylumbelliferone. Findings The enzymatic assay yielded a positive and negative predictive value of 1.0 for genetically confirmed MPS IVA patients (GALNS activity of 0.35 ± 0.21 μmol/L/h) and for controls with normal GALNS activity (23.1 ± 5.3 μmol/L /h). With present enzymatic conditions, the reaction yield in dried blood spots is at least 20 fold higher than any previously reported data with other assays. Interpretation The present LC/MRM-MS based assay for MPS IVA diagnosis provides an easy, highly-standardized, accurate and innovative quantification of the enzymatic product in vitro and distinguishes perfectly between MPS IVA affected patients and normal controls. This technique will significantly simplify the early detection of MPS IVA patients. PMID:26147980

  6. Alginate and chitosan foam combined with electromembrane extraction for dried blood spot analysis.

    PubMed

    Eibak, Lars Erik Eng; Hegge, Anne Bee; Rasmussen, Knut Einar; Pedersen-Bjergaard, Stig; Gjelstad, Astrid

    2012-10-16

    Samples of 10 μL of whole blood containing citalopram, loperamide, methadone, and sertraline as model substances were spotted on alginate and chitosan foams as sampling media. After drying and storage at room temperature, the punched out dried blood spot and the foam was dissolved in 300 μL of 1 mM HCl. With alginate foam as sampling medium, the analytes dissolved completely after 3 min. Enrichment and cleanup was performed with electromembrane extraction for 10 min. The analytes were collected in 21 μL of 10 mM formic acid as the acceptor phase, and the extracts were analyzed by liquid chromatography-mass spectrometry (LC-MS). Sample preparation of blood spots on commercial cards was also performed (Whatman FTA DMPK and Agilent Bond Elut DMS) using elution procedures recommended by the manufacturers. The recoveries obtained with the commercial cards were lower for most of the model analytes compared to the recoveries obtained with alginate and chitosan foams as sampling media. The procedure used for Agilent Bond Elut DMS showed higher recoveries than the procedure used for Whatman FTA DMPK-A, but the time needed for sample preparation was significantly longer (nearly 2 h). The stability of the model substances on the alginate foam was acceptable within 50 days of storage. The limit of quantification (LOQ) defined as S/N = 10, was 1.2, 5.5, 2.0, and 5.3 ng/mL for citalopram, loperamide, methadone, and sertraline, respectively. Linear calibration graphs were obtained in the range 17.5-560 ng/mL with r(2) values 0.983-0.995, and the relative standard deviations were below 20%. PMID:22991973

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

    PubMed

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

    2014-07-01

    The anti-estrogenic effect of tamoxifen is suggested to be mainly attributable to its metabolite (Z)-endoxifen, and a minimum therapeutic threshold for (Z)-endoxifen in serum has been proposed. The objective of this research was to establish the relationship between dried blood spot (DBS) and serum concentrations of tamoxifen and (Z)-endoxifen to allow the use of DBS sampling, a simple and patient-friendly alternative to venous sampling, in clinical practice. Paired DBS and serum samples were obtained from 50 patients using tamoxifen and analyzed using HPLC-MS/MS. Serum concentrations were calculated from DBS concentrations using the formula calculated serum concentration = DBS concentration/([1-haematocrit (Hct)] + blood cell-to-serum ratio × Hct). The blood cell-to-serum ratio was determined ex vivo by incubating a batch of whole blood spiked with both analytes. The average Hct for female adults was imputed as a fixed value. Calculated and analyzed serum concentrations were compared using weighted Deming regression. Weighted Deming regression analysis comparing 44 matching pairs of DBS and serum samples showed a proportional bias for both analytes. Serum concentrations were calculated using [Tamoxifen] serum, calculated  = [Tamoxifen] DBS /0.779 and [(Z)-Endoxifen] serum, calculated = [(Z)-Endoxifen] DBS /0.663. Calculated serum concentrations were within 20 % of analyzed serum concentrations in 84 and 100 % of patient samples for tamoxifen and (Z)-endoxifen, respectively. In conclusion, DBS concentrations of tamoxifen and (Z)-endoxifen were equal to serum concentrations after correction for Hct and blood cell-to-serum ratio. DBS sampling can be used in clinical practice. PMID:24859000

  8. Validation of the Use of Dried Blood Spot (DBS) Method to Assess Vitamin A Status

    PubMed Central

    Fallah, Elham; Peighambardoust, Seyed Hadi

    2012-01-01

    Background: Vitamin A deficiency is an important dietary deficiency in the world. Thus, the ne¬cessity of screening for deficient populations is obvious. This paper introduces a fast, cheap and relatively reliable method called “dried blood spot” (DBS) method in screening the deficient populations. The validity of this method for retinol measurement was investigated. Method: The “precision” and “agreement” criteria of the DBS method were assessed. The preci¬sion was calculated and compared with those of plasma using F-test. The agreement was eva¬luated using Bland-Altman plot. Results: The imprecision of retinol measurements in dried spots was not significantly different from those of the control (plasma). A good correlation coefficient (r2=0.78) was obtained for dried spots’ retinol measurements versus plasma’s retinol analysis (P < 0.01). Paired t-test showed no significant difference between the DBS and retinol methods on a group level. Imprecision of DBS measurement was acceptable, compared to that of the plasma method. The difference be¬tween these two methods was not statistically significant on a group level. Conclusion: Application of DBS standard samples, in which a part of the plasma was replaced with the artificial plasma, was shown to be a reliable calibration mean for retinol measurements in DBS samples. Retinol in dried spots was stable for 90 days. Overall, the DBS method provided a precise measurement of retinol, showing results that were comparable with the measurement of retinol in plasma. PMID:24688932

  9. Analysis of ochratoxin A in dried blood spots - Correlation between venous and finger-prick blood, the influence of hematocrit and spotted volume.

    PubMed

    Osteresch, Bernd; Cramer, Benedikt; Humpf, Hans-Ulrich

    2016-05-01

    We report the improvement of a method for the detection of ochratoxin A (OTA) and its thermal degradation product 2'R-ochratoxin A in dried blood spots (DBS) by high performance liquid chromatographic (HPLC) tandem mass spectrometry (MS/MS). The DBS technique was advanced for the analysis of these two compounds in DBS with unknown amounts of blood as well as varying hematocrit values. Furthermore the comparability of venous vs. capillary blood was investigated. Human whole blood samples were spotted, dried, and extracted with a solvent consisting of acetone, acetonitrile and water for analysis by HPLC-MS/MS. Quantification was carried out by stable isotope labelled internal standards. Blood samples of volunteers (n=50) were used to further optimize and simplify the procedure. Ochratoxin A and 2'R-ochratoxin A concentrations found in the entire spots (approx. 100 μL blood) were compared with punched DBS discs of 8.8mm size containing approximately 20 μL blood. As a result the amounts of both toxins in a punched 8.8mm disc correlate well with the entire DBS. Also the use of capillary blood from finger-pricks versus venous blood was evaluated. The analyte levels correlate as well indicating that the less invasive finger-prick sampling gives also reliable results. The influence of hematocrit was investigated in a range of 25-55% according to the hematocrit in the used real blood samples (34-46% hematocrit). However no significant hematocrit effect was observed for the utilized real blood samples. Moreover different blood volumes were spotted and punched as a minimal spot size is usually recommended for accurate analysis. In this experiment finger-prick samples typically consist of about 90 μL blood. Therefore spots of 75, 100 and 125 μL blood were prepared and analyzed. Similar to the hematocrit effect, no considerable influence was observed. PMID:27046696

  10. Field Evaluation of Capillary Blood Samples as a Collection Specimen for the Rapid Diagnosis of Ebola Virus Infection During an Outbreak Emergency

    PubMed Central

    Strecker, Thomas; Palyi, Bernadett; Ellerbrok, Heinz; Jonckheere, Sylvie; de Clerck, Hilde; Bore, Joseph Akoi; Gabriel, Martin; Stoecker, Kilian; Eickmann, Markus; van Herp, Michel; Formenty, Pierre; Di Caro, Antonino; Becker, Stephan

    2015-01-01

    Background. Reliable reverse transcription polymerase chain reaction (RT-PCR)–based diagnosis of Ebola virus infection currently requires a blood sample obtained by intravenous puncture. During the current Ebola outbreak in Guinea, we evaluated the usability of capillary blood samples collected from fingersticks of patients suspected of having Ebola virus disease (EVD) for field diagnostics during an outbreak emergency. Methods. A total of 120 venous and capillary blood samples were collected from 53 patients admitted to the Ebola Treatment Centre in Guéckédou, Guinea, between July and August 2014. All sample specimens were analyzed by RT-PCR using the RealStar Filovirus Screen RT-PCR Kit 1.0 from altona Diagnostics (Germany). We compared samples obtained by venipuncture and those obtained by capillary blood sampling absorbed onto swab devices. Results. The resulting sensitivity and specificity of tests performed with capillary blood samples were 86.8% (95% confidence interval [CI], 71.9%–95.6%; 33/38 patients) and 100% (95% CI, 84.6%–100%; 22/22 patients), respectively. Conclusions. Our data suggest that capillary blood samples could serve as an alternative to venous blood samples for the diagnosis of EVD in resource-limited settings during a crisis. This can be of particular advantage in cases when venipuncture is difficult to perform—for example, with newborns and infants or when adult patients reject venipuncture for cultural or religious reasons. PMID:25991465

  11. Bridging the gap between sample collection and laboratory analysis: using dried blood spots to identify human exposure to chemical agents

    NASA Astrophysics Data System (ADS)

    Hamelin, Elizabeth I.; Blake, Thomas A.; Perez, Jonas W.; Crow, Brian S.; Shaner, Rebecca L.; Coleman, Rebecca M.; Johnson, Rudolph C.

    2016-05-01

    Public health response to large scale chemical emergencies presents logistical challenges for sample collection, transport, and analysis. Diagnostic methods used to identify and determine exposure to chemical warfare agents, toxins, and poisons traditionally involve blood collection by phlebotomists, cold transport of biomedical samples, and costly sample preparation techniques. Use of dried blood spots, which consist of dried blood on an FDA-approved substrate, can increase analyte stability, decrease infection hazard for those handling samples, greatly reduce the cost of shipping/storing samples by removing the need for refrigeration and cold chain transportation, and be self-prepared by potentially exposed individuals using a simple finger prick and blood spot compatible paper. Our laboratory has developed clinical assays to detect human exposures to nerve agents through the analysis of specific protein adducts and metabolites, for which a simple extraction from a dried blood spot is sufficient for removing matrix interferents and attaining sensitivities on par with traditional sampling methods. The use of dried blood spots can bridge the gap between the laboratory and the field allowing for large scale sample collection with minimal impact on hospital resources while maintaining sensitivity, specificity, traceability, and quality requirements for both clinical and forensic applications.

  12. Rapid Genotyping of Cytomegalovirus in Dried Blood Spots by Multiplex Real-Time PCR Assays Targeting the Envelope Glycoprotein gB and gH Genes

    PubMed Central

    Wessels, Els; Korver, Anna M. H.; van der Eijk, Annemiek A.; Rusman, Lisette G.; Kroes, Aloys C. M.; Vossen, Ann C. T. M.

    2012-01-01

    Genotyping of cytomegalovirus (CMV) is useful to examine potential differences in the pathogenicity of strains and to demonstrate coinfection with multiple strains involved in CMV disease in adults and congenitally infected newborns. Studies on genotyping of CMV in dried blood spots (DBS) are rare and have been hampered by the small amount of dried blood available. In this study, two multiplex real-time PCR assays for rapid gB and gH genotyping of CMV in DBS were developed. Validation of the assays with 39 CMV-positive plasma samples of transplant recipients and 21 urine specimens of congenitally infected newborns was successful in genotyping 100% of the samples, with gB1 and gB3 being the most prevalent genotypes. Multiple gB and gH genotypes were detected in 36% and 33% of the plasma samples, respectively. One urine sample from a newborn with symptomatic congenital CMV was positive for gB1 and gB2. DBS of congenitally infected newborns (n = 41) were tested using 9 μl of dried blood, and genotypes were detected in 81% (gB) and 73% (gH) of the samples, with gB3 being the most prevalent genotype. No clear association of specific genotypes with clinical outcome was observed. In conclusion, the CMV gB and gH PCR assays were found to be rapid, sensitive for detecting mixed infections, and suitable for direct usage on DBS. These assays are efficient tools for genotyping of CMV in DBS of congenitally infected newborns. PMID:22116158

  13. Detection of hemagglutinins in dried saliva stains and their potential use in blood typing.

    PubMed

    Harrington, J J; Martin, R; Kobilinsky, L

    1988-05-01

    Since 1928, hemagglutinins have been known to exist in saliva; however, they have not been utilized as evidence in criminal investigations because in the past, techniques for measuring them have not been sufficiently sensitive. In this paper we describe improved techniques for detecting salivary hemagglutinins and report initial results obtained with these methods. The stability of salivary hemagglutinins at several different temperatures was examined in liquid samples and in dried stains on filter paper, cigarette butts, and envelope flaps. Our observations indicate that salivary hemagglutinins may be sufficiently stable, over periods of one to several days at ambient room temperatures, to be of value to forensic science investigators. The results of the hemagglutinin assay are not affected by the age or sex of the sample donor. Because salivary hemagglutinins can be used to determine ABO blood type, analyses of this kind can serve as an important confirmatory test which the forensic serologist can use in conjunction with salivary agglutinogen determinations. PMID:3385376

  14. Use of dried blood spots to define antibody response to the Strongyloides stercoralis recombinant antigen NIE

    PubMed Central

    Mounsey, Kate; Kearns, Therese; Rampton, Melanie; Llewellyn, Stacey; King, Mallory; Holt, Deborah; Currie, Bart J.; Andrews, Ross; Nutman, Thomas; McCarthy, James

    2015-01-01

    An approach to improve the diagnosis of Strongyloides stercoralis infection is the use of serologic assays utilising the NIE antigen from S. stercoralis, with good diagnostic sensitivity and excellent specificity reported. Detection of antibody eluted from dried blood spots (DBS) has shown utility in large-scale seroepidemiological studies for a range of conditions and is appealing for use with children where sample collection is difficult. We adapted an existing NIE-enzyme linked immunosorbent assay (ELISA) for the testing of strongyloides antibody response on DBS, and evaluated it in a population screening and mass drug administration programme (MDA) for strongyloidiasis conducted in an Australian indigenous community. Study participants were treated with 200 μg/kg ivermectin (>15 kg) or 3× 400 mg albendazole (<15 kg). The sensitivity of the NIE DBS-ELISA was determined by receiver operator characteristic (ROC) analysis to be 85.7%. A total of 214 DBS were collected from 184 participants across two screening and MDA encounters. A total of 27 of 164 participants (16.5%) tested positive for S. stercoralis NIE-DBS prior to MDA treatment, and 6 of 50 participants (12.0%) tested positive after treatment. These prevalence values are similar to those documented by standard serology in the same community. For 30 participants where a DBS was collected at both MDA 1 and 2, a significant decline in ELISA values was evident post treatment (0.12–0.02, p = 0.0012). These results are in agreement with previous studies documenting the high seroprevalence of S. stercoralis in remote Australian Indigenous communities, and suggest that collection of dried blood spots may be a useful approach for field diagnosis of S. stercoralis seroprevalence. PMID:25051188

  15. Sulfatide Analysis by Mass Spectrometry for Screening of Metachromatic Leukodystrophy in Dried Blood and Urine Samples

    PubMed Central

    Spacil, Zdenek; Kumar, Arun Babu; Liao, Hsuan-Chieh; Auray-Blais, Christiane; Stark, Samantha; Suhr, Teryn R.; Scott, C. Ronald; Turecek, Frantisek; Gelb, Michael H.

    2016-01-01

    BACKGROUND Metachromatic leukodystrophy (MLD) is an autosomal recessive disorder caused by deficiency in arylsulfatase A activity, leading to accumulation of sulfatide substrates. Diagnostic and monitoring procedures include demonstration of reduced arylsulfatase A activity in peripheral blood leukocytes or detection of sulfatides in urine. However, the development of a screening test is challenging because of instability of the enzyme in dried blood spots (DBS), the widespread occurrence of pseudodeficiency alleles, and the lack of available urine samples from newborn screening programs. METHODS We measured individual sulfatide profiles in DBS and dried urine spots (DUS) from MLD patients with LC-MS/MS to identify markers with the discriminatory power to differentiate affected individuals from controls. We also developed a method for converting all sulfatide molecular species into a single species, allowing quantification in positive-ion mode upon derivatization. RESULTS In DBS from MLD patients, we found up to 23.2-fold and 5.1-fold differences in total sulfatide concentrations for early- and late-onset MLD, respectively, compared with controls and pseudodeficiencies. Corresponding DUS revealed up to 164-fold and 78-fold differences for early- and late-onset MLD patient samples compared with controls. The use of sulfatides converted to a single species simplified the analysis and increased detection sensitivity in positive-ion mode, providing a second option for sulfatide analysis. CONCLUSIONS This study of sulfatides in DBS and DUS suggests the feasibility of the mass spectrometry method for newborn screening of MLD and sets the stage for a larger-scale newborn screening pilot study. PMID:26585924

  16. Use of dried blood spots to define antibody response to the Strongyloides stercoralis recombinant antigen NIE.

    PubMed

    Mounsey, Kate; Kearns, Therese; Rampton, Melanie; Llewellyn, Stacey; King, Mallory; Holt, Deborah; Currie, Bart J; Andrews, Ross; Nutman, Thomas; McCarthy, James

    2014-10-01

    An approach to improve the diagnosis of Strongyloides stercoralis infection is the use of serologic assays utilising the NIE antigen from S. stercoralis, with good diagnostic sensitivity and excellent specificity reported. Detection of antibody eluted from dried blood spots (DBS) has shown utility in large-scale seroepidemiological studies for a range of conditions and is appealing for use with children where sample collection is difficult. We adapted an existing NIE-enzyme linked immunosorbent assay (ELISA) for the testing of strongyloides antibody response on DBS, and evaluated it in a population screening and mass drug administration programme (MDA) for strongyloidiasis conducted in an Australian indigenous community. Study participants were treated with 200 μg/kg ivermectin (>15 kg) or 3× 400 mg albendazole (<15kg). The sensitivity of the NIE DBS-ELISA was determined by receiver operator characteristic (ROC) analysis to be 85.7%. A total of 214 DBS were collected from 184 participants across two screening and MDA encounters. A total of 27 of 164 participants (16.5%) tested positive for S. stercoralis NIE-DBS prior to MDA treatment, and 6 of 50 participants (12.0%) tested positive after treatment. These prevalence values are similar to those documented by standard serology in the same community. For 30 participants where a DBS was collected at both MDA 1 and 2, a significant decline in ELISA values was evident post treatment (0.12-0.02, p=0.0012). These results are in agreement with previous studies documenting the high seroprevalence of S. stercoralis in remote Australian Indigenous communities, and suggest that collection of dried blood spots may be a useful approach for field diagnosis of S. stercoralis seroprevalence. PMID:25051188

  17. Second-tier test for quantification of underivatized amino acids in dry blood spot for metabolic diseases in newborn screening.

    PubMed

    Wang, Chunyan; Zhu, Hongbin; Zhang, Wenyan; Song, Fengrui; Liu, Zhiqiang; Liu, Shuying

    2013-02-01

    The quantitative analysis of amino acids (AAs) in single dry blood spot (DBS) samples is an important issue for metabolic diseases as a second-tier test in newborn screening. An analytical method for quantifying underivatized AAs in DBS was developed by using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). The sample preparation in this method is simple and ion-pairing agent is not used in the mobile phase that could avoid ion suppression, which happens in mass spectrometry and avoids damage to the column. Through chromatographic separation, some isomeric compounds could be identified and quantified, which cannot be solved through only appropriate multiple reactions monitoring transitions by MS/MS. The concentrations of the different AAs were determined using non-deuterated internal standard. All calibration curves showed excellent linearity within test ranges. For most of the amino acids the accuracy of extraction recovery was between 85.3 and 115 %, and the precision of relative standard deviation was <7.0 %. The 35 AAs could be identified in DBS specimens by the developed LC-MS/MS method in 17-19 min, and eventually 24 AAs in DBS were quantified. The results of the present study prove that this method as a second-tier test in newborn screening for metabolic diseases could be performed by the quantification of free AAs in DBS using the LC-MS/MS method. The assay has advantages of high sensitive, specific, and inexpensive merits because non-deuterated internal standard and acetic acid instead of ion-pairing agent in mobile phase are used in this protocol. PMID:22932943

  18. The opisthonephric blood vascular system of the chicken embryo as studied by scanning electron microscopy of microvascular corrosion casts and critical point dried preparations.

    PubMed

    Ditrich, H; Splechtna, H

    1989-06-01

    Microvascular corrosion casts of chicken embryos between four and 19 days after fertilization have been prepared. The developing kidney was investigated with scanning electron microscopy (SEM). The injection technique and resin composition were modified in order to facilitate the complete replication of native blood vascular systems of specimens as small as 15 mm body length. The development of the opisthonephros was followed from near the beginning of its function until a vascular development comparable to the adult situation was reached. Critical point dried glomeruli show the differentiation of the glomerular visceral epithelium (podocytes) from initially epithelioid to highly branched forms. The embryonic kidney (cranial part of the opisthonephros-mesonephros) shows a construction-principle resembling amphibians that is entirely different from the definitive excretory organ (caudal part of the opisthonephros-metanephros). PMID:2814402

  19. Specimen shipment

    USGS Publications Warehouse

    Franson, J. Christian

    1987-01-01

    Procedures for shipping specimens vary with different disease diagnostic laboratories. Therefore, it is important to contact the receiving laboratory and obtain specific instructions. This will facilitate processing of specimens when they reach the laboratory and assure that the quality of the specimens is not compromised. Time spent on field investigation, specimen collection, and obtaining an adequate history will be of little clue is specimens become contaminated, decomposed, or otherwise spoiled enroute to the diagnostic laboratory. There are five bases of proper specimen shipment: (1) prevent cross-contamination from specimen to specimen, (2) prevent decomposition of the specimen, (3) prevent leakage of fluids, (4) preserve individual specimen identity, and (5) properly label the package. Basic supplies needed for specimen shipment are shown in Fig. 3.1.

  20. Does volumetric absorptive microsampling eliminate the hematocrit bias for caffeine and paraxanthine in dried blood samples? A comparative study.

    PubMed

    De Kesel, Pieter M M; Lambert, Willy E; Stove, Christophe P

    2015-06-30

    Volumetric absorptive microsampling (VAMS) is a novel sampling technique that allows the straightforward collection of an accurate volume of blood (approximately 10μL) from a drop or pool of blood by dipping an absorbent polymeric tip into it. The resulting blood microsample is dried and analyzed as a whole. The aim of this study was to evaluate the potential of VAMS to overcome the hematocrit bias, an important issue in the analysis of dried blood microsamples. An LC-MS/MS method for analysis of the model compounds caffeine and paraxanthine in VAMS samples was fully validated and fulfilled all pre-established criteria. In conjunction with previously validated procedures for dried blood spots (DBS) and blood, this allowed us to set up a meticulous comparative study in which both compounds were determined in over 80 corresponding VAMS, DBS and liquid whole blood samples. These originated from authentic human patient samples, covering a wide hematocrit range (0.21-0.50). By calculating the differences with reference whole blood concentrations, we found that analyte concentrations in VAMS samples were not affected by a bias that changed over the evaluated hematocrit range, in contrast to DBS results. However, VAMS concentrations tend to overestimate whole blood concentrations, as a consistent positive bias was observed. A different behavior of VAMS samples prepared from incurred and spiked blood, combined with a somewhat reduced recovery of caffeine and paraxanthine from VAMS tips at high hematocrit values, an effect that was not observed for DBS using a very similar extraction procedure, was found to be at the basis of the observed VAMS-whole blood deviations. Based on this study, being the first in which the validity and robustness of VAMS is evaluated by analyzing incurred human samples, it can be concluded that VAMS effectively assists in eliminating the effect of hematocrit. PMID:26041521

  1. Quantitation of brinzolamide in dried blood spots by a novel LC-QTOF-MS/MS method.

    PubMed

    Foivas, Anargyros; Malenović, Anđelija; Kostić, Nađa; Božić, Marija; Knežević, Miroslav; Loukas, Yannis L; Dotsikas, Yannis

    2016-02-01

    In the current study, a rapid and sensitive LC-QTOF-MS/MS method for the determination of brinzolamide in dried blood spots (DBS) was developed and validated. This novel sample collection, storage and transfer technique was suitable for analyzing a drug with high distribution into red blood cells and negligible plasma levels. The method included an isocratic mobile phase consisting of methanol and 10mM ammonium formate (90:10, v/v) and detection in positive electrospray mode (ESI+). The flow rate was adjusted to 0.350mL/min yielding retention times of 1.7min for both brinzolamide and internal standard (IS) rabeprazole on a Cyano analytical column, respectively. The validation of the proposed method over the concentration range 0.500-20.0μg/mL was performed in compliance with EMEA and FDA guidelines, assessing all major performance characteristics. Inter- and intra- assay precisions were less than 14%, while inter- and intra- assay accuracies varied from 92.2 to 111%. No matrix effect was observed and the mean brinzolamide extraction recovery was 93.5%. The method was successfully applied to real DBS samples from patients in steady state condition, receiving brinzolamide ophthalmic suspension 1% (w/v) for several months. Initial concentrations were corrected due to hematocrit effect, using image processing algorithm written in Matlab. PMID:26669612

  2. Use of dried blood spots in doping control analysis of anabolic steroid esters.

    PubMed

    Tretzel, Laura; Thomas, Andreas; Geyer, Hans; Gmeiner, Günter; Forsdahl, Guro; Pop, Valentin; Schänzer, Wilhelm; Thevis, Mario

    2014-08-01

    Dried blood spot (DBS) sampling, a technique for whole blood sampling on a piece of filter paper, has more than 50-years tradition, particularly in the diagnostic analysis of metabolic disorders in neonatal screening. Due to the minimal invasiveness, straightforwardness, robustness against manipulation and fastness DBS sampling recommends itself as an advantageous technique in doping control analysis. The present approach highlights the development of a screening assay for the analysis of eight anabolic steroid esters (nandrolone phenylpropionate, trenbolone enanthate, testosterone acetate, testosterone cypionate, testosterone isocaproate, testosterone phenylpropionate, testosterone decanoate and testosterone undecanoate) and nandrolone in DBS. The detection of the intact esters allows an unequivocal proof of the administration of conjugates of exogenous testosterone and its derivatives. Precise, specific and linear conditions were obtained by means of liquid chromatography high resolution/high accuracy mass spectrometry. Sensitivity in the low ppb range was accomplished by the preparation of the methyloxime derivatives of the target compounds. Labeled internal standards (d3-nandrolone, d3-nandrolone caproate and d3-nandrolone undecanoate) were applied to compensate for the broad range in chain length of the esters. The assay presented here outlines the application of DBS for the analysis of anabolic steroid esters in doping controls for the first time providing great potential to simplify the proof of exogenous administration of testosterone. PMID:24713476

  3. Heart rate and blood pressure time courses during prolonged dry apnoea in breath-hold divers.

    PubMed

    Perini, Renza; Tironi, Adelaide; Gheza, Alberto; Butti, Ferdinando; Moia, Christian; Ferretti, Guido

    2008-09-01

    To define the dynamics of cardiovascular adjustments to apnoea, beat-to-beat heart rate (HR) and blood pressure and arterial oxygen saturation (SaO(2)) were recorded during prolonged breath-holding in air in 20 divers. Apnoea had a mean duration of 210 +/- 70 s. In all subjects, HR attained a value 14 beats min(-1) lower than control within the initial 30 s (phase I). HR did not change for the following 2-2.5 min (phase II). Then, nine subjects interrupted the apnoea (group A), whereas 11 subjects (group B) could prolong the breath-holding for about 100 s, during which HR continuously decreased (phase III). In both groups, mean blood pressure was 8 mmHg above control at the end of phase I; it then further increased by additional 12 mmHg at the end of the apnoea. In both groups, SaO(2) did not change in the initial 100-140 s of apnoea; then, it decreased to 95% at the end of phase II. In group B, SaO(2) further diminished to 84% at the end of phase III. A typical pattern of cardiovascular readjustments was identified during dry apnoea. This pattern was not compatible with a role for baroreflexes in phase I and phase II. Further readjustment in group B may imply a role for both baroreflexes and chemoreflexes. Hypothesis has been made that the end of phase II corresponds to physiological breakpoint. PMID:18496707

  4. Robust measurement of vitamin A status in plasma and blood dried on paper.

    PubMed

    Huang, Yichao; Clements, Peter Roy; Gibson, Robert Alan

    2015-12-01

    Vitamin A deficiency is the leading cause of preventable blindness in children and increases the risk of disease and death from severe infections. In addition, fat soluble vitamin A and associated retinoids directly regulate the expression of genes involved in fatty acid metabolism. Conventional methods for measuring vitamin A involve venipuncture, centrifugation and refrigeration all of which make measuring vitamin A in nutritional surveys expensive. We aimed to develop a simple and robust system for measurement of retinol (biomarker for vitamin A) using dried blood spot (DBS) samples. Low recoveries and inconsistent results reported by others were found to be due to poor extraction efficiency rather than retinol instability. Maintaining acid conditions during extraction resulted in recoveries >95% with <6.5% of coefficient of variation. Using isocratic high performance liquid chromatography, separation was achieved in <3.5 min. Detector response was linear (R(2)=0.9939) within a range of 0.05-2 μg/mL, with a limit of quantification of 0.05 μg/mL. Retinol in DBS was shown to be stable (>95%) at room temperature for up to 10 weeks. DBS values for retinol were highly correlated with venous blood samples from 24 healthy subjects (r=0.9724) and were consistent with results from a commercial laboratory. This simple and reliable method for the determination of vitamin A status should prove particularly valuable for population studies and large clinical trials. PMID:26489594

  5. Evaluation of DNA extraction methods for the detection of Cytomegalovirus in dried blood spots

    PubMed Central

    Koontz, D.; Baecher, K.; Amin, M.; Nikolova, S.; Gallagher, M.; Dollard, S.

    2015-01-01

    Background Dried blood spots (DBS) are collected universally from newborns and may be valuable for the diagnosis of congenital Cytomegalovirus (CMV) infection. The reported analytical sensitivity for DBS testing compared to urine or saliva varies greatly across CMV studies. The purpose of this study was to directly compare the performance of various DNA extraction methods for identification of CMV in DBS including those used most often in CMV studies. Study design Whatman® Grade 903 filter paper cards were spotted with blood samples from 25 organ transplant recipients who had confirmed CMV viremia. Six DNA extraction methods were compared for relative yield of viral and cellular DNA: 2 manual solution-based methods (Gentra Puregene, thermal shock), 2 manual silica column-based methods (QIAamp DNA Mini, QIAamp DNA Investigator), and 2 automated methods (M48 MagAttract Mini, QIAcube Investigator). DBS extractions were performed in triplicate followed by real-time quantitative PCR (qPCR). Results For extraction of both viral and cellular DNA, two methods (QIAamp DNA Investigator and thermal shock) consistently gave the highest yields, and two methods (M48 MagAttract Mini and QIAamp DNA Mini) consistently gave the lowest yields. There was an average 3-fold difference in DNA yield between the highest and lowest yield methods. Conclusion The choice of DNA extraction method is a major factor in the ability to detect low levels of CMV in DBS and can largely account for the wide range of DBS sensitivities reported in studies to date. PMID:25866346

  6. Screening for late neonatal vitamin K deficiency by acarboxyprothrombin in dried blood spots.

    PubMed Central

    Motohara, K; Endo, F; Matsuda, I

    1987-01-01

    Acarboxyprothrombin (protein induced by vitamin K absence or antagonist-II (PIVKA-II] concentrations in dried blood spots were determined in 19,029 infants at about 1 month of age as an indicator of vitamin K deficiency. We observed 51 cases with raised blood concentrations of PIVKA-II (greater than 4 AU/ml), nine of whom showed very high concentrations (greater than 20 AU/ml). For infants who did not receive vitamin K prophylaxis at birth, the incidence of the PIVKA-II test yielding positive results was significantly higher in those solely breast fed (0.51%) compared with those fed formula milk (0.18%). Among solely breast fed infants, the incidence of a very high result of the PIVKA-II test was 0.14% in those who had not received vitamin K prophylaxis at birth, 0.04% in those who received 2 mg orally, and 0.03% in those who received 2 mg orally plus a further dose of 2-4 mg orally at 7 days. Thus vitamin K prophylaxis at birth did not completely prevent vitamin K deficiency at 1 month. We administered vitamin K therapeutically to all infants whose PIVKA-II test yielded a positive result at 1 month. Only one infant with a positive result developed late neonatal intracranial haemorrhage. PMID:3592727

  7. Multiplexed Quantitation of Endogenous Proteins in Dried Blood Spots by Multiple Reaction Monitoring - Mass Spectrometry

    PubMed Central

    Chambers, Andrew G.; Percy, Andrew J.; Yang, Juncong; Camenzind, Alexander G.; Borchers, Christoph H.

    2013-01-01

    Dried blood spot (DBS) sampling, coupled with multiple reaction monitoring mass spectrometry (MRM-MS), is a well-established approach for quantifying a wide range of small molecule biomarkers and drugs. This sampling procedure is simpler and less-invasive than those required for traditional plasma or serum samples enabling collection by minimally trained personnel. Many analytes are stable in the DBS format without refrigeration, which reduces the cost and logistical challenges of sample collection in remote locations. These advantages make DBS sample collection desirable for advancing personalized medicine through population-wide biomarker screening. Here we expand this technology by demonstrating the first multiplexed method for the quantitation of endogenous proteins in DBS samples. A panel of 60 abundant proteins in human blood was targeted by monitoring proteotypic tryptic peptides and their stable isotope-labeled analogs by MRM. Linear calibration curves were obtained for 40 of the 65 peptide targets demonstrating multiple proteins can be quantitatively extracted from DBS collection cards. The method was also highly reproducible with a coefficient of variation of <15% for all 40 peptides. Overall, this assay quantified 37 proteins spanning a range of more than four orders of magnitude in concentration within a single 25 min LC/MRM-MS analysis. The protein abundances of the 33 proteins quantified in matching DBS and whole blood samples showed an excellent correlation, with a slope of 0.96 and an R2 value of 0.97. Furthermore, the measured concentrations for 80% of the proteins were stable for at least 10 days when stored at −20 °C, 4 °C and 37 °C. This work represents an important first step in evaluating the integration of DBS sampling with highly-multiplexed MRM for quantitation of endogenous proteins. PMID:23221968

  8. Performance and storage integrity of dried blood spots for PCB, BFR and pesticide measurements.

    PubMed

    Batterman, Stuart; Chernyak, Sergei

    2014-10-01

    Dried blood spots (DBSs) can provide accurate and valuable estimates of exposure to environmental toxicants, and the use of information derived from archived newborn DBSs has enormous potential to open up new research on the impacts of early chemical exposure on disease. Broad application of DBS for the purpose of quantitative exposure estimation requires robust and validated methods. This study investigates the suitability of DBS analyses for population studies of exposure to three chemical groups: polychlorinated biphenyls (PCBs), brominated flame retardants (BFRs), and chlorinated pesticides. It examines background (matrix) contamination, recovery and extraction variability, sensitivity, and storage stability. DBS samples prepared using 50 μL of adult blood were analyzed by GC/MS, and method performance was confirmed by using certified materials and paired DBS-blood samples from six volunteers. Several of the target compounds and their degradation products have not been previously measured in DBSs. All target compounds were detected in DBS samples collected from the volunteers. Sample DBS cards showed background contamination of several compounds. When stored at room temperature, target compounds, excluding PBDEs, were stable for up to one month. When refrigerated or frozen, stability was acceptable for all compounds up to one year, and multiyear storage appears acceptable at colder (e.g., -80°C) temperatures. Multicompartment models may be used to estimate or correct for storage losses. Considering concentrations of contaminants for adults and children reported in the literature, and experimental values of detection limits and background contamination, DBS samples are suitable for quantifying exposures to many PCBs, BFRs and persistent pesticides. PMID:25058892

  9. Performance and Storage Integrity of Dried Blood Spots for PCB, BFR and Pesticide Measurements

    PubMed Central

    Batterman, Stuart; Chernyak, Sergei

    2014-01-01

    Dried blood spots (DBS) can provide accurate and valuable estimates of exposure to environmental toxicants, and the use of information derived from archived newborn DBS information has enormous potential to open up new research on the impacts of early chemical exposure on disease. Broad application of DBS for the purpose of quantitative exposure estimation requires robust and validated methods. This study investigates the suitability of DBS analyses for population studies of exposure to three chemical groups: polychlorinated biphenyls (PCBs), brominated flame retardants (BFRs), and chlorinated pesticides. It examines background (matrix) contamination, recovery and extraction variability, sensitivity, and storage stability. DBS samples prepared using 50 μL of adult blood were analyzed by GC/MS, and method performance was confirmed by using certified materials and paired DBS-blood samples from six volunteers. Several of the target compounds and their degradation products have not been previously measured in DBS. All target compounds were detected in DBS samples collected from the volunteers. Sample DBS cards showed background contamination of several compounds. When stored at room temperature, target compounds, excluding PBDEs, were stable for up to one month. When refrigerated or frozen, stability was acceptable for all compounds up to one year, and multiyear storage appears acceptable at colder (e.g., −80 °C) temperatures. Multicompartment models may be used to estimate or correct for storage losses. Considering concentrations of contaminants for adults and children reported in the literature, and experimental values of detection limits and background contamination, DBS samples are suitable for quantifying exposures to many PCBs, BFRs and persistent pesticides. PMID:25058892

  10. Dried blood spots versus plasma for the quantitation of HIV-1 RNA using a real-Time PCR, m2000rt assay.

    PubMed

    Vidya, Madhavan; Saravanan, Shanmugam; Rifkin, Samara; Solomon, Sunil S; Waldrop, Greer; Mayer, Kenneth H; Solomon, Suniti; Balakrishnan, Pachamuthu

    2012-05-01

    High costs and stringent requirements for storage and transport of plasma, often prohibit the availability of HIV viral load quantification in resource-limited settings. Dried blood spots (DBS) represent a better method of specimen collection that removes many of these logistical and technical limitations. The present study aimed to assess the performance of the Abbott m2000rt assay for quantitation of HIV-1 RNA in DBS specimens using plasma as a "gold standard" for comparison. One hundred paired DBS and plasma specimens were collected from patients infected with HIV, who were 18 years and older during routine visits to a private tertiary-care clinic in Chennai, India. HIV-1 RNA was extracted manually and then detected using the m2000rt assay. The mean plasma and DBS viral loads were 4.27 (95% CI: 2.65, 5.88) and 4.14 (95% CI: 1.96, 6.32) log copies/mL, respectively. The overall sensitivity of DBS reached 95%; with sensitivities of 62%, 88% and 100% when stratified by viral load ranges of ≤1000, 1000-3000 and >3000 copies/mL, respectively. An over quantitation of the viral load with DBS was observed in pairs with plasma viral load<3000 copies/mL [d=-0.3 log copies/mL (ranging from -0.1 to 0.6 log copies/mL)]. The study showed a strong concordance in RNA levels between plasma and DBS. The use of DBS specimens should be considered for HIV monitoring and for detection of virologic failure in resource-limited settings. PMID:22401801

  11. Impact of the phlebotomy training based on CLSI/NCCLS H03-A6 - procedures for the collection of diagnostic blood specimens by venipuncture.

    PubMed Central

    Lima-Oliveira, Gabriel; Lippi, Giuseppe; Salvagno, Gian Luca; Montagnana, Martina; Picheth, Geraldo; Guidi, Gian Cesare

    2012-01-01

    Introduction: The activities involving phlebotomy, a critical task for obtaining diagnostic blood samples, are poorly studied as regards the major sources of errors and the procedures related to laboratory quality control. The aim of this study was to verify the compliance with CLSI documents of clinical laboratories from South America and to assess whether teaching phlebotomists to follow the exact procedure for blood collection by venipuncture from CLSI/NCCLS H03-A6 - Procedures for the Collection of Diagnostic Blood Specimens by Venipuncture might improve the quality of the process. Materials and methods: A survey was sent by mail to 3674 laboratories from South America to verify the use of CLSI documents. Thirty skilled phlebotomists were trained with the CLSI H03-A6 document to perform venipuncture procedures for a period of 20 consecutive working days. The overall performances of the phlebotomists were further compared before and after the training program. Results: 2622 from 2781 laboratories that did answer our survey used CLSI documents to standardize their procedures and process. The phlebotomists’ training for 20 days before our evaluation completely eliminated non-conformity procedures for: i) incorrect friction of the forearm, during the cleaning of the venipuncture site to ease vein location; ii) incorrect sequence of vacuum tubes collection; and iii) inadequate mixing of the blood in primary vacuum tubes containing anticoagulants or clot activators. Unfortunately the CLSI H03-A6 document does not caution against both unsuitable tourniquet application time (i.e., for more than one minute) and inappropriate request to clench the fist repeatedly. These inadequate procedures were observed for all phlebotomists. Conclusion: We showed that strict observance of the CLSI H03-A6 document can remarkably improve quality, although the various steps for collecting diagnostic blood specimens are not a gold standard, since they may still permit errors. Tourniquet

  12. Sr/Ca proxy sea-surface temperature reconstructions from modern and holocene Montastraea faveolata specimens from the Dry Tortugas National Park

    USGS Publications Warehouse

    Flannery, Jennifer A.; Poore, Richard Z.

    2013-01-01

    Sr/Ca ratios from skeletal samples from two Montastraea faveolata corals (one modern, one Holocene, ~6 Ka) from the Dry Tortugas National Park were measured as a proxy for sea-surface temperature (SST). We sampled coral specimens with a computer-driven triaxial micromilling machine, which yielded an average of 15 homogenous samples per annual growth increment. We regressed Sr/Ca values from resulting powdered samples against a local SST record to obtain a calibration equation of Sr/Ca = -0.0392 SST + 10.205, R = -0.97. The resulting calibration was used to generate a 47-year modern (1961-2008) and a 7-year Holocene (~6 Ka) Sr/Ca subannually resolved proxy record of SST. The modern M. faveolata yields well-defined annual Sr/Ca cycles ranging in amplitude from ~0.3 and 0.5 mmol/mol. The amplitude of ~0.3 to 0.5 mmol/mol equates to a 10-15°C seasonal SST amplitude, which is consistent with available local instrumental records. Summer maxima proxy SSTs calculated from the modern coral Sr/ Ca tend to be fairly stable: most SST maxima from 1961–2008 are 29°C ± 1°C. In contrast, winter minimum SST calculated in the 47-year modern time-series are highly variable, with a cool interval in the early to mid-1970s. The Holocene (~6 Ka) Montastraea faveolata coral also yields distinct annual Sr/Ca cycles with amplitudes ranging from ~0.3 to 0.6 mmol/mol. Absolute Sr/Ca values and thus resulting SST estimates over the ~7-year long record are similar to those from the modern coral. We conclude that Sr/Ca from Montastraea faveolata has high potential for developing subannually resolved Holocene SST records.

  13. Development and Validation of a GC-MS Method for the Detection and Quantification of Clotiapine in Blood and Urine Specimens and Application to a Postmortem Case

    PubMed Central

    Mannocchi, Giulio; Pantano, Flaminia; Tittarelli, Roberta; Catanese, Miriam; Umani Ronchi, Federica; Busardò, Francesco Paolo

    2015-01-01

    Introduction. Clotiapine is an atypical antipsychotic of the dibenzothiazepine class introduced in a few European countries since 1970, efficient in treatment-resistant schizophrenic patients. There is little published data on the therapeutic and toxic concentrations of this drug. Aims. The aim of the present study is the development and validation of a method that allows the detection and quantification of clotiapine in blood and urine specimens by gas chromatography-mass spectrometry (GC-MS). Methods. Validation was performed working on spiked postmortem blood and urine samples. Samples were extracted with liquid-liquid extraction (LLE) technique at pH 8.5 with n-hexane/dichloromethane (85/15 v/v) and analysis was followed by GC-MS. Methadone-d9 was used as internal standard. Results. The limit of detection (LOD) was 1.2 and 1.3 ng/mL for urine and blood, respectively, while the lower limit of quantification (LLOQ) was 3.9 and 4.3 ng/mL, respectively. Linearity, precision, selectivity, accuracy, and recovery were also determined. The method was applied to a postmortem case. The blood and urine clotiapine concentrations were 1.32 and 0.49 μg/mL, respectively. Conclusions. A reliable GC-MS method for the detection and quantification of clotiapine in blood and urine samples has been developed and fully validated and then applied to a postmortem case. PMID:26236337

  14. A gas chromatography-thermal conductivity detection method for helium detection in postmortem blood and tissue specimens.

    PubMed

    Schaff, Jason E; Karas, Roman P; Marinetti, Laureen

    2012-03-01

    In cases of death by inert gas asphyxiation, it can be difficult to obtain toxicological evidence supporting assignment of a cause of death. Because of its low mass and high diffusivity, and its common use as a carrier gas, helium presents a particular challenge in this respect. We describe a rapid and simple gas chromatography-thermal conductivity detection method to qualitatively screen a variety of postmortem biological specimens for the presence of helium. Application of this method is demonstrated with three case examples, encompassing an array of different biological matrices. PMID:22337780

  15. Setup and validation of a convenient sampling procedure to promptly and effectively stabilize vitamin C in blood and plasma specimens stored at routine temperatures.

    PubMed

    Rossi, Barbara; Tittone, Francesca; Palleschi, Simonetta

    2016-07-01

    Vitamin C (ascorbic acid, AA) is very labile in nature and decays quickly after blood withdrawal. To ensure AA stability, the current procedure prescribes immediate plasma acidification followed by sample storage at ultra-low temperature. The aim of this study was to set up a pre-analytical procedure to promptly stabilize AA at routine temperatures while minimizing both specimen manipulation and instrumental requirement. Blood from healthy subjects was collected in lithium-heparin gel separator tubes containing or not different reducing agents (dithioerythritol, tris(2-carboxyethyl)phosphine, n-acetylcysteine and sodium thiosulfate). Plasma AA stability during blood and plasma storage at routine temperatures was evaluated. Plasma AA concentration was assayed by RP-HPLC-UV under ion suppression conditions. Each of the reductants tested was able to slow down the ex vivo degradation of plasma AA; dithioerythritol was the most effective. Five to 10 mmol/L dithioerythritol did not interfere with blood separation and allowed plasma AA to be stabilized up to 6 h, 24 h and 60 days at room temperature, +4 °C and -25 °C, respectively. The method worked well even in case of delayed blood separation and/or incomplete vacutainer filling. The procedure is feasible and reliable. Of special usefulness in clinical and epidemiological studies, prompt plasma manipulation after blood withdrawal or special storage equipments are not required. Graphical Abstract Collecting blood in tubes containing a reducing agent is a feasible method to promptly and effectively stabilize plasma vitamin C at routine temperature. PMID:27113458

  16. Use of Blood Smears and Dried Blood Spots for Polymerase Chain Reaction–Based Detection and Quantification of Bacterial Infection and Plasmodium falciparum in Severely Ill Febrile African Children

    PubMed Central

    Wihokhoen, Benchawan; Dondorp, Arjen M.; Turner, Paul; Woodrow, Charles J.; Imwong, Mallika

    2016-01-01

    Molecular approaches offer a means of testing archived samples stored as dried blood spots in settings where standard blood cultures are not possible. Peripheral blood films are one suggested source of material, although the sensitivity of this approach has not been well defined. Thin blood smears and dried blood spots from a severe pediatric malaria study were assessed using specific polymerase chain reaction (PCR) primers to detect non-typhoidal Salmonella (NTS; MisL gene), Streptococcus pneumoniae (lytA), and Plasmodium falciparum (18S rRNA). Of 16 cases of NTS and S. pneumoniae confirmed on blood culture, none were positive by PCR using DNA extracts from blood films or dried blood spots. In contrast, four of 36 dried blood spots and two of 178 plasma samples were PCR positive for S. pneumoniae, despite negative bacterial blood cultures, suggesting false positives. Quantitative assessment revealed that the effective concentration of P. falciparum DNA in blood films was three log orders of magnitude lower than for dried blood spots. The P. falciparum kelch13 gene could not be amplified from blood films. These findings question the value of blood PCR-based approaches for detection of NTS and S. pneumoniae, and show that stored blood films are an inefficient method of studying P. falciparum. PMID:26711525

  17. Use of Blood Smears and Dried Blood Spots for Polymerase Chain Reaction-Based Detection and Quantification of Bacterial Infection and Plasmodium falciparum in Severely Ill Febrile African Children.

    PubMed

    Wihokhoen, Benchawan; Dondorp, Arjen M; Turner, Paul; Woodrow, Charles J; Imwong, Mallika

    2016-02-01

    Molecular approaches offer a means of testing archived samples stored as dried blood spots in settings where standard blood cultures are not possible. Peripheral blood films are one suggested source of material, although the sensitivity of this approach has not been well defined. Thin blood smears and dried blood spots from a severe pediatric malaria study were assessed using specific polymerase chain reaction (PCR) primers to detect non-typhoidal Salmonella (NTS; MisL gene), Streptococcus pneumoniae (lytA), and Plasmodium falciparum (18S rRNA). Of 16 cases of NTS and S. pneumoniae confirmed on blood culture, none were positive by PCR using DNA extracts from blood films or dried blood spots. In contrast, four of 36 dried blood spots and two of 178 plasma samples were PCR positive for S. pneumoniae, despite negative bacterial blood cultures, suggesting false positives. Quantitative assessment revealed that the effective concentration of P. falciparum DNA in blood films was three log orders of magnitude lower than for dried blood spots. The P. falciparum kelch13 gene could not be amplified from blood films. These findings question the value of blood PCR-based approaches for detection of NTS and S. pneumoniae, and show that stored blood films are an inefficient method of studying P. falciparum. PMID:26711525

  18. Dried blood spots for monitoring and individualization of antiepileptic drug treatment.

    PubMed

    Milosheska, Daniela; Grabnar, Iztok; Vovk, Tomaž

    2015-07-30

    Therapeutic drug monitoring (TDM) is a multi-disciplinary clinical specialty used for optimization and individualization of drug therapy in the general and special populations. Since most antiepileptic drugs (AEDs) are characterized by pronounced intra- and inter-individual variability, it can be especially valuable as an aid for dosing adjustments in patients with epilepsy. Dried blood spots (DBS) sampling technique is recognized as a suitable alternative for conventional sampling methods as TDM interventions should be applied in the most cost-effective, rational and clinically useful manner. In the present review we summarize the latest trends and applications of DBS in TDM of epilepsy. Quantification of AEDs in DBS was employed in various clinical settings and has been already reported for phenobarbital, phenytoin, valproic acid, clonazepam, clobazam, carbamazepine, topiramate, rufinamide, lamotrigine, 10-hydroxycarbazepine and levetiracetam. The major limitation of the published studies are restricted evaluation of critical parameters such as the impact of spotted blood volume, spot homogeneity and haematocrit effect, limited clinical validation and non-established correlations between the DBS and plasma concentrations of AEDs. Standardization of critical technical aspects for appropriate sampling, sample preparation and validation of the analytical procedures for quantification of the drugs, as well as appropriate interpretation of the results are the fields which should get more attention in upcoming studies. Limited data on clinical validation and the fact that this technique has been used in practice only for a few AEDs makes the routine implementation of TDM of AEDs using DBS method a big challenge that should be faced by the pharmaceutical scientists in the future. PMID:25896371

  19. [Optimisation of retrospective diagnosis of cytomegalovirus congenital infection from dried blood spots].

    PubMed

    Vauloup-Fellous, C; Dubreuil, P; Grangeot-Keros, L

    2006-12-01

    Out of the 90% of cytomegalovirus (CMV) congenitally infected children that are asymptomatic at birth, 5 to 15% will later develop complications, mainly neurodevelopmental defects and/or deafness. Unfortunately, after the first 2 weeks of life, usual diagnostic techniques for CMV detection (viral culture and serology) are useless to differentiate congenital infection from post-natal acquired infection, whereas detection of viral DNA from dried blood spots (DBS; Guthrie cards), systematically collected from all newborns in the first days of life, has been described for late diagnosis of CMV congenital infection. The aim of our study was to choose and optimise a viral DNA extraction method from DBS and to study if CMV DNA detection is reliable when DBS are stored for 1 year at room temperature or 2 months at 37 degrees C. 10 reference cards (blood collected from CMV seronegative newborns (IgG/IgM negative) were "infected" with serial dilutions of virus and spotted on Guthrie cards) were tested. 3 extraction methods were evaluated, products of PCR were analyzes by agarose gel electrophoresis and quantification of CMV from DBS was also performed. Analysis of the results obtained from reference cards showed higher sensitivity of phenol/chloroform extraction following treatment with proteinase K, compared to heat extraction in cell culture medium or extraction with a commercial kit. We did not observe quantitative loss of viral DNA after 1 year storage at room temperature. CMV DNA detection from Guthrie cards could become a very useful tool for retrospective diagnosis of congenital CMV infection when sequelae are diagnosed in the first years of life. We are pursuing this study with DBS from congenitally infected children. PMID:17027198

  20. Efficient IDUA Gene Mutation Detection with Combined Use of dHPLC and Dried Blood Samples

    PubMed Central

    Duarte, Ana Joana; Vieira, Luis

    2013-01-01

    Objectives. Development of a simple mutation directed method in order to allow lowering the cost of mutation testing using an easily obtainable biological material. Assessment of the feasibility of such method was tested using a GC-rich amplicon. Design and Methods. A method of denaturing high-performance liquid chromatography (dHPLC) was improved and implemented as a technique for the detection of variants in exon 9 of the IDUA gene. The optimized method was tested in 500 genomic DNA samples obtained from dried blood spots (DBS). Results. With this dHPLC approach it was possible to detect different variants, including the common p.Trp402Ter mutation in the IDUA gene. The high GC content did not interfere with the resolution and reliability of this technique, and discrimination of G-C transversions was also achieved. Conclusion. This PCR-based dHPLC method is proved to be a rapid, a sensitive, and an excellent option for screening numerous samples obtained from DBS. Furthermore, it resulted in the consistent detection of clearly distinguishable profiles of the common p.Trp402Ter IDUA mutation with an advantageous balance of cost and technical requirements. PMID:27335677

  1. Free thyroxin measured in dried blood spots from normal, low-birth-weight, and hypothyroid neonates.

    PubMed

    Lemonnier, F; Masson, J; Laroche, D; Travert, J; Travert, G

    1991-12-01

    We have adapted a new radioimmunoassay for free thyroxin (FT4) measurement in dried blood spots for use in neonatal screening for hypothyroidism. The method is easy, fast, and cheap. Within-assay and between-assay CVs are respectively 9.6% and 13.2%. In 997 neonates three days postpartum with normal thyrotropin concentrations, the mean FT4 concentration was 27.2 pmol/L (SD 7.3 pmol/L). There was no significant difference in mean FT4 concentration between boys and girls. FT4 concentrations increased linearly with birth weight or with gestational age, as expressed by multiple linear regression: FT4 (pmol/L) = 0.0016 birth weight (g) + 0.6931 gestational age (weeks) - 4.8772. Only gestational age significantly affected the FT4 value. For five hypothyroid infants tested on day three postpartum, FT4 values were all below the 1st percentile of values from healthy neonates. Thus, when the neonatal concentration of thyrotropin is above normal, FT4 measured in the same sample can provide a reliable earlier diagnosis of hypothyroidism. PMID:1764786

  2. Geneva cocktail for cytochrome p450 and P-glycoprotein activity assessment using dried blood spots.

    PubMed

    Bosilkovska, M; Samer, C F; Déglon, J; Rebsamen, M; Staub, C; Dayer, P; Walder, B; Desmeules, J A; Daali, Y

    2014-09-01

    The suitability of the capillary dried blood spot (DBS) sampling method was assessed for simultaneous phenotyping of cytochrome P450 (CYP) enzymes and P-glycoprotein (P-gp) using a cocktail approach. Ten volunteers received an oral cocktail capsule containing low doses of the probes bupropion (CYP2B6), flurbiprofen (CYP2C9), omeprazole (CYP2C19), dextromethorphan (CYP2D6), midazolam (CYP3A), and fexofenadine (P-gp) with coffee/Coke (CYP1A2) on four occasions. They received the cocktail alone (session 1), and with the CYP inhibitors fluvoxamine and voriconazole (session 2) and quinidine (session 3). In session 4, subjects received the cocktail after a 7-day pretreatment with the inducer rifampicin. The concentrations of probes/metabolites were determined in DBS and plasma using a single liquid chromatography-tandem mass spectrometry method. The pharmacokinetic profiles of the drugs were comparable in DBS and plasma. Important modulation of CYP and P-gp activities was observed in the presence of inhibitors and the inducer. Minimally invasive one- and three-point (at 2, 3, and 6 h) DBS-sampling methods were found to reliably reflect CYP and P-gp activities at each session. PMID:24722393

  3. Direct Blood Dry LAMP: A Rapid, Stable, and Easy Diagnostic Tool for Human African Trypanosomiasis

    PubMed Central

    Hayashida, Kyoko; Kajino, Kiichi; Hachaambwa, Lottie; Namangala, Boniface; Sugimoto, Chihiro

    2015-01-01

    Loop-mediated isothermal amplification (LAMP) is a rapid and sensitive tool used for the diagnosis of a variety of infectious diseases. One of the advantages of this method over the polymerase chain reaction is that DNA amplification occurs at a constant temperature, usually between 60–65°C; therefore, expensive devices are unnecessary for this step. However, LAMP still requires complicated sample preparation steps and a well-equipped laboratory to produce reliable and reproducible results, which limits its use in resource-poor laboratories in most developing countries. In this study, we made several substantial modifications to the technique to carry out on-site diagnosis of Human African Trypanosomiasis (HAT) in remote areas using LAMP. The first essential improvement was that LAMP reagents were dried and stabilized in a single tube by incorporating trehalose as a cryoprotectant to prolong shelf life at ambient temperature. The second technical improvement was achieved by simplifying the sample preparation step so that DNA or RNA could be amplified directly from detergent-lysed blood samples. With these modifications, diagnosis of HAT in local clinics or villages in endemic areas becomes a reality, which could greatly impact on the application of diagnosis not only for HAT but also for other tropical diseases. PMID:25769046

  4. High-Quality Exome Sequencing of Whole-Genome Amplified Neonatal Dried Blood Spot DNA

    PubMed Central

    Grove, Jakob; Bækvad-Hansen, Marie; Christiansen, Michael; Hagen, Christian Munch; Maller, Julian; Stevens, Christine; Li, Shenting; Li, Qibin; Sun, Jihua; Wang, Jun; Nordentoft, Merete; Werge, Thomas Mears; Mortensen, Preben Bo; Børglum, Anders Dupont; Daly, Mark; Hougaard, David Michael

    2016-01-01

    Stored neonatal dried blood spot (DBS) samples from neonatal screening programmes are a valuable diagnostic and research resource. Combined with information from national health registries they can be used in population-based studies of genetic diseases. DNA extracted from neonatal DBSs can be amplified to obtain micrograms of an otherwise limited resource, referred to as whole-genome amplified DNA (wgaDNA). Here we investigate the robustness of exome sequencing of wgaDNA of neonatal DBS samples. We conducted three pilot studies of seven, eight and seven subjects, respectively. For each subject we analysed a neonatal DBS sample and corresponding adult whole-blood (WB) reference sample. Different DNA sample types were prepared for each of the subjects. Pilot 1: wgaDNA of 2x3.2mm neonatal DBSs (DBS_2x3.2) and raw DNA extract of the WB reference sample (WB_ref). Pilot 2: DBS_2x3.2, WB_ref and a WB_ref replica sharing DNA extract with the WB_ref sample. Pilot 3: DBS_2x3.2, WB_ref, wgaDNA of 2x1.6 mm neonatal DBSs and wgaDNA of the WB reference sample. Following sequencing and data analysis, we compared pairwise variant calls to obtain a measure of similarity—the concordance rate. Concordance rates were slightly lower when comparing DBS vs WB sample types than for any two WB sample types of the same subject before filtering of the variant calls. The overall concordance rates were dependent on the variant type, with SNPs performing best. Post-filtering, the comparisons of DBS vs WB and WB vs WB sample types yielded similar concordance rates, with values close to 100%. WgaDNA of neonatal DBS samples performs with great accuracy and efficiency in exome sequencing. The wgaDNA performed similarly to matched high-quality reference—whole-blood DNA—based on concordance rates calculated from variant calls. No differences were observed substituting 2x3.2 with 2x1.6 mm discs, allowing for additional reduction of sample material in future projects. PMID:27089011

  5. High-Quality Exome Sequencing of Whole-Genome Amplified Neonatal Dried Blood Spot DNA.

    PubMed

    Poulsen, Jesper Buchhave; Lescai, Francesco; Grove, Jakob; Bækvad-Hansen, Marie; Christiansen, Michael; Hagen, Christian Munch; Maller, Julian; Stevens, Christine; Li, Shenting; Li, Qibin; Sun, Jihua; Wang, Jun; Nordentoft, Merete; Werge, Thomas Mears; Mortensen, Preben Bo; Børglum, Anders Dupont; Daly, Mark; Hougaard, David Michael; Bybjerg-Grauholm, Jonas; Hollegaard, Mads Vilhelm

    2016-01-01

    Stored neonatal dried blood spot (DBS) samples from neonatal screening programmes are a valuable diagnostic and research resource. Combined with information from national health registries they can be used in population-based studies of genetic diseases. DNA extracted from neonatal DBSs can be amplified to obtain micrograms of an otherwise limited resource, referred to as whole-genome amplified DNA (wgaDNA). Here we investigate the robustness of exome sequencing of wgaDNA of neonatal DBS samples. We conducted three pilot studies of seven, eight and seven subjects, respectively. For each subject we analysed a neonatal DBS sample and corresponding adult whole-blood (WB) reference sample. Different DNA sample types were prepared for each of the subjects. Pilot 1: wgaDNA of 2x3.2mm neonatal DBSs (DBS_2x3.2) and raw DNA extract of the WB reference sample (WB_ref). Pilot 2: DBS_2x3.2, WB_ref and a WB_ref replica sharing DNA extract with the WB_ref sample. Pilot 3: DBS_2x3.2, WB_ref, wgaDNA of 2x1.6 mm neonatal DBSs and wgaDNA of the WB reference sample. Following sequencing and data analysis, we compared pairwise variant calls to obtain a measure of similarity--the concordance rate. Concordance rates were slightly lower when comparing DBS vs WB sample types than for any two WB sample types of the same subject before filtering of the variant calls. The overall concordance rates were dependent on the variant type, with SNPs performing best. Post-filtering, the comparisons of DBS vs WB and WB vs WB sample types yielded similar concordance rates, with values close to 100%. WgaDNA of neonatal DBS samples performs with great accuracy and efficiency in exome sequencing. The wgaDNA performed similarly to matched high-quality reference--whole-blood DNA--based on concordance rates calculated from variant calls. No differences were observed substituting 2x3.2 with 2x1.6 mm discs, allowing for additional reduction of sample material in future projects. PMID:27089011

  6. Quantification of rifapentine, a potent antituberculosis drug, from dried blood spot samples using liquid chromatographic-tandem mass spectrometric analysis.

    PubMed

    Parsons, Teresa L; Marzinke, Mark A; Hoang, Thuy; Bliven-Sizemore, Erin; Weiner, Marc; Mac Kenzie, William R; Dorman, Susan E; Dooley, Kelly E

    2014-11-01

    The quantification of antituberculosis drug concentrations in multinational trials currently requires the collection of modest blood volumes, centrifugation, aliquoting of plasma, freezing, and keeping samples frozen during shipping. We prospectively enrolled healthy individuals into the Tuberculosis Trials Consortium Study 29B, a phase I dose escalation study of rifapentine, a rifamycin under evaluation in tuberculosis treatment trials. We developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for quantifying rifapentine in whole blood on dried blood spots (DBS) to facilitate pharmacokinetic/pharmacodynamic analyses in clinical trials. Paired plasma and whole-blood samples were collected by venipuncture, and whole blood was spotted on Whatman protein saver 903 cards. The methods were optimized for plasma and then validated for DBS. The analytical measuring range for quantification of rifapentine and its metabolite was 50 to 80,000 ng/ml in whole-blood DBS. The analyte was stable on the cards for 11 weeks with a desiccant at room temperature and protected from light. The method concordance for paired plasma and whole-blood DBS samples was determined after correcting for participant hematocrit or population-based estimates of bias from Bland-Altman plots. The application of either correction factor resulted in acceptable correlation between plasma and whole-blood DBS (Passing-Bablok regression corrected for hematocrit; y = 0.98x + 356). Concentrations of rifapentine may be determined from whole-blood DBS collected via venipuncture after normalization in order to account for the dilutional effects of red blood cells. Additional studies are focused on the application of this methodology to capillary blood collected by finger stick. The simplicity of processing, storage, shipping, and low blood volume makes whole-blood DBS attractive for rifapentine pharmacokinetic evaluations, especially in international and pediatric trials. PMID:25182637

  7. Quantification of Rifapentine, a Potent Antituberculosis Drug, from Dried Blood Spot Samples Using Liquid Chromatographic-Tandem Mass Spectrometric Analysis

    PubMed Central

    Parsons, Teresa L.; Marzinke, Mark A.; Hoang, Thuy; Bliven-Sizemore, Erin; Weiner, Marc; Mac Kenzie, William R.; Dorman, Susan E.

    2014-01-01

    The quantification of antituberculosis drug concentrations in multinational trials currently requires the collection of modest blood volumes, centrifugation, aliquoting of plasma, freezing, and keeping samples frozen during shipping. We prospectively enrolled healthy individuals into the Tuberculosis Trials Consortium Study 29B, a phase I dose escalation study of rifapentine, a rifamycin under evaluation in tuberculosis treatment trials. We developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for quantifying rifapentine in whole blood on dried blood spots (DBS) to facilitate pharmacokinetic/pharmacodynamic analyses in clinical trials. Paired plasma and whole-blood samples were collected by venipuncture, and whole blood was spotted on Whatman protein saver 903 cards. The methods were optimized for plasma and then validated for DBS. The analytical measuring range for quantification of rifapentine and its metabolite was 50 to 80,000 ng/ml in whole-blood DBS. The analyte was stable on the cards for 11 weeks with a desiccant at room temperature and protected from light. The method concordance for paired plasma and whole-blood DBS samples was determined after correcting for participant hematocrit or population-based estimates of bias from Bland-Altman plots. The application of either correction factor resulted in acceptable correlation between plasma and whole-blood DBS (Passing-Bablok regression corrected for hematocrit; y = 0.98x + 356). Concentrations of rifapentine may be determined from whole-blood DBS collected via venipuncture after normalization in order to account for the dilutional effects of red blood cells. Additional studies are focused on the application of this methodology to capillary blood collected by finger stick. The simplicity of processing, storage, shipping, and low blood volume makes whole-blood DBS attractive for rifapentine pharmacokinetic evaluations, especially in international and pediatric trials. PMID:25182637

  8. Supplementation of Dried Mealworm (Tenebrio molitor larva) on Growth Performance, Nutrient Digestibility and Blood Profiles in Weaning Pigs.

    PubMed

    Jin, X H; Heo, P S; Hong, J S; Kim, N J; Kim, Y Y

    2016-07-01

    This experiment was conducted to investigate the effects of dried mealworm (Tenebrio molitor larva) on growth performance, nutrient digestibility and blood profiles in weaning pigs. A total of 120 weaning pigs (28±3 days and 8.04±0.08 kg of body weight) were allotted to one of five treatments, based on sex and body weight, in 6 replicates with 4 pigs per pen by a randomized complete block design. Supplementation level of dried mealworm was 0%, 1.5%, 3.0%, 4.5%, or 6.0% in experimental diet as treatment. Two phase feeding programs (phase I from 0 day to 14 day, phase II from 14 day to 35 day) were used in this experiment. All animals were allowed to access diet and water ad libitum. During phase I, increasing level of dried mealworm in diet linearly improved the body weight (p<0.01), average daily gain (ADG) (p<0.01) and average daily feed intake (ADFI) (p<0.01). During phase II, ADG also tended to increase linearly when pigs were fed higher level of dried mealworm (p = 0.08). In addition, increasing level of dried mealworm improved the ADG (p<0.01), ADFI (p<0.05) and tended to increase gain to feed ratio (p = 0.07) during the whole experimental period. As dried mealworm level was increased, nitrogen retention and digestibility of dry matter as well as crude protein were linearly increased (p = 0.05). In the results of blood profiles, decrease of blood urea nitrogen (linear, p = 0.05) and increase of insulin-like growth factor (linear, p = 0.03) were observed as dried mealworm was increased in diet during phase II. However, there were no significant differences in immunoglobulin A (IgA) and IgG concentration by addition of dried mealworm in the growth trial. Consequently, supplementation of dried mealworm up to 6% in weaning pigs' diet improves growth performance and nutrient digestibility without any detrimental effect on immune responses. PMID:27282974

  9. Supplementation of Dried Mealworm (Tenebrio molitor larva) on Growth Performance, Nutrient Digestibility and Blood Profiles in Weaning Pigs

    PubMed Central

    Jin, X. H.; Heo, P. S.; Hong, J. S.; Kim, N. J.; Kim, Y. Y.

    2016-01-01

    This experiment was conducted to investigate the effects of dried mealworm (Tenebrio molitor larva) on growth performance, nutrient digestibility and blood profiles in weaning pigs. A total of 120 weaning pigs (28±3 days and 8.04±0.08 kg of body weight) were allotted to one of five treatments, based on sex and body weight, in 6 replicates with 4 pigs per pen by a randomized complete block design. Supplementation level of dried mealworm was 0%, 1.5%, 3.0%, 4.5%, or 6.0% in experimental diet as treatment. Two phase feeding programs (phase I from 0 day to 14 day, phase II from 14 day to 35 day) were used in this experiment. All animals were allowed to access diet and water ad libitum. During phase I, increasing level of dried mealworm in diet linearly improved the body weight (p<0.01), average daily gain (ADG) (p<0.01) and average daily feed intake (ADFI) (p<0.01). During phase II, ADG also tended to increase linearly when pigs were fed higher level of dried mealworm (p = 0.08). In addition, increasing level of dried mealworm improved the ADG (p<0.01), ADFI (p<0.05) and tended to increase gain to feed ratio (p = 0.07) during the whole experimental period. As dried mealworm level was increased, nitrogen retention and digestibility of dry matter as well as crude protein were linearly increased (p = 0.05). In the results of blood profiles, decrease of blood urea nitrogen (linear, p = 0.05) and increase of insulin-like growth factor (linear, p = 0.03) were observed as dried mealworm was increased in diet during phase II. However, there were no significant differences in immunoglobulin A (IgA) and IgG concentration by addition of dried mealworm in the growth trial. Consequently, supplementation of dried mealworm up to 6% in weaning pigs’ diet improves growth performance and nutrient digestibility without any detrimental effect on immune responses. PMID:27282974

  10. Application to cows and horses of Spotchem, a dry-chemistry blood analyzer for use in veterinary clinics.

    PubMed

    Hoshi, F; Satho, M; Koyama, S; Nakadaka, K; Chiba, M; Ikeda, N; Hakamada, R; Higuchi, S; Kawamura, S

    1994-02-01

    The usefulness of a dry-chemistry blood analyzer, Spotchem SP-4410 (SP-4410) in a veterinary clinic for analysis of bovine and equine blood chemistry was studied. We quantitated total protein (TP), albumin (Alb), total bilirubin (T-Bil), blood urea nitrogen (BUN), total cholesterol (T-Cho), glucose (Glu), calcium (Ca), aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT), creatinine phosphokinase (CPK), and alkaline phosphatase (ALP) in bovine sera. Each sample was assayed with both the SP-4410 and an automated blood analyzer which served as a wet-chemistry reference system, and the data were analyzed with regression analysis. The correlation coefficient for AST was 0.997 being the highest for all the parameters, and all the correlation coefficients were 0.93 or higher. The coefficients of variation were lower than 5.0 except in the case of bovine T-Bil where it was 5,756. The ranges of normal reference values measured by SP-4410 were the same as those reported by other investigators in most cases, but those for GGT and CPK were slightly higher. The strongest interference was observed with hemoglobin. It seems that dry-chemical-analysis of blood serum using the SP-4410 is useful for analysis of bovine and equine blood. PMID:8085395

  11. Evaluation of the Broad-Range PCR/ESI-MS Technology in Blood Specimens for the Molecular Diagnosis of Bloodstream Infections

    PubMed Central

    Jordana-Lluch, Elena; Giménez, Montserrat; Quesada, Mª Dolores; Rivaya, Belén; Marcó, Clara; Domínguez, Mª Jesús; Arméstar, Fernando; Martró, Elisa; Ausina, Vicente

    2015-01-01

    Background Rapid identification of the etiological agent in bloodstream infections is of vital importance for the early administration of the most appropriate antibiotic therapy. Molecular methods may offer an advantage to current culture-based microbiological diagnosis. The goal of this study was to evaluate the performance of IRIDICA, a platform based on universal genetic amplification followed by mass spectrometry (PCR/ESI-MS) for the molecular diagnosis of sepsis-related pathogens directly from the patient’s blood. Methods A total of 410 whole blood specimens from patients admitted to Emergency Room (ER) and Intensive Care Unit (ICU) with clinical suspicion of sepsis were tested with the IRIDICA BAC BSI Assay (broad identification of bacteria and Candida spp.). Microorganisms grown in culture and detected by IRIDICA were compared considering blood culture as gold standard. When discrepancies were found, clinical records and results from other cultures were taken into consideration (clinical infection criterion). Results The overall positive and negative agreement of IRIDICA with blood culture in the analysis by specimen was 74.8% and 78.6%, respectively, rising to 76.9% and 87.2% respectively, when compared with the clinical infection criterion. Interestingly, IRIDICA detected 41 clinically significant microorganisms missed by culture, most of them from patients under antimicrobial treatment. Of special interest were the detections of one Mycoplasma hominis and two Mycobacterium simiae in immunocompromised patients. When ICU patients were analyzed separately, sensitivity, specificity, positive and negative predictive values compared with blood culture were 83.3%, 78.6%, 33.9% and 97.3% respectively, and 90.5%, 87.2%, 64.4% and 97.3% respectively, in comparison with the clinical infection criterion. Conclusions IRIDICA is a promising technology that offers an early and reliable identification of a wide variety of pathogens directly from the patient’s blood

  12. High quality methylome-wide investigations through next-generation sequencing of DNA from a single archived dry blood spot.

    PubMed

    Aberg, Karolina A; Xie, Lin Y; Nerella, Srilaxmi; Copeland, William E; Costello, E Jane; van den Oord, Edwin J C G

    2013-05-01

    The potential importance of DNA methylation in the etiology of complex diseases has led to interest in the development of methylome-wide association studies (MWAS) aimed at interrogating all methylation sites in the human genome. When using blood as biomaterial for a MWAS the DNA is typically extracted directly from fresh or frozen whole blood that was collected via venous puncture. However, DNA extracted from dry blood spots may also be an alternative starting material. In the present study, we apply a methyl-CpG binding domain (MBD) protein enrichment-based technique in combination with next generation sequencing (MBD-seq) to assess the methylation status of the ~27 million CpGs in the human autosomal reference genome. We investigate eight methylomes using DNA from blood spots. This data are compared with 1,500 methylomes previously assayed with the same MBD-seq approach using DNA from whole blood. When investigating the sequence quality and the enrichment profile across biological features, we find that DNA extracted from blood spots gives comparable results with DNA extracted from whole blood. Only if the amount of starting material is ≤ 0.5µg DNA we observe a slight decrease in the assay performance. In conclusion, we show that high quality methylome-wide investigations using MBD-seq can be conducted in DNA extracted from archived dry blood spots without sacrificing quality and without bias in enrichment profile as long as the amount of starting material is sufficient. In general, the amount of DNA extracted from a single blood spot is sufficient for methylome-wide investigations with the MBD-seq approach. PMID:23644822

  13. A Pilot Study Using Residual Newborn Dried Blood Spots to Assess the Potential Role of Cytomegalovirus and Toxoplasma gondii in the Etiology of Congenital Hydrocephalus

    PubMed Central

    Simeone, Regina M.; Rasmussen, Sonja A.; Mei, Joanne V.; Dollard, Sheila C.; Frias, Jaime L.; Shaw, Gary M.; Canfield, Mark A.; Meyer, Robert E.; Jones, Jeffrey L.; Lorey, Fred; Honein, Margaret A.

    2015-01-01

    BACKGROUND Congenital hydrocephalus is a condition characterized by accumulation of cerebrospinal fluid in the ventricles of the brain. Prenatal infections are risk factors for some birth defects. This pilot study investigated whether residual dried blood spots (DBS) could be used to assess infections as risk factors for birth defects by examining the associations between prenatal infection with Toxoplasma gondii (T. gondii) or cytomegalovirus (CMV) with congenital hydrocephalus. METHODS Case-infants with hydrocephalus (N = 410) were identified among live-born infants using birth defects surveillance systems in California, North Carolina, and Texas. Control-infants without birth defects were randomly selected from the same geographic areas and time periods as case-infants (N = 448). We tested residual DBS from case- and control-infants for T. gondii immunoglobulin M and CMV DNA. When possible, we calculated crude odds ratios (cORs) and confidence intervals (CIs). RESULTS Evidence for prenatal T. gondii infection was more common among case-infants (1.2%) than control-infants (0%; p = 0.11), and evidence for prenatal CMV infection was higher among case-infants (1.5%) than control-infants (0.7%; cOR: 2.3; 95% CI: 0.48, 13.99). CONCLUSIONS Prenatal infections with T. gondii and CMV occurred more often among infants with congenital hydrocephalus than control-infants, although differences were not statistically significant. This pilot study highlighted some challenges in using DBS to examine associations between certain infections and birth defects, particularly related to reduced sensitivity and specimen storage conditions. Further study with increased numbers of specimens and higher quality specimens should be considered to understand better the contribution of these infections to the occurrence of congenital hydrocephalus. PMID:23716471

  14. The Relevance of a Novel Quantitative Assay to Detect up to 40 Major Streptococcus pneumoniae Serotypes Directly in Clinical Nasopharyngeal and Blood Specimens

    PubMed Central

    Albrich, Werner C.; van der Linden, Mark P. G.; Bénet, Thomas; Chou, Monidarin; Sylla, Mariam; Barreto Costa, Patricia; Richard, Nathalie; Klugman, Keith P.; Endtz, Hubert P.; Paranhos-Baccalà, Gláucia; Telles, Jean-Noël

    2016-01-01

    For epidemiological and surveillance purposes, it is relevant to monitor the distribution and dynamics of Streptococcus pneumoniae serotypes. Conventional serotyping methods do not provide rapid or quantitative information on serotype loads. Quantitative serotyping may enable prediction of the invasiveness of a specific serotype compared to other serotypes carried. Here, we describe a novel, rapid multiplex real-time PCR assay for identification and quantification of the 40 most prevalent pneumococcal serotypes and the assay impacts in pneumonia specimens from emerging and developing countries. Eleven multiplex PCR to detect 40 serotypes or serogroups were optimized. Quantification was enabled by reference to standard dilutions of known bacterial load. Performance of the assay was evaluated to specifically type and quantify S. pneumoniae in nasopharyngeal and blood samples from adult and pediatric patients hospitalized with pneumonia (n = 664) from five different countries. Serogroup 6 was widely represented in nasopharyngeal specimens from all five cohorts. The most frequent serotypes in the French, South African, and Brazilian cohorts were 1 and 7A/F, 3 and 19F, and 14, respectively. When both samples were available, the serotype in blood was always present as carriage with other serotypes in the nasopharynx. Moreover, the ability of a serotype to invade the bloodstream may be linked to its nasopharyngeal load. The mean nasopharyngeal concentration of the serotypes that moved to the blood was 3 log-fold higher than the ones only found in the nasopharynx. This novel, rapid, quantitative assay may potentially predict some of the S. pneumoniae serotypes invasiveness and assessment of pneumococcal serotype distribution. PMID:26986831

  15. The Relevance of a Novel Quantitative Assay to Detect up to 40 Major Streptococcus pneumoniae Serotypes Directly in Clinical Nasopharyngeal and Blood Specimens.

    PubMed

    Messaoudi, Melina; Milenkov, Milen; Albrich, Werner C; van der Linden, Mark P G; Bénet, Thomas; Chou, Monidarin; Sylla, Mariam; Barreto Costa, Patricia; Richard, Nathalie; Klugman, Keith P; Endtz, Hubert P; Paranhos-Baccalà, Gláucia; Telles, Jean-Noël

    2016-01-01

    For epidemiological and surveillance purposes, it is relevant to monitor the distribution and dynamics of Streptococcus pneumoniae serotypes. Conventional serotyping methods do not provide rapid or quantitative information on serotype loads. Quantitative serotyping may enable prediction of the invasiveness of a specific serotype compared to other serotypes carried. Here, we describe a novel, rapid multiplex real-time PCR assay for identification and quantification of the 40 most prevalent pneumococcal serotypes and the assay impacts in pneumonia specimens from emerging and developing countries. Eleven multiplex PCR to detect 40 serotypes or serogroups were optimized. Quantification was enabled by reference to standard dilutions of known bacterial load. Performance of the assay was evaluated to specifically type and quantify S. pneumoniae in nasopharyngeal and blood samples from adult and pediatric patients hospitalized with pneumonia (n = 664) from five different countries. Serogroup 6 was widely represented in nasopharyngeal specimens from all five cohorts. The most frequent serotypes in the French, South African, and Brazilian cohorts were 1 and 7A/F, 3 and 19F, and 14, respectively. When both samples were available, the serotype in blood was always present as carriage with other serotypes in the nasopharynx. Moreover, the ability of a serotype to invade the bloodstream may be linked to its nasopharyngeal load. The mean nasopharyngeal concentration of the serotypes that moved to the blood was 3 log-fold higher than the ones only found in the nasopharynx. This novel, rapid, quantitative assay may potentially predict some of the S. pneumoniae serotypes invasiveness and assessment of pneumococcal serotype distribution. PMID:26986831

  16. Four to seven random casual urine specimens are sufficient to estimate 24-h urinary sodium/potassium ratio in individuals with high blood pressure.

    PubMed

    Iwahori, T; Ueshima, H; Torii, S; Saito, Y; Fujiyoshi, A; Ohkubo, T; Miura, K

    2016-05-01

    This study was done to clarify the optimal number and type of casual urine specimens required to estimate urinary sodium/potassium (Na/K) ratio in individuals with high blood pressure. A total of 74 individuals with high blood pressure, 43 treated and 31 untreated, were recruited from the Japanese general population. Urinary sodium, potassium and Na/K ratio were measured in both casual urine samples and 7-day 24-h urine samples and then analyzed by correlation and Bland-Altman analyses. Mean Na/K ratio from random casual urine samples on four or more days strongly correlated with the Na/K ratio of 7-day 24-h urine (r=0.80-0.87), which was similar to the correlation between 1 and 2-day 24-h urine and 7-day 24-h urine (r=0.75-0.89). The agreement quality for Na/K ratio of seven random casual urine for estimating the Na/K ratio of 7-day 24-h urine was good (bias: -0.26, limits of agreements: -1.53-1.01), and it was similar to that of 2-day 24-h urine for estimating 7-day 24-h values (bias: 0.07, limits of agreement: -1.03 to 1.18). Stratified analyses comparing individuals using antihypertensive medication and individuals not using antihypertensive medication showed similar results. Correlations of the means of casual urine sodium or potassium concentrations with 7-day 24-h sodium or potassium excretions were relatively weaker than those for Na/K ratio. The mean Na/K ratio of 4-7 random casual urine specimens on different days provides a good substitute for 1-2-day 24-h urinary Na/K ratio for individuals with high blood pressure. PMID:26310187

  17. A microsphere-based assay for mutation analysis of the biotinidase gene using dried blood spots

    PubMed Central

    Lindau-Shepard, Barbara; Janik, David K.; Pass, Kenneth A.

    2012-01-01

    Biotinidase deficiency is an autosomal recessive syndrome caused by defects in the biotinidase gene, the product of which affects biotin metabolism. Newborn screening (NBS) for biotinidase deficiency can identify affected infants prior to onset of symptoms; biotin supplementation can resolve or prevent the clinical features. In NBS, dry blood spots (DBS) are usually tested for biotinidase enzyme activity by colorimetric analysis. By taking advantage of the multiplexing capabilities of the Luminex platform, we have developed a microsphere-based array genotyping method for the simultaneous detection of six disease causing mutations in the biotinidase gene, thereby permitting a second tier of molecular analysis. Genomic DNA was extracted from 3.2 mm DBS. Biotinidase gene sequences, containing the mutations of interest, were amplified by multiplexed polymerase chain reaction, followed by multiplexed allele-specific primer extension using universally tagged genotyping primers. The products were then hybridized to anti-tag carrying xTAG microspheres and detected on the Luminex platform. Genotypes were verified by sequencing. Genotyping results of 22 known biotinidase deficient samples by our xTAG biotinidase assay was in concordance with the results obtained from DNA sequencing, for all 6 mutations used in our panel. These results indicate that genotyping by an xTAG microsphere-based array is accurate, flexible, and can be adapted for high-throughput. Since NBS for biotinidase deficiency is by enzymatic assay, less than optimal quality of the DBS itself can compromise enzyme activity, while the DNA from these samples mostly remains unaffected. This assay warrants evaluation as a viable complement to the biotinidase semi-quantitative colorimetric assay.

  18. Analysis of methylcitrate in dried blood spots by liquid chromatography-tandem mass spectrometry.

    PubMed

    Al-Dirbashi, Osama Y; McIntosh, Nathan; McRoberts, Christine; Fisher, Larry; Rashed, Mohamed S; Makhseed, Nawal; Geraghty, Michael T; Santa, Tomofumi; Chakraborty, Pranesh

    2014-01-01

    Accumulation of propionylcarnitine (C3) in neonatal dried blood spots (DBS) is indicative of inborn errors of propionate metabolism including propionic acidemia (PA), methylmalonic aciduria (MMA), and cobalamin (Cbl) metabolic defects. Concentrations of C3 in affected newborns overlap with healthy individuals rendering this marker neither specific nor sensitive. While a conservative C3 cutoff together with relevant acylcarnitines ratios improve screening sensitivity, existing mass spectrometric methods in newborn screening laboratories are inadequate at improving testing specificity. Therefore, using the original screening DBS, we sought to measure 2-methylcitric acid (MCA), a pathognomonic hallmark of C3 disorders to decrease the false positive rate and improve the positive predictive value of C3 disorders. MCA was derivatized with 4-[2-(N,N-dimethylamino)ethylaminosulfonyl]-7-(2-aminoethylamino)-2,1,3-benzoxadiazole (DAABD-AE). No separate extraction step was required and derivatization was performed directly using a 3.2-mm disc of DBS as a sample (65°C for 45 min). The reaction mixture was analyzed by liquid chromatography tandem mass spectrometry. MCA was well separated and eluted at 2.3 min with a total run time of 7 min. The median and (range) of MCA of 0.06 μmol/L (0-0.63) were in excellent agreement with the literature. The method was applied retrospectively on DBS samples from established patients with PA, MMA, Cbl C, Cbl F, maternal vitamin B12 deficiency (n = 20) and controls (n = 337). Comparison with results obtained by another method was satisfactory (n = 252). This method will be applied as a second tier test for samples which trigger positive PA or MMA results by the primary newborn screening method. PMID:24997714

  19. Galactose-1-phosphate Uridyltransferase Dried Blood Spot Quality Control Materials for Newborn Screening Tests

    PubMed Central

    Adam, Barbara W.; Flores, Sharon R.; Hou, Yu; Allen, Todd W.; De Jesus, Victor R.

    2015-01-01

    Objectives We aimed to prepare dried-blood-spot (DBS) quality control (QC) materials for galactose-1-phosphate uridyltransferase (GALT), to evaluate their stability during storage and use, and to evaluate their performance in five DBS GALT test methods. Design and Methods We prepared and characterized GALT-normal and GALT-deficient DBS materials and compared GALT activities in DBSs after predetermined storage intervals at controlled temperatures and humidities. External evaluators documented the suitability of the DBS QC materials for use in five GALT test methods. Results GALT activity losses from DBSs stored in low (<30%) humidity for 14 days at 45°C, 35 days at 37°C, 91 days at room temperature, 182 days at 4°C, and 367 days at −20°C were 54%, 53%, 52% 23%, and 7% respectively. In paired DBSs stored in high humidity (>50%) for identical intervals, losses were: 45°C—68%; 37°C—79%; room temperature—72%, and 4°C—63%. GALT activities in DBSs stored at 4°C were stable throughout 19 excursions to room temperature. Twenty-five of 26 external evaluators, using five different GALT test methods, classified the GALT-deficient DBSs as “outside normal limits”. All evaluators classified the GALT-normal DBSs as “within normal limits”. Conclusions Most of the GALT activity loss from DBSs stored at elevated or room temperature was attributable to the effects of storage temperature. Most of the loss from DBSs stored at 4°C was attributable to the effects of elevated humidity. Loss from DBSs stored at −20°C was insignificant. The DBS materials were suitable for monitoring performance of all five GALT test methods. PMID:25528144

  20. Hepatitis B viral load in dried blood spots: a validation study in Zambia

    PubMed Central

    Vinikoor, Michael J.; Zürcher, Samuel; Musukuma, Kalo; Kachuwaire, Obert; Rauch, Andri; Chi, Benjamin H.; Gorgievski, Meri; Zwahlen, Marcel; Wandeler, Gilles

    2016-01-01

    Background Access to hepatitis B viral load (VL) testing is poor in sub-Saharan Africa (SSA) due to economic and logistical reasons. Objectives To demonstrate the feasibility of testing dried blood spots (DBS) for hepatitis B virus (HBV) VL in a laboratory in Lusaka, Zambia, and to compare HBV VLs between DBS and plasma samples. Study design Paired plasma and DBS samples from HIV-HBV co-infected Zambian adults were analyzed for HBV VL using the COBAS AmpliPrep/COBAS TaqMan HBV test (Version 2.0) and for genotype by direct sequencing. We used Bland-Altman analysis to compare VLs between sample types and by HBV genotype. Logistic regression analysis was conducted to assess the probability of an undetectable DBS result by plasma VL. Results Among 68 participants, median age was 34 years, 61.8% were men, and median plasma HBV VL was 3.98 log IU/ml (interquartile range, 2.04–5.95). Among sequenced viruses, 28 were genotype A1 and 27 were genotype E. Bland-Altman plots suggested strong agreement between DBS and plasma VLs. DBS VLs were on average 1.59 log IU/ml lower compared to plasma with 95% limits of agreement of −2.40 to −0.83 log IU/ml. At a plasma VL ≥2,000 IU/ml, the probability of an undetectable DBS result was 1.8% (95% CI: 0.5–6.6). At plasma VL ≥20,000 IU/ml this probability reduced to 0.2% (95% CI: 0.03–1.7). Conclusions In a Zambian laboratory, we observed strong agreement between DBS and plasma VLs and high sensitivity in DBS at plasma VL ≥2,000 IU/ml. As HBV treatment expands, DBS could increase access to HBV VL testing in SSA settings. PMID:26356987

  1. African Swine Fever Diagnosis Adapted to Tropical Conditions by the Use of Dried-blood Filter Papers.

    PubMed

    Randriamparany, T; Kouakou, K V; Michaud, V; Fernández-Pinero, J; Gallardo, C; Le Potier, M-F; Rabenarivahiny, R; Couacy-Hymann, E; Raherimandimby, M; Albina, E

    2016-08-01

    The performance of Whatman 3-MM filter papers for the collection, drying, shipment and long-term storage of blood at ambient temperature, and for the detection of African swine fever virus and antibodies was assessed. Conventional and real-time PCR, viral isolation and antibody detection by ELISA were performed on paired samples (blood/tissue versus dried-blood 3-MM filter papers) collected from experimentally infected pigs and from farm pigs in Madagascar and Côte d'Ivoire. 3-MM filter papers were used directly in the conventional and real-time PCR without previous extraction of nucleic acids. Tests that performed better with 3-MM filter papers were in descending order: virus isolation, real-time UPL PCR and conventional PCR. The analytical sensitivity of real-time UPL PCR on filter papers was similar to conventional testing (virus isolation or conventional PCR) on organs or blood. In addition, blood-dried filter papers were tested in ELISA for antibody detection and the observed sensitivity was very close to conventional detection on serum samples and gave comparable results. Filter papers were stored up to 9 months at 20-25°C and for 2 months at 37°C without significant loss of sensitivity for virus genome detection. All tests on 3-MM filter papers had 100% specificity compared to the gold standards. Whatman 3-MM filter papers have the advantage of being cheap and of preserving virus viability for future virus isolation and characterization. In this study, Whatman 3-MM filter papers proved to be a suitable support for the collection, storage and use of blood in remote areas of tropical countries without the need for a cold chain and thus provide new possibilities for antibody testing and virus isolation. PMID:25430732

  2. The effectiveness of acetic acid wash protocol and the interpretation patterns of blood contaminated cervical cytology ThinPrep® specimens

    PubMed Central

    Frisch, Nora K.; Ahmed, Yasin; Sethi, Seema; Neill, Daniel; Kalinicheva, Tatyana; Shidham, Vinod

    2015-01-01

    Background: ThinPrep® (TP) cervical cytology, as a liquid-based method, has many benefits but also a relatively high unsatisfactory rate due to debris/lubricant contamination and the presence of blood. These contaminants clog the TP filter and prevent the deposition of adequate diagnostic cells on the slide. An acetic acid wash (AAW) protocol is often used to lyse red blood cells, before preparing the TP slides. Design: From 23,291 TP cervical cytology specimens over a 4-month period, 2739 underwent AAW protocol due to initial unsatisfactory smear (UNS) with scant cellularity due to blood or being grossly bloody. Randomly selected 2739 cervical cytology specimens which did not undergo AAW from the same time period formed the control (non-AAW) group. Cytopathologic interpretations of AAW and non-AAW groups were compared using the Chi-square test. Results: About 94.2% of the 2739 cases which underwent AAW were subsequently satisfactory for evaluation with interpretations of atypical squamous cells of undetermined significance (ASCUS) 4.9% (135), low-grade squamous intraepithelial lesions (LSIL) 3.7% (102), and high-grade squamous intraepithelial lesions (HSIL) 1% (28). From the 2739 control cases, 96.3% were satisfactory with ASCUS 5.5% (151), LSIL 5.1% (139), and HSIL 0.7% (19). The prevalence of ASCUS interpretations was similar (P = 0.33). Although there were 32% more HSIL interpretations in the AAW group (28 in AAW vs. 19 in non-AAW), the difference was statistically insignificant (P = 0.18). AAW category; however, had significantly fewer LSIL interpretations (P = 0.02). The percentage of UNS cases remained higher in the AAW group with statistical significance (P < 0.01). Conclusions: While AAW had a significantly higher percent of UNS interpretations, the protocol was effective in rescuing 94.2% of specimens which otherwise may have been reported unsatisfactory. This improved patient care by avoiding a repeat test. The prevalence of ASCUS and HSIL

  3. Screening of high-risk Gaucher disease patients in Brazil using miniaturized dried blood spots and leukocyte techniques.

    PubMed

    Goldim, Mariana Pereira de Souza; Garcia, Cristina da Silva; de Castilhos, Cristina Dickie; Daitx, Vanessa Vitcoski; Mezzalira, Jamila; Breier, Ana Carolina; Cé, Jaqueline; Mello, Alexandre; Andrade, Carla Vieira; Sartori, Nicole; Coelho, Janice Carneiro

    2012-10-25

    This study investigates the miniaturization of the screening technique using dried blood spots on filter paper (DBS) to measure GBA and CT activities, and GBA and β-galactosidase activities in leukocytes. 274 DBS from individuals with suspected GD were screened for 1.5 years. Of these, we confirmed the diagnosis in 13.5%. The miniaturization of the DBS and leukocyte techniques afforded to reduce costs and sample size appropriate for a reliable diagnosis. PMID:22884741

  4. Further studies on the effects of diets containing dried coffee pulp: growth performance, blood and carcass characteristics of pigs.

    PubMed

    Okai, D B; Dabo, P

    1991-01-01

    4 groups of 5 pigs each were fed rations containing 0, 10, 20, or 30% of dried coffee pulp over a period of 10 weeks. The inclusion of these rations had no significant influence on the feed intake, growth rate and feed conversion efficiency. There were no significant differences in the blood parameters either (glucose, protein, P, Ca, cholesterol) or in the slaughter weight. Pigs fed the coffee pulp had less backfat and higher liver weights. PMID:1793431

  5. Dry period cooling ameliorates physiological variables and blood acid base balance, improving milk production in murrah buffaloes

    NASA Astrophysics Data System (ADS)

    Aarif, Ovais; Aggarwal, Anjali

    2016-03-01

    This study aimed to evaluate the impact of evaporative cooling during late gestation on physiological responses, blood gas and acid base balance and subsequent milk production of Murrah buffaloes. To investigate this study sixteen healthy pregnant dry Murrah buffaloes (second to fourth parity) at sixty days prepartum were selected in the months of May to June and divided into two groups of eight animals each. One group of buffaloes (Cooled/CL) was managed under fan and mist cooling system during dry period. Group second buffaloes (Noncooled/NCL) remained as control without provision of cooling during dry period. The physiological responses viz. Rectal temperature (RT), Respiratory rate (RR) and Pulse rate were significantly ( P < 0.05) lower in group 2, with the provision of cooling. Skin surface temperature at thorax was significantly lower in cooled group relative to noncooled group. Blood pH and pO2 were significantly ( P < 0.05) higher in heat stressed group as compared to the cooled group. pCO2, TCO2, HCO3, SBC, base excess in extracellular fluid (BEecf), base excess in blood (BEb), PCV and Hb were significantly ( P < 0.05) higher in cooled group as compared to noncooled group. DMI was significantly ( P < 0.05) higher in cooled relative to noncooled animals. Milk yield, FCM, fat yield, lactose yield and total solid yield was significantly higher ( P < 0.05) in cooled group of Murrah buffaloes.

  6. Dry period cooling ameliorates physiological variables and blood acid base balance, improving milk production in murrah buffaloes.

    PubMed

    Aarif, Ovais; Aggarwal, Anjali

    2016-03-01

    This study aimed to evaluate the impact of evaporative cooling during late gestation on physiological responses, blood gas and acid base balance and subsequent milk production of Murrah buffaloes. To investigate this study sixteen healthy pregnant dry Murrah buffaloes (second to fourth parity) at sixty days prepartum were selected in the months of May to June and divided into two groups of eight animals each. One group of buffaloes (Cooled/CL) was managed under fan and mist cooling system during dry period. Group second buffaloes (Noncooled/NCL) remained as control without provision of cooling during dry period. The physiological responses viz. Rectal temperature (RT), Respiratory rate (RR) and Pulse rate were significantly (P < 0.05) lower in group 2, with the provision of cooling. Skin surface temperature at thorax was significantly lower in cooled group relative to noncooled group. Blood pH and pO2 were significantly (P < 0.05) higher in heat stressed group as compared to the cooled group. pCO2, TCO2, HCO3, SBC, base excess in extracellular fluid (BEecf), base excess in blood (BEb), PCV and Hb were significantly (P < 0.05) higher in cooled group as compared to noncooled group. DMI was significantly (P < 0.05) higher in cooled relative to noncooled animals. Milk yield, FCM, fat yield, lactose yield and total solid yield was significantly higher (P < 0.05) in cooled group of Murrah buffaloes. PMID:26232368

  7. Validation and development of an immunonephelometric assay for the determination of alpha-1 antitrypsin levels in dried blood spots from patients with COPD*

    PubMed Central

    Zillmer, Laura Russo; Russo, Rodrigo; Manzano, Beatriz Martins; Ivanaga, Ivan; Nascimento, Oliver Augusto; de Souza, Altay Alves Lino; Santos, Gildo; Rodriguez, Francisco; Miravitlles, Marc; Jardim, José Roberto

    2013-01-01

    OBJECTIVE: To validate and develop an immunonephelometric assay for the determination of alpha-1 antitrypsin (AAT) levels in dried blood spots from COPD patients in Brazil. METHODS: We determined AAT levels in serum samples and dried blood spots from 192 COPD patients. For the preparation of dried blood spots, a disk (diameter, 6 mm) was placed into a tube, eluted with 200 µL of PBS, and stored overnight at 4ºC. All of the samples were analyzed by immunonephelometry in duplicate. We used the bootstrap resampling method in order to determine a cut-off point for AAT levels in dried blood spots. RESULTS: The correlation coefficient between the AAT levels in serum samples and those in dried blood spots was r = 0.45. For dried blood spots, the cut-off value was 2.02 mg/dL (97% CI: 1.45-2.64 mg/dL), with a sensitivity, specificity, positive predictive value, and negative predictive value of 100%, 95.7%, 27.2%, and 100%, respectively. CONCLUSIONS: This method for the determination of AAT levels in dried blood spots appears to be a reliable screening tool for patients with AAT deficiency. PMID:24310627

  8. Short communication: Effect of estrogen supplemented at dry-off on temporal changes in concentrations of lactose in blood plasma of Holstein cows.

    PubMed

    Gulay, M S; Hayen, M J; Head, H H; Bachman, K C

    2009-08-01

    The objective was to determine the effect of supplemental estrogen (estradiol cypionate, ECP) at dry-off on temporal changes in concentrations of lactose in blood plasma of Holstein cows as an indicator of rate of mammary involution. Thirty-two Holstein cows (8/group) were assigned randomly to 4 treatment groups: 30-d dry, 30-d dry + ECP, 60-d dry, and 60-d dry + ECP. A single injection (7.5 mL) of cottonseed oil (30- and 60-d dry) or ECP (15 mg) in oil (30- and 60-d dry + ECP) was administered intramuscularly at dry-off. Blood samples were collected from the coccygeal vein of all cows 24 h before dry-off and at dry-off, and then 8 samples were collected throughout the subsequent 48 h to monitor concentrations of lactose in blood plasma. No significant effects of ECP on the overall mean concentrations of lactose were detected. Concentrations of lactose increased and were greatest in blood collected 20 h (520.4 +/- 54.1, 268.1 +/- 48.2, 345.0 +/- 52.3, 418.4 +/- 49.8 microM, for the 4 treatment groups respective to the order listed above) after supplemental ECP and final milk removal. At 40 h, concentrations approached those observed 24 h before dry-off (140.5 +/- 52.1, 57.6 +/- 47.1, 90.1 +/- 51.4, 61.2 +/- 48.4 microM, respectively). Concentrations of lactose at 20 h were positively correlated with milk yield of cows at dry-off. Similar temporal profiles of lactose in blood plasma of cows supplemented or not with ECP indicated that ECP at dry-off did not markedly alter the course of tight junction leakage that typically occurs in mammary epithelial tissue during progressive early involution when milk removal is discontinued. PMID:19620664

  9. Enzymatic diagnosis of Fabry disease using a fluorometric assay on dried blood spots: An alternative methodology.

    PubMed

    Caudron, Eric; Prognon, Patrice; Germain, Dominique P

    2015-12-01

    Fabry disease (FD, OMIM#301500) is an X-linked lysosomal storage disorder caused by the functional deficiency of α-galactosidase A, a lysosomal enzyme. A method to screen for FD in large populations has been developed using a fluorometric assay of α-galactosidase A activity in dried blood spots (DBS) on filter paper. However, results can be influenced by quenching of fluorescence by haemoglobin which, together with small sample size, may result in a low light emission signal. An alternative, simple and sensitive fluorometric assay was developed for the determination of α-galactosidase A activity in DBS. The assay uses 4-methylumbelliferyl-α-d-galactose as an artificial substrate. To minimize the risk of false-positives, zinc sulfate was used for protein precipitation to stop the enzymatic reaction and eliminate interfering species (hemoglobin). Samples from 209 individuals (60 hemizygotes, 68 heterozygotes, and 81 controls) were tested to establish reference values for the assay. The mean α-galactosidase A activity of the 81 controls was 9.1 ± 3.3 μmol h(-1) L(-1) (mean ± SD). All 60 hemizygotes affected with FD had AGAL activities below 1.7 μmol h(-1) L(-1) (0.2 ± 0.3 μmol h(-1) L(-1)). For the 68 heterozygous females, AGAL activity ranged from 0 to 12.6 μmol h(-1) L(-1) (3.5 ± 2.7 μmol h(-1) L(-1)). Two-thirds of the female patients could be identified using the enzymatic assay and a cut-off level of 40% of the median control value (<3.4 μmol h(-1) L(-1)). Our fluorometric assay using zinc sulfate protein precipitation was shown to have similar sensitivity and robustness while reducing the risk of false positive results due to quenching of 4-MU fluorescence by haemoglobin. PMID:26520229

  10. Development and implementation challenges of a quality assured HIV infant diagnosis program in Nigeria using dried blood spots and DNA polymerase chain reaction.

    PubMed

    Audu, Rosemary; Onwuamah, Chika; Salu, Olumuyiwa; Okwuraiwe, Azuka; Ou, Chin-Yih; Bolu, Omotayo; Bond, Kyle B; Diallo, Karidia; Lu, Lydia; Jelpe, Tapdiyel; Okoye, McPaul; Ngige, Evelyn; Vertefeuille, John

    2015-04-01

    Nigeria has one of the highest HIV burdens as well as mother-to-infant transmission rates in the world. A pilot program using polymerase chain reaction (PCR)-based testing of dried blood spot (DBS) specimens was implemented to enable early identification of HIV-infected infants and timely referral and linkage to care. From February 2007 to October 2008, whole blood was collected by finger prick to prepare DBS from infants <18 months presenting in six public mother-and-child health facilities in Lagos, Nigeria. The DBS were tested using the Roche Amplicor HIV-1 DNA Test, v1.5. To monitor laboratory testing quality, all of the PCR-positive and 10% of the PCR-negative DBS were retested by the same method at another reference laboratory. Three hundred and sixty-five randomly selected infants were screened using HIV rapid tests (RT) according to the national algorithm and RT-negative and PCR-positive specimens were also tested using Genscreen enzyme-linked immunosorbent assay (EIA) (Bio-Rad, France). The turnaround time (TAT) from sample collection, testing, and dispatching of results from each health facility was monitored. A total of 1,273 infants with a median age of 12.6 weeks (1 day to 71.6 weeks) participated in the program and 280 (22.0%) were PCR positive. HIV transmission levels varied greatly in the different health facilities ranging from 7.1% to 38.4%. Infants aged 48 to 72 weeks had the highest level of PCR positivity (41.1%). All PCR-positive specimens were confirmed by retesting. The mean turnaround time from DBS collection to returning of the laboratory result to the health facilities was 25 days. Three infants were found to be HIV antibody negative by rapid tests but were positive by both PCR and the fourth generation EIA. The DBS-based PCR program accurately identified all of the HIV-infected infants. However, many programmatic challenges related to the laboratory and TAT were identified. PMID:25381805

  11. Simplifying sample pretreatment: application of dried blood spot (DBS) method to blood samples, including postmortem, for UHPLC-MS/MS analysis of drugs of abuse.

    PubMed

    Odoardi, Sara; Anzillotti, Luca; Strano-Rossi, Sabina

    2014-10-01

    The complexity of biological matrices, such as blood, requires the development of suitably selective and reliable sample pretreatment procedures prior to their instrumental analysis. A method has been developed for the analysis of drugs of abuse and their metabolites from different chemical classes (opiates, methadone, fentanyl and analogues, cocaine, amphetamines and amphetamine-like substances, ketamine, LSD) in human blood using dried blood spot (DBS) and subsequent UHPLC-MS/MS analysis. DBS extraction required only 100μL of sample, added with the internal standards and then three droplets (30μL each) of this solution were spotted on the card, let dry for 1h, punched and extracted with methanol with 0.1% of formic acid. The supernatant was evaporated and the residue was then reconstituted in 100μL of water with 0.1% of formic acid and injected in the UHPLC-MS/MS system. The method was validated considering the following parameters: LOD and LOQ, linearity, precision, accuracy, matrix effect and dilution integrity. LODs were 0.05-1ng/mL and LOQs were 0.2-2ng/mL. The method showed satisfactory linearity for all substances, with determination coefficients always higher than 0.99. Intra and inter day precision, accuracy, matrix effect and dilution integrity were acceptable for all the studied substances. The addition of internal standards before DBS extraction and the deposition of a fixed volume of blood on the filter cards ensured the accurate quantification of the analytes. The validated method was then applied to authentic postmortem blood samples. PMID:24814508

  12. Comparative Evaluation of Four Real-Time PCR Methods for the Quantitative Detection of Epstein-Barr Virus from Whole Blood Specimens.

    PubMed

    Buelow, Daelynn; Sun, Yilun; Tang, Li; Gu, Zhengming; Pounds, Stanley; Hayden, Randall

    2016-07-01

    Monitoring of Epstein-Barr virus (EBV) load in immunocompromised patients has become integral to their care. An increasing number of reagents are available for quantitative detection of EBV; however, there are little published comparative data. Four real-time PCR systems (one using laboratory-developed reagents and three using analyte-specific reagents) were compared with one another for detection of EBV from whole blood. Whole blood specimens seeded with EBV were used to determine quantitative linearity, analytical measurement range, lower limit of detection, and CV for each assay. Retrospective testing of 198 clinical samples was performed in parallel with all methods; results were compared to determine relative quantitative and qualitative performance. All assays showed similar performance. No significant difference was found in limit of detection (3.12-3.49 log10 copies/mL; P = 0.37). A strong qualitative correlation was seen with all assays that used clinical samples (positive detection rates of 89.5%-95.8%). Quantitative correlation of clinical samples across assays was also seen in pairwise regression analysis, with R(2) ranging from 0.83 to 0.95. Normalizing clinical sample results to IU/mL did not alter the quantitative correlation between assays. Quantitative EBV detection by real-time PCR can be performed over a wide linear dynamic range, using three different commercially available reagents and laboratory-developed methods. EBV was detected with comparable sensitivity and quantitative correlation for all assays. PMID:27157323

  13. Chemical composition and biological value of spray dried porcine blood by-products and bone protein hydrolysate for young chickens.

    PubMed

    Jamroz, D; Wiliczkiewicz, A; Orda, J; Skorupińska, J; Słupczyńska, M; Kuryszko, J

    2011-10-01

    The chemical composition of spray dried porcine blood by-products is characterised by wide variation in crude protein contents. In spray dried porcine blood plasma (SDBP) it varied between 670-780 g/kg, in spray dried blood cells (SDBC) between 830-930 g/kg, and in bone protein hydrolysate (BPH) in a range of 740-780 g/kg. Compared with fish meal, these feeds are poor in Met and Lys. Moreover, in BPH deep deficits of Met, Cys, Thr and other amino acids were found. The experiment comprised 7 dietary treatments: SDBP, SDBC, and BPH, each at an inclusion rate of 20 or 40 g/kg diet, plus a control. The addition of 20 or 40 g/kg of the analysed meals into feeds for very young chickens (1-28 d post hatch) significantly decreased the body weight (BW) of birds. Only the treatments with 40 g/kg of SDBP and SDBC showed no significant difference in BW as compared with the control. There were no significant differences between treatments and type of meal for feed intake, haematocrit and haemoglobin concentrations in blood. Addition of bone protein and blood cell meals to feed decreased the IgG concentration in blood and caused shortening of the femur and tibia bones. However, changes in the mineral composition of bones were not significantly affected by the type of meal used. The blood by-products, which are rich in microelements, improved retention of Ca and Cu only. In comparison to control chickens, significantly better accretion of these minerals was found in treatments containing 20 g/kg of SDBP or 40 g/kg of SDBC. Great variability in apparent ileal amino acid digestibility in chickens was determined. In this respect, some significant differences related to the type of meal fed were confirmed for Asp, Pro, Val, Tyr and His. In general, the apparent ileal digestibility of amino acids was about 2-3 percentage units better in chickens fed on diets containing the animal by products than in control birds. PMID:22029787

  14. Freeze drying of red blood cells: the use of directional freezing and a new radio frequency lyophilization device.

    PubMed

    Arav, Amir; Natan, Dity

    2012-08-01

    Red blood cell (RBC) units are administered routinely into patients expressing a wide range of acute and chronic conditions (e.g., anemia, traumatic bleeding, chronic diseases, and surgery). The modern blood banking system has been designed to answer this need and assure a continuous, high quality blood supply to patients. However, RBCs units can be stored under hypothermic conditions for only up to 42 days, which leads to periodic shortages. Cryopreservation can solve these shortages, but current freezing methods employ high glycerol concentrations, which need to be removed and the cells washed prior to transfusion, resulting in a long (more than 1 hour) and cumbersome washing step. Thus, frozen RBCs have limited use in acute and trauma situations. In addition, transportation of frozen samples is complicated and costly. Freeze drying (lyophilization) of RBCs has been suggested as a solution for these problems, since it will allow for a low weight sample to be stored at room temperature, but reaching this goal is not a simple task. We studied the effect of different solutions (IMT2 and IMT3) containing trehalose and antioxidants or trehalose and human serum albumin, respectively, on freezing/thawing and freeze drying of RBCs. In addition, we evaluated the effect of cells concentrations and cooling rates on the post thaw and post rehydration recoveries of the RBCs. Finally, we developed a new radio frequency (RF) lyophilization device for a more rapid and homogeneous sublimation process of the frozen RBCs samples. Recovery and free Hb were measured as well as oxygen association/dissociation and cell's deformability. We found that IMT3 (0.3 M trehalose and 10% HSA) solution that was directionally frozen at a rapid interface velocity of 1 mm/sec (resulting in a cooling rate of 150°C/min) yielded the best results (better than IMT2 solution and slow interface velocity). Freeze thawing gave 100% survival, while freeze drying followed by rehydration with 20% dextran-40k

  15. Dried Blood Spot Methodology in Combination With Liquid Chromatography/Tandem Mass Spectrometry Facilitates the Monitoring of Teriflunomide

    PubMed Central

    Lunven, Catherine; Turpault, Sandrine; Beyer, Yann-Joel; O'Brien, Amy; Delfolie, Astrid; Boyanova, Neli; Sanderink, Ger-Jan; Baldinetti, Francesca

    2016-01-01

    Background: Teriflunomide, a once-daily oral immunomodulator approved for treatment of relapsing-remitting multiple sclerosis, is eliminated slowly from plasma. If necessary to rapidly lower plasma concentrations of teriflunomide, an accelerated elimination procedure using cholestyramine or activated charcoal may be used. The current bioanalytical assay for determination of plasma teriflunomide concentration requires laboratory facilities for blood centrifugation and plasma storage. An alternative method, with potential for greater convenience, is dried blood spot (DBS) methodology. Analytical and clinical validations are required to switch from plasma to DBS (finger-prick sampling) methodology. Methods: Using blood samples from healthy subjects, an LC-MS/MS assay method for quantification of teriflunomide in DBS over a range of 0.01–10 mcg/mL was developed and validated for specificity, selectivity, accuracy, precision, reproducibility, and stability. Results were compared with those from the current plasma assay for determination of plasma teriflunomide concentration. Results: Method was specific and selective relative to endogenous compounds, with process efficiency ∼88%, and no matrix effect. Inaccuracy and imprecision for intraday and interday analyses were <15% at all concentrations tested. Quantification of teriflunomide in DBS assay was not affected by blood deposit volume and punch position within spot, and hematocrit level had a limited but acceptable effect on measurement accuracy. Teriflunomide was stable for at least 4 months at room temperature, and for at least 24 hours at 37°C with and without 95% relative humidity, to cover sampling, drying, and shipment conditions in the field. The correlation between DBS and plasma concentrations (R2 = 0.97), with an average blood to plasma ratio of 0.59, was concentration independent and constant over time. Conclusions: DBS sampling is a simple and practical method for monitoring teriflunomide

  16. The stability of amitriptyline N-oxide and clozapine N-oxide on treated and untreated dry blood spot cards.

    PubMed

    Temesi, David; Swales, John; Keene, Warren; Dick, Samuel

    2013-03-25

    Procedures for drug monitoring based on Dried Blood Spot (DBS) sampling are gaining acceptance for an increasing number of clinical and preclinical applications, where ease of use, small sample requirement, and improved sample stability have been shown to offer advantages over blood tube sampling. However, to-date, the vast majority of this work has described the analysis of well characterized drugs. Using amitriptyline, clozapine, and their potentially labile N-oxide metabolites as model compounds, we consider the merits of using DBS for discovery pharmacokinetic (PK) studies where the metabolic fate of test compounds are often unknown. Both N-oxide metabolites reverted to parent compound under standard drying (2hr) and extraction conditions. Card type significantly affected the outcome, with 14% and 22% degradation occurring for clozapine-N-oxide and amitriptyline-N-oxide on a brand of untreated DBS cards, compared to 59 and 88% on a brand of treated DBS cards. Enrichment of the parent compound ex vivo leads to overestimation of circulating blood concentration and inaccurate determination of the PK profile. PMID:23333684

  17. Using dried blood spot sampling to improve data quality and reduce animal use in mouse pharmacokinetic studies.

    PubMed

    Wickremsinhe, Enaksha R; Perkins, Everett J

    2015-03-01

    Traditional pharmacokinetic analysis in nonclinical studies is based on the concentration of a test compound in plasma and requires approximately 100 to 200 μL blood collected per time point. However, the total blood volume of mice limits the number of samples that can be collected from an individual animal-often to a single collection per mouse-thus necessitating dosing multiple mice to generate a pharmacokinetic profile in a sparse-sampling design. Compared with traditional methods, dried blood spot (DBS) analysis requires smaller volumes of blood (15 to 20 μL), thus supporting serial blood sampling and the generation of a complete pharmacokinetic profile from a single mouse. Here we compare plasma-derived data with DBS-derived data, explain how to adopt DBS sampling to support discovery mouse studies, and describe how to generate pharmacokinetic and pharmacodynamic data from a single mouse. Executing novel study designs that use DBS enhances the ability to identify and streamline better drug candidates during drug discovery. Implementing DBS sampling can reduce the number of mice needed in a drug discovery program. In addition, the simplicity of DBS sampling and the smaller numbers of mice needed translate to decreased study costs. Overall, DBS sampling is consistent with 3Rs principles by achieving reductions in the number of animals used, decreased restraint-associated stress, improved data quality, direct comparison of interanimal variability, and the generation of multiple endpoints from a single study. PMID:25836959

  18. Using Dried Blood Spot Sampling to Improve Data Quality and Reduce Animal Use in Mouse Pharmacokinetic Studies

    PubMed Central

    Wickremsinhe, Enaksha R; Perkins, Everett J

    2015-01-01

    Traditional pharmacokinetic analysis in nonclinical studies is based on the concentration of a test compound in plasma and requires approximately 100 to 200 µL blood collected per time point. However, the total blood volume of mice limits the number of samples that can be collected from an individual animal—often to a single collection per mouse—thus necessitating dosing multiple mice to generate a pharmacokinetic profile in a sparse-sampling design. Compared with traditional methods, dried blood spot (DBS) analysis requires smaller volumes of blood (15 to 20 µL), thus supporting serial blood sampling and the generation of a complete pharmacokinetic profile from a single mouse. Here we compare plasma-derived data with DBS-derived data, explain how to adopt DBS sampling to support discovery mouse studies, and describe how to generate pharmacokinetic and pharmacodynamic data from a single mouse. Executing novel study designs that use DBS enhances the ability to identify and streamline better drug candidates during drug discovery. Implementing DBS sampling can reduce the number of mice needed in a drug discovery program. In addition, the simplicity of DBS sampling and the smaller numbers of mice needed translate to decreased study costs. Overall, DBS sampling is consistent with 3Rs principles by achieving reductions in the number of animals used, decreased restraint-associated stress, improved data quality, direct comparison of interanimal variability, and the generation of multiple endpoints from a single study. PMID:25836959

  19. Can ultrashort-TE (UTE) MRI sequences on a 3-T clinical scanner detect signal directly from collagen protons: freeze-dry and D2 O exchange studies of cortical bone and Achilles tendon specimens.

    PubMed

    Ma, Ya-Jun; Chang, Eric Y; Bydder, Graeme M; Du, Jiang

    2016-07-01

    Ultrashort-TE (UTE) sequences can obtain signal directly from short-T2 , collagen-rich tissues. It is generally accepted that bound and free water can be detected with UTE techniques, but the ability to detect protons directly on the collagen molecule remains controversial. In this study, we investigated the potential of UTE sequences on a 3-T clinical scanner to detect collagen protons via freeze-drying and D2 O-H2 O exchange studies. Experiments were performed on bovine cortical bone and human Achilles tendon specimens, which were either subject to freeze-drying for over 66 h or D2 O-H2 O exchange for 6 days. Specimens were imaged using two- and three-dimensional UTE with Cones trajectory techniques with a minimum TE of 8 μs at 3 T. UTE images before treatment showed high signal from all specimens with bi-component T2 * behavior. Bovine cortical bone showed a shorter T2 * component of 0.36 ms and a longer T2 * component of 2.30 ms with fractions of 78.2% and 21.8% by volume, respectively. Achilles tendon showed a shorter T2 * component of 1.22 ms and a longer T2 * component of 15.1 ms with fractions of 81.1% and 18.9% by volume, respectively. Imaging after freeze-drying or D2 O-H2 O exchange resulted in either the absence or near-absence of signal. These results indicate that bound and free water are the sole sources of UTE signal in bovine cortical bone and human Achilles tendon samples on a clinical 3-T scanner. Protons on the native collagen molecule are not directly visible when imaged using UTE sequences. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27148693

  20. Quantitative determination of opioids in whole blood using fully automated dried blood spot desorption coupled to on-line SPE-LC-MS/MS.

    PubMed

    Verplaetse, Ruth; Henion, Jack

    2016-01-01

    Opioids are well known, widely used painkillers. Increased stability of opioids in the dried blood spot (DBS) matrix compared to blood/plasma has been described. Other benefits provided by DBS techniques include point-of-care collection, less invasive micro sampling, more economical shipment, and convenient storage. Current methodology for analysis of micro whole blood samples for opioids is limited to the classical DBS workflow, including tedious manual punching of the DBS cards followed by extraction and liquid chromatography-tandem mass spectrometry (LC-MS/MS) bioanalysis. The goal of this study was to develop and validate a fully automated on-line sample preparation procedure for the analysis of DBS micro samples relevant to the detection of opioids in finger prick blood. To this end, automated flow-through elution of DBS cards was followed by on-line solid-phase extraction (SPE) and analysis by LC-MS/MS. Selective, sensitive, accurate, and reproducible quantitation of five representative opioids in human blood at sub-therapeutic, therapeutic, and toxic levels was achieved. The range of reliable response (R(2)  ≥0.997) was 1 to 500 ng/mL whole blood for morphine, codeine, oxycodone, hydrocodone; and 0.1 to 50 ng/mL for fentanyl. Inter-day, intra-day, and matrix inter-lot accuracy and precision was less than 15% (even at lower limits of quantitation (LLOQ) level). The method was successfully used to measure hydrocodone and its major metabolite norhydrocodone in incurred human samples. Our data support the enormous potential of DBS sampling and automated analysis for monitoring opioids as well as other pharmaceuticals in both anti-doping and pain management regimens. PMID:26607771

  1. Estimation and Correction of the Blood Volume Variations of Dried Blood Spots Using a Postcolumn Infused-Internal Standard Strategy with LC-Electrospray Ionization-MS.

    PubMed

    Liao, Hsiao-Wei; Lin, Shu-Wen; Chen, Guan-Yuan; Kuo, Ching-Hua

    2016-06-21

    Dried blood spots (DBSs) have had a long history in disease screening in newborns but have gained attention in recent years in the medical care of adults because of the growing importance of personalized medicine. DBSs have several advantages, such as easy transportation, cost-effectiveness, and minimally invasive biological sampling. There are two strategies to process DBS samples. One method takes a fixed diameter of subsample, and another requires the extraction of the whole spot. The whole-spot extraction method is less affected by hematocrit-caused errors, but it requires calibration of the blood volume. We propose a novel strategy using a postcolumn infused-internal standard (PCI-IS) method with liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) for estimating and correcting blood volume variations on the DBS cards. By using PCI-IS to measure the extent of ion suppression in the first ion suppression zone in the chromatogram, the blood volume on the DBS cards can be calculated and further calibrated. We used reference blood samples with different volumes (5 to 25 μL) to construct a calibration curve between the blood volume and the extent of ion suppression. The calibration curve was used to estimate the blood volume on the DBS cards collected from 6 volunteers, with 5 designated volumes from each volunteer. The estimation accuracy of the PCI-IS method was between 74.5% and 120.3%. The validated PCI-IS method was used to estimate and calibrate blood volume variation and also to quantify the voriconazole concentration for 26 patients undergoing voriconazole therapy. A high correlation was found for the quantification results between the DBS samples and the conventionally used plasma samples (r = 0.97). The PCI-IS method was demonstrated to be a simple and accurate method for estimating and calibrating the blood volume variation on DBS cards, which greatly facilitates using the DBS method for therapeutic drug monitoring (TDM) for

  2. Liquid extraction surface analysis field asymmetric waveform ion mobility spectrometry mass spectrometry for the analysis of dried blood spots.

    PubMed

    Griffiths, Rian L; Dexter, Alex; Creese, Andrew J; Cooper, Helen J

    2015-10-21

    Liquid extraction surface analysis (LESA) is a surface sampling technique that allows electrospray mass spectrometry analysis of a wide range of analytes directly from biological substrates. Here, we present LESA mass spectrometry coupled with high field asymmetric waveform ion mobility spectrometry (FAIMS) for the analysis of dried blood spots on filter paper. Incorporation of FAIMS in the workflow enables gas-phase separation of lipid and protein molecular classes, enabling analysis of both haemoglobin and a range of lipids (phosphatidylcholine or phosphatidylethanolamine, and sphingomyelin species) from a single extraction sample. The work has implications for multiplexed clinical assays of multiple analytes. PMID:26198596

  3. Dried blood spot analysis of (+) and (-) darunavir enantiomers on immobilized amylose tris-(3, 5-dimethylphenylcarbamate) LC and its application to pharmacokinetics.

    PubMed

    Ramisetti, Nageswara Rao; Arnipalli, Manikanta Swamy; Nimmu, Narendra Varma

    2015-12-01

    Dried blood spot analysis is an innovative novel blood sampling technique gaining interest in drug discovery and development processes owing to its inherent advantages over the conventional whole blood, plasma or serum sample collection. The present manuscript describes the development and validation of a highly sensitive and precise method of evaluation of pharmacokinetics of (+) and (-) darunavir enantiomers on rat dried blood spots. The enantiomers on rat dried blood spots were extracted into methanol and separated by LC on a Chiralpak IA column using hexane and ethanol containing 0.1% DEA (75:25, v/v) as a mobile phase at 20°C; both the enantiomers were detected at 266 nm using a photodiode array detector. The method was validated in terms of selectivity, linearity, accuracy, precision and stability as per the US Food and Drug and Administration guidelines. The hematocrit effect on extraction recovery was evaluated and the mean recoveries of (-) and (+) enantiomers of darunavir from dried blood spots were found to be 85.76 and 88.91% respectively. The intra- and inter-day precision and accuracy were 3.1-8.4 and 0.8-4.8% respectively. The developed method was successfully applied to a pharmacokinetic study of (+) and (-) enantiomers of darunavir on rat dried blood spots. PMID:26081678

  4. Comparison of urine analysis and dried blood spot analysis for the detection of ephedrine and methylephedrine in doping control.

    PubMed

    Kojima, Asami; Nishitani, Yasunori; Sato, Mitsuhiko; Kageyama, Shinji; Dohi, Michiko; Okano, Masato

    2016-02-01

    When the misuse of stimulants is determined in doping control tests conducted during the in-competition period, athletes are asked to account for the violation of the rules. This study was designed to evaluate whether the urinary threshold values (10 µg/mL) for ephedrine and methylephedrine set by the World Anti-Doping Agency (WADA) can be exceeded after the oral administration of each substance (25 mg). In addition, the study describes the validity of a liquid chromatography-tandem mass spectrometric method using dried blood spot testing to detect ephedrine and methylephedrine by comparing it to a quantitative laboratory urine assay. After administration of ephedrine, the urinary concentration of ephedrine did not exceed the threshold at 4-10 h in two subjects, whereas the threshold was exceeded in both the subjects at 12 h after administration. For methylephedrine, the urinary concentrations of all the subjects failed to reach the threshold for up to 10 h after administration. The concentrations reached the threshold at 12-24 h after administration in some volunteers. In contrast, the blood concentrations of ephedrine and methylephedrine reached their maximum levels at 2-8 h after administration. The blood concentrations showed a low inter-individual variability, and the results suggested that the urinary excretion of ephedrine and methylephedrine can be strongly affected by urine pH and/or urine volume. These facts suggest that urinary concentrations cannot reflect the psychoactive level of ephedrines in circulation. Thus, dried blood analysis might be suitable for the adequate detection of stimulants during in-competition testing. PMID:25869885

  5. Multicenter Comparison of ESP Culture System II with BACTEC 460TB and with Lowenstein-Jensen Medium for Recovery of Mycobacteria from Different Clinical Specimens, Including Blood

    PubMed Central

    Tortoli, Enrico; Cichero, Paola; Chirillo, M. Gabriella; Gismondo, M. Rita; Bono, Letizia; Gesu, Giampietro; Simonetti, M. Tullia; Volpe, Gisella; Nardi, Giampietro; Marone, Piero

    1998-01-01

    The recently developed ESP Culture System II (AccuMed, Chicago, Ill.) was compared with radiometric BACTEC 460TB (Becton Dickinson, Towson, Md.) and with Lowenstein-Jensen medium for recovery of mycobacteria from over 2,500 clinical specimens both of respiratory and nonrespiratory origin, including blood. The majority of the 219 mycobacterial isolates (129) belonged to the Mycobacterium tuberculosis complex, followed by 37 isolates of the Mycobacterium avium complex (MAC) and 53 isolates of eight other mycobacterial species. Rates of recovery obtained with BACTEC, ESP, and Lowenstein-Jensen medium were 89, 79, and 64%, respectively, with such differences being statistically significant. Different media and systems appeared to behave differently when the more frequently detected organisms were considered: M. tuberculosis complex isolates grew better with BACTEC, and MAC isolates grew better with ESP. An analysis of the combinations of Lowenstein-Jensen medium with BACTEC and with ESP did not reveal significant differences in recovery rates. With regard to the times needed for the detection of positive cultures, they were significantly longer on Lowenstein-Jensen medium (average, 28 days) than with the remaining two systems, between which there was no difference (average, 18 days). We conclude, therefore, that the ESP system, when used in combination with a solid medium, performs as well as the thoroughly validated radiometric BACTEC system and offers the advantages of full automation and absence of radioisotopes. PMID:9574709

  6. Biomonitoring using dried blood spots: Detection of ochratoxin A and its degradation product 2’R‐ochratoxin A in blood from coffee drinkers*

    PubMed Central

    Cramer, Benedikt; Osteresch, Bernd; Muñoz, Katherine A.; Hillmann, Hartmut; Sibrowski, Walter

    2015-01-01

    Scope In this study, human exposure to the mycotoxin ochratoxin A (OTA) and its thermal degradation product 2’R‐ochratoxin A (2’R‐OTA, previously named as 14R‐Ochratoxin A [22]) through coffee consumption was assessed. LC‐MS/MS and the dried blood spot (DBS) technique were used for the analysis of blood samples from coffee and noncoffee drinkers (n = 50), and food frequency questionnaires were used to document coffee consumption. Methods and results For the detection of OTA and 2’R‐OTA in blood, a new sensitive and efficient sample preparation method based on DBS was established and validated. Using this technique 2’R‐OTA was for the first time detected in biological samples. Comparison between coffee drinkers and noncoffee drinkers showed for the first time that 2’R‐OTA was only present in blood from the first group while OTA could be found in both groups in a mean concentration of 0.21 μg/L. 2’R‐OTA mean concentration was 0.11 μg/L with a maximum concentration of 0.414 μg/L. Thus, in average 2’R‐OTA was approx. half the concentration of OTA but in some cases even exceeded OTA levels. No correlation between the amounts of coffee consumption and OTA or 2’R‐OTA levels was observed. Conclusion The results of this study revealed for the first time a high exposure of coffee consumers to 2’R‐OTA, a compound formed from OTA during coffee roasting. Since little information is available regarding toxicity and possible carcinogenicity of this compound, further OTA monitoring in blood including 2’R‐OTA is advisable. PMID:26012425

  7. [Investigations on the usefulness of the dry chemistry blood anaylsis system SPOTCHEM SP-4410in laboratory diagnosis of cattle].

    PubMed

    Lorenz, I; Aigner, M; Klee, W

    2001-01-01

    The usefulness of the dry-chemistry blood analyzer, SPOTCHEM SP-4410, for analysis of bovine blood chemistry was studied in a veterinary clinic. The control serum Precipath-U, Boehringer-Mannheim, was used to measure precision within each run and between days. The coefficients of variation (CV) ranged between 1.54% and 4.86%, with the exception of albumin and creatine phosphokinase showing a CV of 6.3% and 10.03% for between-day precision. For methodological comparison bovine serum samples were assayed with both the SPOTCHEM SP-4410 and the automated blood analyzer HITACHI 705, which served as a wet-chemistry reference system. The following analytes were measured: glucose, urea, creatinine, total protein, albumin, total bilirubin and the enzymes AST, CPK and gamma-GT. For hemoglobin, which was measured in heparinized whole blood, the CO oximeter 855, CIBA-CORNING, was used as a reference system. The comparative analysis showed very good correlation in eight of ten parameters and their correlation coefficients (r) ranged between 0.962 and 0.998. Only the correlation coefficients of the analysis of total bilirubin (r = 0.903) and albumin (r = 0.771) were less satisfactory. The recovery test was carried out with the two parameters glucose and blood urea. The recovery of glucose was 93.7% and of urea 98.8%. The SPOTCHEM SP-4410 is easy to use and proved to be reliable and accurate, and therefore it seems to be useful for analysis of bovine blood samples. PMID:11225499

  8. A New Method to Quantify Ifosfamide Blood Levels Using Dried Blood Spots and UPLC-MS/MS in Paediatric Patients with Embryonic Solid Tumours

    PubMed Central

    Chávez-Pacheco, Juan L.; Navas, Carlos F.; Demetrio, Joel A.; Alemón-Medina, Radamés; Trujillo, Francisca; Pérez, Martín; Zapata, Martha M.; Cárdenas, Rocío; Salinas, Citlaltepetl; Aquino, Arnoldo; Velázquez-Cruz, Rafael; Castillejos, Manuel-de-Jesús

    2015-01-01

    Ifosfamide blood concentrations are necessary to monitor its therapeutic response, avoiding any adverse effect. We developed and validated an analytical method by UPLC-MS/MS to quantify ifosfamide in dried blood spots (DBS). Blood samples were collected on Whatman 903® filter paper cards. Five 3 mm disks were punched out from each dried blood spot. Acetonitrile and ethyl acetate were used for drug extraction. Chromatographic separation was carried out in an Acquity UPLC equipment with a BEH-C18 column, 2.1 x 100 mm, 1.7 μm (Waters®). The mobile phase consisted in 5 mM ammonium formate and methanol:acetonitrile (40:48:12 v/v/v) at 0.2 mL/min. LC-MS/MS detection was done by ESI+ and multiple reaction mode monitoring, ionic transitions were m/z1+ 260.99 > 91.63 for ifosfamide and 261.00 > 139.90 for cyclophosphamide (internal standard). This method was linear within a 100–10000 ng/mL range and it was accurate, precise and selective. Ifosfamide samples in DBS were stable for up to 52 days at -80°C. The procedure was tested in 14 patients, ages 1 month to 17 years (9 males and 5 females), with embryonic tumours treated with ifosfamide, alone or combined, at a public tertiary referral hospital. Ifosfamide blood levels ranged from 11.1 to 39.7 μmol/L at 12 hours after the last infusion, while 24-hour levels ranged from 0.7–19.7 μmol/L. The median at 12 hours was 19.5 μmol/L (Q25 14.4–Q75 29.0) and 3.8 μmol/L (Q25 1.5–Q75 9.9) at 24 hours, p<0.001. This method is feasible to determine ifosfamide plasma levels in paediatric patients. PMID:26600181

  9. Detection of HIV type 1 env subtypes A, B, C, and E in Asia using dried blood spots: a new surveillance tool for molecular epidemiology.

    PubMed

    Cassol, S; Weniger, B G; Babu, P G; Salminen, M O; Zheng, X; Htoon, M T; Delaney, A; O'Shaughnessy, M; Ou, C Y

    1996-10-10

    Global surveillance of HIV-1 subtypes for genetic characterization is hampered by the biohazard of processing and the difficulties of shipping whole blood or cells from many developing country regions. We developed a technique for the direct automated sequencing of viral DNA from dried blood spot (DBS) specimens collected on absorbent paper, which can be mailed unrefrigerated in sturdy paper envelopes with low biohazard risk. DBS were collected nonrandomly from HIV-1-infected, mostly asymptomatic, patients in five Asian countries in 1991, and shipped via airmail or hand carried without refrigeration to Bangkok, and then transshipped to North America for processing. After more than 2 years of storage, including 6 months at ambient temperatures, proviral DNA in the DBS was amplified by nested PCR, and a 389-nucleotide segment of the C2-V3 env gene region was sequenced, from which 287 base pairs were aligned and subtyped by phylogenetic analysis with neighbor-joining and other methods. From southern India, there were 25 infections with subtype C and 2 with subtype A. From Myanmar (Burma), we identified the first subtype E infection, as well as six subtype BB, a distinct cluster within subtype B that was first discovered in Thailand and that has now appeared in China, Malaysia, and Japan. From southwest China, one BB was identified, while a "classical" B typical of North American and European strains was found in Indonesia. From Thailand, five DBS of ambiguous serotype were identified as three B, one BB, and one E. A blinded control serotype E specimen was correctly identified, but a serotype BB control was not tested. Most HIV-1 in southern India appears to be env subtype C, with rare A, as others have reported in western and northern India. The subtypes BB and E in Myanmar, and the BB in China, suggest epidemiological linkage with these subtypes in neighboring Thailand. DBS are a practical, economical technique for conducting large-scale molecular epidemiological

  10. Top-Down Proteomics and Direct Surface Sampling of Neonatal Dried Blood Spots: Diagnosis of Unknown Hemoglobin Variants

    NASA Astrophysics Data System (ADS)

    Edwards, Rebecca L.; Griffiths, Paul; Bunch, Josephine; Cooper, Helen J.

    2012-11-01

    We have previously shown that liquid microjunction surface sampling of dried blood spots coupled with high resolution top-down mass spectrometry may be used for screening of common hemoglobin variants HbS, HbC, and HbD. In order to test the robustness of the approach, we have applied the approach to unknown hemoglobin variants. Six neonatal dried blood spot samples that had been identified as variants, but which could not be diagnosed by current screening methods, were analyzed by direct surface sampling top-down mass spectrometry. Both collision-induced dissociation and electron transfer dissociation mass spectrometry were employed. Four of the samples were identified as β-chain variants: two were heterozygous Hb D-Iran, one was heterozygous Hb Headington, and one was heterozygous Hb J-Baltimore. The fifth sample was identified as the α-chain variant heterozygous Hb Phnom Penh. Analysis of the sixth sample suggested that it did not in fact contain a variant. Adoption of the approach in the clinic would require speed in both data collection and interpretation. To address that issue, we have compared manual data analysis with freely available data analysis software (ProsightPTM). The results demonstrate the power of top-down proteomics for hemoglobin variant analysis in newborn samples.

  11. Blood

    MedlinePlus

    ... solid part of your blood contains red blood cells, white blood cells, and platelets. Red blood cells (RBC) deliver oxygen from your lungs to your tissues and organs. White blood cells (WBC) fight infection and are part of your ...

  12. Transport of viral specimens.

    PubMed Central

    Johnson, F B

    1990-01-01

    The diagnosis of viral infections by culture relies on the collection of proper specimens, proper care to protect the virus in the specimens from environmental damage, and use of an adequate transport system to maintain virus activity. Collection of specimens with swabs that are toxic to either virus or cell culture should be avoided. A variety of transport media have been formulated, beginning with early bacteriological transport media. Certain swab-tube combinations have proven to be both effective and convenient. Of the liquid transport media, sucrose-based and broth-based media appear to be the most widely accepted and used. Studies on virus stability show that most viruses tested are sufficiently stable in transport media to withstand a transport time of 1 to 3 days. Some viruses may withstand longer transport times. In many cases, it is not necessary to store virus specimens in a refrigerator or send them to the laboratory on wet ice or frozen on dry ice. However, the specimen should not be exposed to environmental extremes. Modern viral transport media allow for more effective use of viral culture and culture enhancement techniques for the diagnosis of human viral infections. PMID:2187591

  13. Correlation of serum and dried blood spot results for quantitation of Schistosoma circulating anodic antigen: a proof of principle.

    PubMed

    Downs, Jennifer A; Corstjens, Paul L A M; Mngara, Julius; Lutonja, Peter; Isingo, Raphael; Urassa, Mark; Kornelis, Dieuwke; van Dam, Govert J

    2015-10-01

    Circulating anodic antigen (CAA) testing is a powerful, increasingly-used tool for diagnosis of active schistosome infection. We sought to determine the feasibility and reliability of measuring CAA in blood spots collected on Whatman 903 protein saver cards, which are the predominant filter papers used worldwide for dried blood spot (DBS) research and clinical care. CAA was eluted from blood spots collected from 19 individuals onto Whatman 903 cards in Mwanza, Tanzania, and the assay was optimized to achieve CAA ratios comparable to those obtained from the spots' corresponding serum samples. The optimized assay was then used to determine the correlation of serum samples (n=16) with DBS from cards that had been stored for 8 years at ambient temperature. Using a DBS volume equivalent to approximately four times the quantity of serum, CAA testing in DBS had a sensitivity of 76% and a specificity of 79% compared to CAA testing in serum. CAA testing was reliable in samples eluted from Whatman 903 cards that had been stored for 8 years at ambient temperature. The overall kappa coefficient was 0.53 (standard error 0.17, p<0.001). We conclude that CAA can be reliably and accurately measured in DBS collected onto the filter paper that is most commonly used for clinical care and research, and that can be stored from prolonged periods of time. This finding opens new avenues for future work among more than 700million individuals living in areas worldwide in which schistosomes are endemic. PMID:26149541

  14. External Quality Control for Dried Blood Spot Based C-reactive Protein Assay: Experience from the Indonesia Family Life Survey and the Longitudinal Aging Study in India

    PubMed Central

    Hu, Peifeng; Herningtyas, Elizabeth H.; Kale, Varsha; Crimmins, Eileen M.; Risbud, Arun R.; McCreath, Heather; Lee, Jinkook; Strauss, John; O’Brien, Jennifer C.; Bloom, David E.; Seeman, Teresa E.

    2015-01-01

    Measurement of C-reactive protein, a marker of inflammation, in dried blood spots has been increasingly incorporated in community-based social surveys internationally. Although the dried blood spot based CRP assay protocol has been validated in the United States, it remains unclear whether laboratories in other less developed countries can generate C-reactive protein results of similar quality. We therefore conducted external quality monitoring for dried blood spot based C-reactive protein measurement for the Indonesia Family Life Survey and the Longitudinal Aging Study in India. Our results show that dried blood spot based C-reactive protein results in these two countries have excellent and consistent correlations with serum-based values and dried blood spot based results from the reference laboratory in the United States. Even though the results from duplicate samples may have fluctuations in absolute values over time, the relative order of C-reactive protein levels remains similar and the estimates are reasonably precise for population-based studies that investigate the association between socioeconomic factors and health. PMID:25879265

  15. Field Evaluation of Dried Blood Spots for Routine HIV-1 Viral Load and Drug Resistance Monitoring in Patients Receiving Antiretroviral Therapy in Africa and Asia

    PubMed Central

    Monleau, Marjorie; Eymard-Duvernay, Sabrina; Dagnra, Anoumou; Kania, Dramane; Ngo-Giang-Huong, Nicole; Touré-Kane, Coumba; Truong, Lien X. T.; Chaix, Marie-Laure; Delaporte, Eric; Ayouba, Ahidjo; Peeters, Martine

    2014-01-01

    Dried blood spots (DBS) can be used in developing countries to alleviate the logistic constraints of using blood plasma specimens for viral load (VL) and HIV drug resistance (HIVDR) testing, but they should be assessed under field conditions. Between 2009 and 2011, we collected paired plasma-DBS samples from treatment-experienced HIV-1-infected adults in Burkina Faso, Cameroon, Senegal, Togo, Thailand, and Vietnam. The DBS were stored at an ambient temperature for 2 to 4 weeks and subsequently at −20°C before testing. VL testing was performed on the plasma samples and DBS using locally available methods: the Abbott m2000rt HIV-1 test, generic G2 real-time PCR, or the NucliSENS EasyQ version 1.2 test. In the case of virological failure (VF), i.e., a plasma VL of ≥1,000 copies/ml, HIVDR genotyping was performed on paired plasma-DBS samples. Overall, we compared 382 plasma-DBS sample pairs for DBS VL testing accuracy. The sensitivities of the different assays in different laboratories for detecting VF using DBS varied from 75% to 100% for the m2000rt test in labs B, C, and D, 91% to 93% for generic G2 real-time PCR in labs A and F, and 85% for the NucliSENS test in lab E. The specificities varied from 82% to 97% for the m2000rt and NucliSENS tests and reached only 60% for the generic G2 test. The NucliSENS test showed good agreement between plasma and DBS VL but underestimated the DBS VL. The lowest agreement was observed for the generic G2 test. Genotyping was successful for 96/124 (77%) DBS tested, and 75/96 (78%) plasma-DBS pairs had identical HIVDR mutations. Significant discrepancies in resistance interpretations were observed in 9 cases, 6 of which were from the same laboratory. DBS can be successfully used as an alternative to blood plasma samples for routine VL and HIVDR monitoring in African and Asian settings. However, the selection of an adequate VL measurement method and the definition of the VF threshold should be considered, and laboratory

  16. Direct multiplex assay of enzymes in dried blood spots by tandem mass spectrometry for the newborn screening of lysosomal storage disorders.

    PubMed

    Gelb, Michael H; Turecek, Frantisek; Scott, C Ron; Chamoles, Nestor A

    2006-01-01

    Tandem mass spectrometry is currently used in newborn screening programmes to quantify the level of amino acids and acylcarnitines in dried blood spots for detection of metabolites associated with treatable diseases. We have developed assays for lysosomal enzymes in rehydrated dried blood spots in which a set of substrates is added and the set of corresponding enzymatic products are quantified using tandem mass spectrometry with the aid of mass-differentiated internal standards. We have developed a multiplex assay of the set of enzymes that, when deficient, cause the lysosomal storage disorders Fabry, Gaucher, Hurler, Krabbe, Niemann-Pick A/B and Pompe diseases. These diseases were selected because treatments are now available or expected to emerge shortly. The discovery that acarbose is a selective inhibitor of maltase glucoamylase allows the Pompe disease enzyme, acid alpha-glucosidase, to be selectively assayed in white blood cells and dried blood spots. When tested with dried blood spots from 40 unaffected individuals and 10-12 individuals with the lysosomal storage disorder, the tandem mass spectrometry assay led to the correct identification of the affected individuals with 100% sensitivity. Many of the reagents needed for the new assays are commercially available, and those that are not are being prepared under Good Manufacturing Procedures for approval by the FDA. Our newborn screening assay for Krabbe disease is currently being put in place at the Wadsworth Center in New York State for the analysis of approximately 1000 dried blood spots per day. Summary We have developed tandem mass spectrometry for the direct assay of lysosomal enzymes in rehydrated dried blood spots that can be implemented for newborn screening of lysosomal storage disorders. Several enzymes can be analysed by a single method (multiplex analysis) and in a high-throughput manner appropriate for newborn screening laboratories. PMID:16763908

  17. Identification and quantification of psychoactive drugs in whole blood using dried blood spot (DBS) by ultra-performance liquid chromatography tandem mass spectrometry.

    PubMed

    Kyriakou, Chrystalla; Marchei, Emilia; Scaravelli, Giulia; García-Algar, Oscar; Supervía, August; Graziano, Silvia

    2016-09-01

    A procedure based on ultra-high-pressure liquid chromatography tandem mass spectrometry has been developed for the determination of twenty three psychoactive drugs and metabolites in whole blood using dried blood spot (DBS). Chromatographic separation was achieved at ambient temperature using a reverse-phase column and a linear gradient elution with two solvents: 0.1% formic acid in acetonitrile and 5mM ammonium formate at pH 3. The mass spectrometer was operated in positive ion mode, using multiple reaction monitoring via positive electro-spray ionization. The method was linear from the limit of quantification (5ng/ml for all the analytes apart from 15ng/ml for Δ-9-tetrahydrocannabinol and metabolites) to 500ng/ml, and showed good correlation coefficients (r(2)=0.990) for all substances. Analytical recovery of analytes under investigation was always higher than 75% and intra-assay and inter-assay precision and accuracy always better than 15%. Using the validated method, ten DBS samples, collected at the hospital emergency department in cases of acute drug intoxication, were found positive to one or more psychoactive drugs. Our data support the potential of DBS sampling for non invasive monitoring of exposure/intoxication to psychoactive drugs. PMID:27232151

  18. Fast transmethylation of total lipids in dried blood by microwave irradiation and its application to a population study.

    PubMed

    Lin, Yu Hong; Hanson, Jennifer A; Strandjord, Sarah E; Salem, Nicholas M; Dretsch, Michael N; Haub, Mark D; Hibbeln, Joseph R

    2014-08-01

    A methodology combining finger-pricked blood sampling, microwave accelerated fatty acid assay, fast gas chromatography data acquisition, and automated data processing was developed, evaluated and applied to a population study. Finger-pricked blood was collected on filter paper previously impregnated with 0.05 mg of the antioxidant butylated hydroxytoluene and air-dried at room temperature. Transmethylation was accelerated by microwave irradiation in an explosion-proof multimode microwave reaction system. The chemical procedure was based on a one-step direct transmethylation procedure catalyzed by acetyl chloride. The short-term stability of PUFA in blood dried on filter paper and storage at room temperature was examined using venous blood. The recoveries ranged from 97 to 101 % for the categorized fatty acids as well as the ratios of n-6 to n-3 PUFA and the n-3 % highly unsaturated fatty acid. Specifically, recoveries were 99, 98, 97, and 97 % for linoleic acid (18:2n-6), arachidonic acid (ARA), α-linolenic acid (ALA), and docosahexaenoic acid (DHA), respectively. The mol% (mean ± SD, 95 % confidence interval) of fatty acid composition in subjects from the population study was determined as 36.2 ± 3.8 (35.8, 36.7), 23.2 ± 3.0 (22.8, 23.5), 36.8 ± 3.5 (36.4, 37.2) and 3.79 ± 1.0 (3.68, 3.91) for the saturated, monounsaturated, n-6 and n-3 PUFA, respectively. Individually, the mean mol% (95 % CI) was 22.6 (22.3, 22.9) for 18:2n-6, 9.5 (9.3, 9.7) for ARA, 0.51 (0.49, 0.53) for ALA, 0.42 (0.38, 0.47) for eicosapentaenoic acid (EPA), and 1.67 (1.61, 1.73) for DHA. This methodology provides an accelerated yet high-efficiency, chemically safe, and temperature-controlled transmethylation, with diverse laboratory applications including population studies. PMID:24986160

  19. Fast Transmethylation of Total Lipids in Dried Blood by Microwave Irradiation and its Application to a Population Study

    PubMed Central

    Lin, Yu Hong; Hanson, Jennifer A.; Strandjord, Sarah E.; Salem, Nicholas M.; Dretsch, Michael N.; Haub, Mark D.; Hibbeln, Joseph R.

    2014-01-01

    A methodology combining finger-pricked blood sampling, microwave accelerated fatty acid assay, fast gas chromatography data acquisition, and automated data processing was developed, evaluated and applied to a population study. Finger-pricked blood was collected on filter paper previously impregnated with 0.05 mg of the antioxidant butylated hydroxytoluene and air-dried at room temperature. Transmethylation was accelerated by microwave irradiation in an explosion-proof multimode microwave reaction system. The chemical procedure was based on a one-step direct transmethylation procedure catalyzed by acetyl chloride. The short-term stability of PUFA in blood dried on filter paper and stored overnight at room temperature was examined using venous blood. The recoveries ranged from 97–101 % for the categorized fatty acids as well as the ratios of n-6 to n-3 PUFA and the n-3% highly unsaturated fatty acid. Specifically, recoveries were 99, 98, 97, and 97 % for linoleic acid (18:2n-6), arachidonic acid (ARA), α-linolenic acid (ALA), and docosahexaenoic acid (DHA), respectively. The mol% (mean ± SD, 95% confidence interval) of fatty acid composition in subjects from the population study was determined as 36.2±3.8 (35.8, 36.7), 23.2±3.0 (22.8, 23.5), 36.8±3.5 (36.4, 37.2) and 3.79±1.0 (3.68, 3.91) for the saturated, monounsaturated, n-6 and n-3 PUFA, respectively. Individually, the mean mol% (95% CI) was 22.6 (22.3, 22.9) for 18:2n-6, 9.5 (9.3, 9.7) for ARA, 0.51 (0.49, 0.53) for ALA, 0.42 (0.38, 0.47) for eicosapentaenoic acid (EPA), and 1.67 (1.61, 1.73) for DHA. This methodology provides an accelerated yet high-efficiency, chemically safe, and temperature-controlled transmethylation, with diverse laboratory applications including population studies. PMID:24986160

  20. Influence of Relative Humidity on the Spreading Dynamics of a Drying Drop of Whole Blood

    NASA Astrophysics Data System (ADS)

    Bou Zeid, Wassim; Brutin, David

    2013-11-01

    Newtonian and non-Newtonian fluids start spreading after coming into contact with a solid substrate till the anchoring of the triple line. Our experimental work aims to study the effect of the relative humidity on the spreading dynamics of drops of whole blood. Drops of blood of same volume (+/-4.8%) are injected using a digital micropipette and gently deposited onto microscope ultraclean glass substrates. Experiments are conducted in a glove box at ambient temperature and a range of investigated relative humidities between 13.0% and 80.0%. The recorded images are post-processed using ImageJ in which the position of the contact line is measured every 20 ms. We show that the spreading dynamics is, in a first time, governed by relative humidity and later no more influence by relative humidity. Two spreading regimes are observed and analyzed compared to classical viscous drops. In previous work, we show that relative humidity influences the contact angle and the initial wetting radius. In the first regime, we find a spreading power law exponent that decreases for an increasing relative humidity. In the second regime, all the data collapse on each other and the evolution of the dimensionless radius no more depend on relative humidity.

  1. Determination of Synacthen(®) in dried blood spots for doping control analysis using liquid chromatography tandem mass spectrometry.

    PubMed

    Tretzel, Laura; Thomas, Andreas; Geyer, Hans; Delahaut, Philippe; Schänzer, Wilhelm; Thevis, Mario

    2015-06-01

    Dried blood spot (DBS) sampling, a technique used for taking whole blood samples dried on a filter paper, was initially reported in 1963 by Robert Guthrie. While the diagnostic analysis of metabolic disorders in newborns was the focus of investigations at that time, the number of established applications for preclinical drug development, toxicological studies, and therapeutic drug monitoring increased enormously in the last decades. As a consequence of speed, simplicity, and minimal invasiveness, DBS recommends itself as the preferential technique in sports drug testing. The present approach highlights for the first time the development of a screening assay for the analysis of the synthetic human adrenocorticotropic hormone tetracosactide hexaacetate (Synacthen(®)) in DBS using liquid chromatography tandem mass spectrometry. Highly purified sample extracts were obtained by an advanced sample preparation procedure including the addition of an internal standard (d8-tetracosactide) and immunoaffinity purification. The method's overall recovery was 27.6 %, and the assay's imprecision was calculated between 8.1 and 17.9 % for intraday and 12.9 to 20.5 % for interday measurements. Stability of the synthetic peptide in DBS was shown for at least 10 days at room temperature and presents a major benefit, since a rapid degradation in conventionally applied matrices such as urine or plasma is well known. With a limit of detection of 50 pg/mL, a detection window of several hours is expected considering reported steady-state plasma levels of 300 pg/mL after intramuscular application of Synacthen(®) Depot (1 mg). The analysis of authentic DBS samples within the scope of an administration study with 250 μg Synacthen(®) (short stimulation test) demonstrated the great potential of the developed assay to simplify the analysis of Synacthen(®) for doping control purposes. PMID:25863802

  2. Dry Electrodes for ECG and Pulse Transit Time for Blood Pressure: A Wearable Sensor and Smartphone Communication Approach

    NASA Astrophysics Data System (ADS)

    Shyamkumar, Prashanth

    Cardiovascular Diseases (CVDs) have been a major cause for deaths in both men and women in United States. Cerebrovascular Diseases like Strokes are known to have origins in CVDs as well. Moreover, nearly 18 Million Americans have a history of myocardial infarction and are currently undergoing cardiac rehabilitation. Consequently, CVDs are the highest costing disease groups and cost more than all types of cancer combined. However, significant cost reduction is possible through the effective use of the vast advances in embedded and pervasive electronic devices for healthcare. These devices can automate and move a significant portion of disease management to the patient's home through cyber connectivity, a concept known as point-of-care (POC) diagnostics and healthcare services. POC can minimize hospital visits and potentially avoid admission altogether with prognostic tools that give advanced notice of any abnormalities or chronic illnesses so that the treatment can be planned in advance. The POC concept requires continuous remote health monitoring. Therefore, the various sensors needed for comprehensive monitoring need to be worn daily and throughout the day. Moreover, true "roaming" capability is necessary so that it does not restrict the user's travel or his/her quotidian activities. Two biomedical signals namely, Electrocardiogram (ECG) and Blood Pressure are important diagnostic tests in assessing the cardiac health of a person. To that end, the research presented in this thesis: First , describes the development of a remote monitoring solution based on Bluetooth(TM), smartphones and cyber infrastructure for cardiac care called e-nanoflex. Second, Sensors for ECG that are compatible with everyday life style namely, (a) dry, gel-less vertically aligned gold nanowire electrodes, (b) dry textile-based conductive sensor electrodes to address the need for this technology to monitor cardiovascular diseases in women are tested with e-nanoflex and discussed. Third, non

  3. Clinical Evaluation of an Affordable Qualitative Viral Failure Assay for HIV Using Dried Blood Spots in Uganda

    PubMed Central

    Balinda, Sheila N.; Ondoa, Pascale; Obuku, Ekwaro A.; Kliphuis, Aletta; Egau, Isaac; Bronze, Michelle; Kasambula, Lordwin; Schuurman, Rob; Spieker, Nicole; Rinke de Wit, Tobias F.; Kityo, Cissy

    2016-01-01

    Background WHO recommends regular viral load (VL) monitoring of patients on antiretroviral therapy (ART) for timely detection of virological failure, prevention of acquired HIV drug resistance (HIVDR) and avoiding unnecessary switching to second-line ART. However, the cost and complexity of routine VL testing remains prohibitive in most resource limited settings (RLS). We evaluated a simple, low–cost, qualitative viral–failure assay (VFA) on dried blood spots (DBS) in three clinical settings in Uganda. Methods We conducted a cross–sectional diagnostic accuracy study in three HIV/AIDS treatment centres at the Joint Clinical Research Centre in Uganda. The VFA employs semi-quantitative detection of HIV–1 RNA amplified from the LTR gene. We used paired dry blood spot (DBS) and plasma with the COBASAmpliPrep/COBASTaqMan, Roche version 2 (VLref) as the reference assay. We used the VFA at two thresholds of viral load, (>5,000 or >1,000 copies/ml). Results 496 paired VFA and VLref results were available for comparative analysis. Overall, VFA demonstrated 78.4% sensitivity, (95% CI: 69.7%–87.1%), 93% specificity (95% CI: 89.7%–96.4%), 89.3% accuracy (95% CI: 85%–92%) and an agreement kappa = 0.72 as compared to the VLref. The predictive values of positivity and negativity among patients on ART for >12 months were 72.7% and 99.3%, respectively. Conclusions VFA allowed 89% of correct classification of VF. Only 11% of the patients were misclassified with the potential of unnecessary or late switch to second–line ART. Our findings present an opportunity to roll out simple and affordable VL monitoring for HIV–1 treatment in RLS. PMID:26824465

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

    PubMed Central

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

    2003-01-01

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

  5. 50 CFR 14.24 - Scientific specimens.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 50 Wildlife and Fisheries 1 2013-10-01 2013-10-01 false Scientific specimens. 14.24 Section 14.24....24 Scientific specimens. Except for wildlife requiring a permit pursuant to parts 16, 17, 18, 21, 22 or 23 of this subchapter, dead, preserved, dried, or embedded scientific specimens or parts...

  6. 50 CFR 14.24 - Scientific specimens.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 50 Wildlife and Fisheries 1 2014-10-01 2014-10-01 false Scientific specimens. 14.24 Section 14.24....24 Scientific specimens. Except for wildlife requiring a permit pursuant to parts 16, 17, 18, 21, 22 or 23 of this subchapter, dead, preserved, dried, or embedded scientific specimens or parts...

  7. 50 CFR 14.24 - Scientific specimens.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 50 Wildlife and Fisheries 1 2011-10-01 2011-10-01 false Scientific specimens. 14.24 Section 14.24....24 Scientific specimens. Except for wildlife requiring a permit pursuant to parts 16, 17, 18, 21, 22 or 23 of this subchapter, dead, preserved, dried, or embedded scientific specimens or parts...

  8. 50 CFR 14.24 - Scientific specimens.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 50 Wildlife and Fisheries 1 2012-10-01 2012-10-01 false Scientific specimens. 14.24 Section 14.24....24 Scientific specimens. Except for wildlife requiring a permit pursuant to parts 16, 17, 18, 21, 22 or 23 of this subchapter, dead, preserved, dried, or embedded scientific specimens or parts...

  9. 50 CFR 14.24 - Scientific specimens.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 50 Wildlife and Fisheries 1 2010-10-01 2010-10-01 false Scientific specimens. 14.24 Section 14.24....24 Scientific specimens. Except for wildlife requiring a permit pursuant to parts 16, 17, 18, 21, 22 or 23 of this subchapter, dead, preserved, dried, or embedded scientific specimens or parts...

  10. Are higher blood mercury levels associated with dry eye symptoms in adult Koreans? A population-based cross-sectional study

    PubMed Central

    Chung, So-Hyang

    2016-01-01

    Objectives The purpose of this study was to investigate whether blood mercury concentrations associated with the presence of dry eye symptoms in a nationally representative Korean population. Methods Population-based prospective cross-sectional study using the heavy metal data set of the 2010–2012 Korean National Health and Nutrition Examination Survey (KNHANES). A total of 4761 adult Koreans were the eligible population in this study. Of the 7162 survey participants, 2401 were excluded because they were <19 years of age, there were missing data in the heavy metal data set, or they had diabetes, rheumatoid arthritis, thyroid disease, asthma, depression and/or under-the-eye surgery. Blood mercury levels were measured on the day the participants completed a questionnaire regarding the presence of dry eye symptoms (persistent dryness or eye irritation). The population was divided into low and high groups by median level (4.26 and 2.89 µg/L for males and females, respectively). Results Self-reported dry eye symptoms were present in 13.0% of the cohort. Participants with dry eye symptoms were significantly more likely to have blood mercury levels exceeding the median than those without dry eye symptoms (45.7% vs 51.7%, p=0.021). Logistic regression analysis showed that, after adjusting for age, gender, education, total household income, smoking status, heavy alcohol use, sleep time, perceived stress status, total cholesterol levels and atopy history, dry eye symptoms were significantly associated with blood mercury levels that exceeded the median (reference: lower mercury group; OR, 1.324; 95% CI 1.059 to 1.655; p<0.05). Conclusions High blood mercury levels were associated with dry eye symptoms in a nationally representative Korean population. PMID:27121705

  11. Dried Blood Spot Technique for the Monitoring of Ambrisentan, Bosentan, Sildenafil, and Tadalafil in Patients with Pulmonary Arterial Hypertension.

    PubMed

    Enderle, Yeliz; Meid, Andreas D; Friedrich, Jörg; Grünig, Ekkehard; Wilkens, Heinrike; Haefeli, Walter E; Burhenne, Jürgen

    2015-12-15

    Endothelin receptor antagonists (ERA) and phosphodiesterase 5 inhibitors (PDE5I) are long-term therapeutics for the treatment of pulmonary arterial hypertension (PAH). Their interindividual pharmacokinetic variability is remarkably large, and despite the seriousness of the disease, nonadherence is occurring. Therefore, methods to monitor sufficient circulating drug levels are essential. The objectives of this study were to develop and validate dried blood spot (DBS) assays for the quantification of ambrisentan, bosentan, sildenafil, tadalafil, and their main metabolites. We also quantified the influence of different hematocrit levels and assessed the correlation of simultaneously taken capillary whole blood (DBS) and venous plasma samples. The aliquot punches were extracted by liquid/liquid extraction followed by liquid chromatography/tandem mass spectrometry (LC/MS/MS) quantification methods. All assays fulfilled the requirements of the FDA and EMA guidelines for assay validation with a lower limit of quantification of 2.5 ng/mL for the ERAs, 5 ng/mL for sildenafil, and 10 ng/mL for tadalafil. All analytes were stable for at least 147 days when stored on DBS filter paper cards at room temperature in the dark. Due to poor distribution into erythrocytes, drug concentrations in DBS were always lower than in plasma, resulting in conversion factors of 1.58 for ambrisentan and sildenafil and 1.52 for bosentan and tadalafil. PMID:26583764

  12. Validation of an Optimized ELISA for Quantitative Assessment of Epstein-Barr Virus Antibodies from Dried Blood Spots.

    PubMed

    Eick, Geeta; Urlacher, Samuel S; McDade, Thomas W; Kowal, Paul; Snodgrass, J Josh

    2016-01-01

    Our objective was to validate a commercially available ELISA to measure antibody titers against Epstein-Barr virus (EBV) in dried blood spots (DBS) to replace a previously validated assay for DBS that is no longer available. We evaluated the precision, reliability, and stability of the assay for the measurement of EBV antibodies in matched plasma, fingerprick DBS, and venous blood DBS samples from 208 individuals. Effects of hematocrit and DBS sample matrix on EBV antibody determination were also investigated, and the cutoff for seropositivity in DBS was determined. A conversion equation was derived to enable comparison of results generated using this method with the former DBS method. There was a high correlation between plasma and DBS EBV antibody titers (R(2) = 0.93) with very little bias (-0.07 based on Bland-Altman analysis). The assay showed good linearity and did not appear to be affected by the DBS matrix, and physiological hematocrit levels had no effect on assay performance. There was reasonable agreement between DBS EBV titer estimates obtained using this assay and the previously validated assay (R(2) = 0.72). The commercially available ELISA assay for EBV antibody titers that we validated for use with DBS will facilitate continued investigation of EBV antibody titers in DBS. PMID:27337556

  13. Comparison of fresh, dried and stir-frying gingers in decoction with blood stasis syndrome in rats based on a GC-TOF/MS metabolomics approach.

    PubMed

    Han, YanQuan; Li, YuXin; Wang, YongZhong; Gao, JiaRong; Xia, LunZhu; Hong, Yan

    2016-09-10

    In China, ginger (Zingiberofficinale Rosc.) and its processed products, such as dried ginger and stir-frying ginger are commonly applied in traditional Chinese medicine (TCM). The paper presents the research on the effects of fresh ginger, dried ginger and stir-frying ginger extracts in blood stasis syndrome. First, a blood stasis syndrome rats model was established and then the hemorheological and blood coagulation activities were analyzed. Third, a sensitive, simple, and valid gas chromatography combined with time-of-flight mass spectrometry (GC-TOF/MS) method was established to compare the metabolic fingerprint coupled with multivariate analysis. The total 27 metabolites (16 in serum and 11 in urine) were identified and contributed to the blood stasis progress. These metabolites mainly involve six metabolism pathways in different impact-value. The altered efficacy index and metabolites can be regulated to normal levels by fresh ginger (FG), dried ginger (DG) and stir-frying ginger (SG). FG is the most effective as shown by the efficacy index, similarity analysis and peak intensity. The result presented here shows that metabolomics equipped with efficacy index makes it possible to study the blood stasis syndrome and to compare the effect and metabolites in fresh, dried and stir-frying gingers. The metabolomics approach can be recommended to study the pharmacological effect and mechanism of herbal drugs. PMID:27454085

  14. Simultaneous measurement of cyclosporin A and tacrolimus from dried blood spots by ultra high performance liquid chromatography tandem mass spectrometry.

    PubMed

    Hinchliffe, Edward; Adaway, Joanne E; Keevil, Brian G

    2012-02-01

    Cyclosporin A (CsA) and tacrolimus are immunosuppressant drugs principally used in solid organ transplant recipients. Therapeutic drug monitoring (TDM) of both drugs is essential to avoid toxicity related to overdosage, and transplant rejection from underdosage. This necessitates frequent hospital visits to phlebotomy services. Capillary blood sampling onto dried blood spots (DBS) provides numerous advantages to venous whole blood sampling, including the ability for patients to send DBS to the laboratory by post, significantly reducing the number of unnecessary hospital visits. We have developed a novel, simple and rapid method for the extraction and simultaneous UPLC-MS/MS measurement of both CsA and tacrolimus from DBS. The extraction method involved a simple 30 min hot solvent extraction with ultrasonication. Extract (10 μL) was injected onto a Waters Acquity UPLC column filter unit security frit, coupled to a Waters Acquity BEH C18 UPLC column, with methanolic mobile phase gradient elution. Eluant was connected to a Waters Quattro Premier XE tandem mass spectrometer operating in ES+ mode. We detected multiple reaction monitoring (MRM) transitions of m/z 1220>1203 and 1231.9>1215.1 for CsA and d12 CsA respectively which co-eluted at 1.30min, and 821.6>768.5 and 809.6>756.5 for tacrolimus and ascomycin respectively which co-eluted at 1.17 min. Ion suppression was negligible. Mean recovery was 95.5% for CsA and 92.8% for tacrolimus. Limit of detection and limit of quantitation were both 8.5 μg/L for CsA, and 0.5 and 2.3 μg/L respectively for tacrolimus. The assay was linear up to 1500μg/L for CsA (r(2)=0.9999), and up to 50 μg/L for tacrolimus (r(2)=0.9994). Mean intra assay imprecision, inter assay imprecision and bias were all <10% for both CsA and tacrolimus. DBS were stable for at least 14 days at room temperature. Comparison of the DBS UPLC-MS/MS method and the routine venous whole blood LC-MS/MS assay demonstrated good agreement between the two methods

  15. Dried blood spots and sparse sampling: a practical approach to estimating pharmacokinetic parameters of caffeine in preterm infants

    PubMed Central

    Patel, Parul; Mulla, Hussain; Kairamkonda, Venkatesh; Spooner, Neil; Gade, Sonya; Della Pasqua, Oscar; Field, David J.; Pandya, Hitesh C.

    2013-01-01

    Aims Dried blood spots (DBS) alongside micro‐analytical techniques are a potential solution to the challenges of performing pharmacokinetic (PK) studies in children. However, DBS methods have received little formal evaluation in clinical settings relevant to children. The aim of the present study was to determine a PK model for caffeine using a ‘DBS/microvolume platform’ in preterm infants. Methods DBS samples were collected prospectively from premature babies receiving caffeine for treatment of apnoea of prematurity. A non‐linear mixed effects approach was used to develop a population PK model from measured DBS caffeine concentrations. Caffeine PK parameter estimates based on DBS data were then compared with plasma estimates for agreement. Results Three hundred and thirty‐eight DBS cards for caffeine measurement were collected from 67 preterm infants (birth weight 0.6–2.11 kg). 88% of cards obtained were of acceptable quality and no child had more than 10 DBS samples or more than 0.5 ml of blood taken over the study period. There was good agreement between PK parameters estimated using caffeine concentrations from DBS samples (CL = 7.3 ml h−1 kg−1; V = 593 ml kg−1; t1/2 = 57 h) and historical caffeine PK parameter estimates based on plasma samples (CL = 4.9–7.9 ml h−1 kg−1; V = 640–970 ml kg−1; t1/2 = 101–144 h). We also found that changes in blood haematocrit may significantly confound estimates of caffeine PK parameters based on DBS data. Conclusions This study demonstrates that DBS methods can be applied to PK studies in a vulnerable population group and are a practical alternative to wet matrix sampling techniques. PMID:22822712

  16. Validation and Clinical Evaluation of a Novel Method To Measure Miltefosine in Leishmaniasis Patients Using Dried Blood Spot Sample Collection

    PubMed Central

    Rosing, H.; Hillebrand, M. J. X.; Blesson, S.; Mengesha, B.; Diro, E.; Hailu, A.; Schellens, J. H. M.; Beijnen, J. H.

    2016-01-01

    To facilitate future pharmacokinetic studies of combination treatments against leishmaniasis in remote regions in which the disease is endemic, a simple cheap sampling method is required for miltefosine quantification. The aims of this study were to validate a liquid chromatography-tandem mass spectrometry method to quantify miltefosine in dried blood spot (DBS) samples and to validate its use with Ethiopian patients with visceral leishmaniasis (VL). Since hematocrit (Ht) levels are typically severely decreased in VL patients, returning to normal during treatment, the method was evaluated over a range of clinically relevant Ht values. Miltefosine was extracted from DBS samples using a simple method of pretreatment with methanol, resulting in >97% recovery. The method was validated over a calibration range of 10 to 2,000 ng/ml, and accuracy and precision were within ±11.2% and ≤7.0% (≤19.1% at the lower limit of quantification), respectively. The method was accurate and precise for blood spot volumes between 10 and 30 μl and for Ht levels of 20 to 35%, although a linear effect of Ht levels on miltefosine quantification was observed in the bioanalytical validation. DBS samples were stable for at least 162 days at 37°C. Clinical validation of the method using paired DBS and plasma samples from 16 VL patients showed a median observed DBS/plasma miltefosine concentration ratio of 0.99, with good correlation (Pearson's r = 0.946). Correcting for patient-specific Ht levels did not further improve the concordance between the sampling methods. This successfully validated method to quantify miltefosine in DBS samples was demonstrated to be a valid and practical alternative to venous blood sampling that can be applied in future miltefosine pharmacokinetic studies with leishmaniasis patients, without Ht correction. PMID:26787691

  17. Dried Blood as an Alternative to Plasma or Serum for Trypanosoma cruzi IgG Detection in Screening Programs

    PubMed Central

    Holguín, Africa; Norman, Francesca; Martín, Leticia; Mateos, María Luisa; Chacón, Jesús; López-Vélez, Rogelio

    2013-01-01

    Trypanosoma cruzi serological screening is recommended for people potentially exposed to this parasite in countries where Trypanosoma cruzi is endemic and those where it is not endemic. Blood samples on filter paper may be a practical alternative to plasma/serum for antibody detection. Using the Architect Chagas assay, we detected the presence of IgG against T. cruzi in matched serum and dried blood spots (DBS) collected from 147 patients residing in Madrid, Spain, who had potential previous exposure to T. cruzi. The κ statistic for the DBS/serum proportion of agreement for the detection of antibodies against T. cruzi was 0.803, considering an S/CO (assay result unit; chemiluminescent signal from the sample [S] divided by the mean chemiluminescent signal for the three calibrators used in the test [CO]) cutoff value of ≥1.00. The relative sensitivity of the Architect test using DBS increased from 95.2% to 98.8% when the cutoff was lowered from ≥1.00 to ≥0.88, while the relative specificity decreased from 84.1% to 71.6%. Overall, the median S/CO values for DBS were significantly lower than those for serum (2.6 versus 6.5; P < 0.001). Discrepancies that occurred with the use of DBS included 10 false positives (with low S/CO values in 9 cases [median, 2.13]) and 4 false negatives, with mean S/CO values of 0.905 (gray zone). Using DBS plus a highly sensitive and specific enzyme-linked immunosorbent assay (ELISA) may be a simple and reliable method for detecting IgG against T. cruzi when blood sampling by venipuncture is not feasible. This method may also reduce the false-negative rates observed with some rapid diagnostic tests. The lower relative sensitivity compared to the reference method may be increased by lowering the optical density threshold. PMID:23740927

  18. Estimation of HIV-1 DNA Level Interfering with Reliability of HIV-1 RNA Quantification Performed on Dried Blood Spots Collected from Successfully Treated Patients.

    PubMed

    Zida, Sylvie; Tuaillon, Edouard; Barro, Makoura; Kwimatouo Lekpa Franchard, Arnaud; Kagoné, Thérèse; Nacro, Boubacar; Ouedraogo, Abdoul Salam; Bolloré, Karine; Sanosyan, Armen; Plantier, Jean-Christophe; Meda, Nicolas; Sangaré, Lassana; Rouzioux, Christine; Rouet, François; Kania, Dramane

    2016-06-01

    The impact of HIV-1 DNA coamplification during HIV-1 RNA quantification on dried blood spots (DBS) was explored. False-positive HIV RNA detection (22/62, 35%) was associated with high HIV-1 DNA levels. Specificity of HIV-1 RNA assays on DBS should be evaluated following manufacturer protocols on samples with HIV-1 DNA levels of ≥1,000 copies/10(6) peripheral blood mononuclear cells. PMID:27008874

  19. Enhanced Diagnosis of Pneumococcal Bacteremia Using Antigen- and Molecular-Based Tools on Blood Specimens in Mali and Thailand: A Prospective Surveillance Study.

    PubMed

    Moïsi, Jennifer C; Moore, Matthew; Carvalho, Maria da Gloria; Sow, Samba O; Siludjai, Duangkamon; Knoll, Maria Deloria; Tapia, Milagritos; Baggett, Henry C

    2016-02-01

    Prior antibiotic use, contamination, limited blood volume, and processing delays reduce yield of blood cultures for detection of Streptococcus pneumoniae. We performed immunochromatographic testing (ICT) on broth from incubated blood culture bottles and real-time lytA polymerase chain reaction (PCR) on broth and whole blood and compared findings to blood culture in patients with suspected bacteremia. We selected 383 patients in Mali and 586 patients in Thailand based on their blood culture results: 75 and 31 were positive for pneumococcus, 100 and 162 were positive for other pathogens, and 208 and 403 were blood culture negative, respectively. ICT and PCR of blood culture broth were at least 87% sensitive and 97% specific compared with blood culture; whole blood PCR was 75-88% sensitive and 96-100% specific. Pneumococcal yields in children < 5 years of age increased from 2.9% to 10.7% in Mali with > 99% of additional cases detected by whole blood PCR, and from 0.07% to 5.1% in Thailand with two-thirds of additional cases identified by ICT. Compared with blood culture, ICT and lytA PCR on cultured broth were highly sensitive and specific but their ability to improve pneumococcal identification varied by site. Further studies of these tools are needed before widespread implementation. PMID:26643535

  20. Enhanced Diagnosis of Pneumococcal Bacteremia Using Antigen- and Molecular-Based Tools on Blood Specimens in Mali and Thailand: A Prospective Surveillance Study

    PubMed Central

    Moïsi, Jennifer C.; Moore, Matthew; da Gloria Carvalho, Maria; Sow, Samba O.; Siludjai, Duangkamon; Knoll, Maria Deloria; Tapia, Milagritos; Baggett, Henry C.

    2016-01-01

    Prior antibiotic use, contamination, limited blood volume, and processing delays reduce yield of blood cultures for detection of Streptococcus pneumoniae. We performed immunochromatographic testing (ICT) on broth from incubated blood culture bottles and real-time lytA polymerase chain reaction (PCR) on broth and whole blood and compared findings to blood culture in patients with suspected bacteremia. We selected 383 patients in Mali and 586 patients in Thailand based on their blood culture results: 75 and 31 were positive for pneumococcus, 100 and 162 were positive for other pathogens, and 208 and 403 were blood culture negative, respectively. ICT and PCR of blood culture broth were at least 87% sensitive and 97% specific compared with blood culture; whole blood PCR was 75–88% sensitive and 96–100% specific. Pneumococcal yields in children < 5 years of age increased from 2.9% to 10.7% in Mali with > 99% of additional cases detected by whole blood PCR, and from 0.07% to 5.1% in Thailand with two-thirds of additional cases identified by ICT. Compared with blood culture, ICT and lytA PCR on cultured broth were highly sensitive and specific but their ability to improve pneumococcal identification varied by site. Further studies of these tools are needed before widespread implementation. PMID:26643535

  1. Analysis of tacrolimus and creatinine from a single dried blood spot using liquid chromatography tandem mass spectrometry

    PubMed Central

    Koop, Dennis R.; Bleyle, Lisa A.; Munar, Myrna; Cherala, Ganesh; Al-Uzri, Amira

    2014-01-01

    Long term therapeutic drug monitoring and assessment of renal function are required in renal transplant recipients on immunosuppressant therapy such as tacrolimus. Dry blood spots (DBS) have been used successfully in the clinic for many years and offers a convenient, simple and non-invasive method for repeated blood tests. We developed and performed a preliminary validation of a method for the analysis of tacrolimus and creatinine from a single DBS using liquid chromatography-tandem mass spectrometric (LC–MS/MS). Tacrolimus and creatinine were extracted from a 6 mm punch with a mixture of methanol/acetonitrile containing ascomycin and deuterated creatinine as internal standards. A 10 μl aliquot of the extract was analyzed directly after dilution for creatinine with normal phase high performance liquid chromatography and multiple reaction monitoring. The remainder of the extract was processed and analyzed for tacrolimus. The lower limit of quantification for tacrolimus was 1 ng/ml with accuracy of 0.34% bias and precision (CV) of 11.1%. The precision ranged from 1.33% to 7.68% and accuracy from −4.44% to 11.6% bias for the intra- and inter-day analysis. The lower limit of quantification of creatinine was 0.01 mg/dL with precision of 7.94%. Accuracy was based on recovery of additional creatinine spiked into whole blood samples and ranged from −2.45% bias at 5 mg/dL to 3.75% bias at 0.5 mg/dL. Intra- and inter-day precision was from 3.48 to 4.11%. The assay was further validated with DBS prepared from pediatric renal transplant recipients. There was excellent correlation between the levels of tacrolimus and creatinine obtained from the clinical laboratory and the DBS method developed. After additional validation, this assay may have a significant impact on compliance with medication intake as well as potentially lowering the cost associated with intravenous blood draws in clinical laboratories. PMID:23548676

  2. Blood

    MedlinePlus

    ... fight infection and are part of your body's defense system. Platelets help blood to clot when you have a cut or wound. Bone marrow, the spongy material inside your bones, makes new blood cells. Blood cells ...

  3. Sequencing from Dried Blood Spots in Infants with “False Positive” Newborn Screen for MCAD Deficiency

    PubMed Central

    McCandless, Shawn E.; Chandrasekar, Ram; Linard, Sharon; Kikano, Sandra; Rice, Lorrie

    2012-01-01

    Background Newborn screening (NBS) for medium chain acyl-CoA dehydrogenase deficiency (MCADD), one of the most common disorders identified, uses measurement of octanoylcarnitine (C8) from dried blood spots. In the state of Ohio, as in many places, primary care providers, with or without consultation from a metabolic specialist, may perform “confirmatory testing”, with the final diagnostic decision returned to the state. Confirmatory testing may involve measurement of metabolites, enzyme analysis, mutation screening, or sequencing. We now report sequencing results for infants said to have “false positive” NBS results for MCAD deficiency, or who died before confirmatory testing could be performed. Methods Dried blood spots (DBS) were obtained from all 18 available NBS cards identified as “false positive” by NBS for the 3 year period after screening began in Ohio in 2003 (N=20, thus 2 had no DBS available), and from all 6 infants with abnormal screens who died before confirmatory testing could be obtained. DNA extracted from DBS was screened for the common c.985A>G mutation in exon 11 of the ACADM gene, using a specific restriction digest method, followed by sequencing of the 12 exons, intron-exon junctions, and several hundred base pairs of the 5′ untranslated region. Results The NBS cut-off value for C8 used was 0.7 μmol/L. Sequencing of ACADM in six neonates with elevated C8 on NBS who died before confirmatory testing was obtained did not identify any significant variants in the coding region of the gene, suggesting that MCADD was not a contributing factor in these deaths. The mean C8 for the 18 surviving infants labeled as “False Positives” was 0.90 (95%CI 0.77-1.15), much lower than the mean value for confirmed cases. Ten of the 18 were premature births weighing <1200 g, the rest were normal sized and full term. Eight infants, mostly full term with appropriate birth weight, were heterozygous for the common c.985A>G mutation; one of those also

  4. Short Communication: Tenofovir Diphosphate in Dried Blood Spots As an Objective Measure of Adherence in HIV-Infected Women

    PubMed Central

    Castillo-Mancilla, Jose R.; Searls, Kristina; Caraway, Patricia; Zheng, Jia-Hua; Gardner, Edward M.; Predhomme, Julie; Bushman, Lane R.; Meditz, Amie L.

    2015-01-01

    Abstract Simple and reproducible tools to assess antiretroviral adherence are needed. A level of tenofovir diphosphate (TFV-DP) in dried blood spots (DBS) <1,250 fmol/punch is predicted to identify imperfect adherence. Herein we evaluated TFV-DP in DBS as a measure of adherence among HIV-infected women. DBS and peripheral blood mononuclear cells (PBMCs) were collected twice (∼1 week apart) in 35 well-controlled HIV-infected women [median age 42 years, 14 African American/black (AA)] receiving daily coformulated tenofovir/emtricitabine and either atazanavir/ritonavir (n=20) or raltegravir (n=16). TFV-DP in DBS and PBMCs was quantified by LC-MS/MS. Six-month adherence was measured as average days between monthly pharmacy refills. Data were loge transformed for analysis and presented as median (range); the correlation between continuous variables was analyzed using the Pearson correlation coefficient. The average TFV-DP between the two visits (aTFV-DP) in DBS and PBMCs was 1,874 (706–3,776) fmol/punch and 125 (1–278) fmol/106 cells, respectively. AA women had lower levels of aTFV-DP in DBS compared to whites (1,660 vs. 1,970 fmol/punch; p=0.04), with a viremic patient having the lowest drug levels (706 fmol/punch). Days between pharmacy refills were 34 (30–54) vs. 30 (26–40) in women with TFV-DP in DBS <1,250 vs. ≥1,250 fmol/punch (p=0.006). TFV-DP in DBS was negatively correlated with an increasing number of days between refills (r=−0.56, p=0.002). TFV-DP DBS was a reliable and objective measure of adherence in HIV-infected women based on a strong inverse relationship with pharmacy refill adherence. PMID:25328112

  5. Quantification of insulin-like growth factor-1 in dried blood spots for detection of growth hormone abuse in sport.

    PubMed

    Cox, Holly D; Rampton, Jessica; Eichner, Daniel

    2013-02-01

    There is significant evidence that athletes are using recombinant human growth hormone (rhGH) to enhance performance, and its use is banned by the World Anti-Doping Agency and professional sports leagues. Insulin-like growth factor-1 (IGF-1) is the primary mediator of growth hormone action and is used as a biomarker for the detection of rhGH abuse. The current biomarker-based method requires collection and expedited shipment of venous blood which is costly and may decrease the number of tests performed. Measurement of GH biomarkers in dried blood spots (DBS) would considerably simplify sample collection and shipping methods to allow testing of a greater number of samples regardless of location. A method was developed to quantify intact IGF-1 protein in DBS by liquid chromatography-tandem mass spectrometry. A step-wise acid-acetonitrile extraction was optimized to achieve a sensitive assay with a lower limit of quantification of 50 ng/mL. IGF-1 remained stable at room temperature for up to 8 days, which would allow shipment of DBS cards at ambient temperature. In a comparison between plasma concentrations of IGF-1 and concentrations measured from venous and finger prick DBS, there was good correlation and agreement, r(2) of 0.8551 and accuracy of 86-113 % for venous DBS and r(2) of 0.9586 and accuracy of 89-122 % for finger prick DBS. The method is intended for use as a rapid screening method for IGF-1 to be used in the biomarker method of rhGH abuse detection. PMID:23263515

  6. Short communication: Tenofovir diphosphate in dried blood spots as an objective measure of adherence in HIV-infected women.

    PubMed

    Castillo-Mancilla, Jose R; Searls, Kristina; Caraway, Patricia; Zheng, Jia-Hua; Gardner, Edward M; Predhomme, Julie; Bushman, Lane R; Anderson, Peter L; Meditz, Amie L

    2015-04-01

    Simple and reproducible tools to assess antiretroviral adherence are needed. A level of tenofovir diphosphate (TFV-DP) in dried blood spots (DBS) <1,250 fmol/punch is predicted to identify imperfect adherence. Herein we evaluated TFV-DP in DBS as a measure of adherence among HIV-infected women. DBS and peripheral blood mononuclear cells (PBMCs) were collected twice (∼1 week apart) in 35 well-controlled HIV-infected women [median age 42 years, 14 African American/black (AA)] receiving daily coformulated tenofovir/emtricitabine and either atazanavir/ritonavir (n=20) or raltegravir (n=16). TFV-DP in DBS and PBMCs was quantified by LC-MS/MS. Six-month adherence was measured as average days between monthly pharmacy refills. Data were loge transformed for analysis and presented as median (range); the correlation between continuous variables was analyzed using the Pearson correlation coefficient. The average TFV-DP between the two visits (aTFV-DP) in DBS and PBMCs was 1,874 (706-3,776) fmol/punch and 125 (1-278) fmol/10(6) cells, respectively. AA women had lower levels of aTFV-DP in DBS compared to whites (1,660 vs. 1,970 fmol/punch; p=0.04), with a viremic patient having the lowest drug levels (706 fmol/punch). Days between pharmacy refills were 34 (30-54) vs. 30 (26-40) in women with TFV-DP in DBS <1,250 vs. ≥1,250 fmol/punch (p=0.006). TFV-DP in DBS was negatively correlated with an increasing number of days between refills (r=-0.56, p=0.002). TFV-DP DBS was a reliable and objective measure of adherence in HIV-infected women based on a strong inverse relationship with pharmacy refill adherence. PMID:25328112

  7. Stability of HIV-1 Nucleic Acids in Dried Blood Spot Samples for HIV-1 Drug Resistance Genotyping

    PubMed Central

    Aitken, Susan C.; Wallis, Carole L.; Stevens, Wendy; de Wit, Tobias Rinke; Schuurman, Rob

    2015-01-01

    Dried blood spots (DBS) are an easy to collect sample-type that can stabilize biological material at ambient temperature for transport and storage, making them ideal for use in resource-limited settings (RLS). We investigated the effect of storage temperature and duration on ability to detect mixed HIV-1 viral RNA populations, and subsequently viral RNA populations in a background of proviral DNA. Part one of the study used DBS samples of whole blood spiked with specific quantities of HIV-1 subtype-B and -C RNA to study mixed virus population detection. Part two used DBS comprising of HIV-1 subtype-B proviral DNA containing U1 cells combined with HIV-1 subtype-C RNA to mimic HIV-1 infected clinical samples as a model system to study the relative stability of HIV-1 RNA and DNA in DBS. Prepared DBS were stored at -20°C and +30°C for periods of one day, one, two, and four weeks. Samples were genotyped to determine changes in the detection of mixtures in the sample over time. From two weeks onwards, storage at +30°C resulted in gradual, time-related reduction in the detection of mixed virus population at log10 VL 4.0 but not at log10 5.0. Proviral DNA and viral RNA were both stable for at least 52 weeks when stored at -20°C, compared to progressive RNA decay over time at +30°C. DBS storage conditions and duration had a significant effect on HIV-1 RNA amplification. Our results demonstrate that DBS storage at ambient temperature (+30°C) should not exceed two weeks, with long-term storage at -20°C or lower. PMID:26147689

  8. Stability of HIV-1 Nucleic Acids in Dried Blood Spot Samples for HIV-1 Drug Resistance Genotyping.

    PubMed

    Aitken, Susan C; Wallis, Carole L; Stevens, Wendy; de Wit, Tobias Rinke; Schuurman, Rob

    2015-01-01

    Dried blood spots (DBS) are an easy to collect sample-type that can stabilize biological material at ambient temperature for transport and storage, making them ideal for use in resource-limited settings (RLS). We investigated the effect of storage temperature and duration on ability to detect mixed HIV-1 viral RNA populations, and subsequently viral RNA populations in a background of proviral DNA. Part one of the study used DBS samples of whole blood spiked with specific quantities of HIV-1 subtype-B and -C RNA to study mixed virus population detection. Part two used DBS comprising of HIV-1 subtype-B proviral DNA containing U1 cells combined with HIV-1 subtype-C RNA to mimic HIV-1 infected clinical samples as a model system to study the relative stability of HIV-1 RNA and DNA in DBS. Prepared DBS were stored at -20 °C and +30 °C for periods of one day, one, two, and four weeks. Samples were genotyped to determine changes in the detection of mixtures in the sample over time. From two weeks onwards, storage at +30 °C resulted in gradual, time-related reduction in the detection of mixed virus population at log10 VL 4.0 but not at log10 5.0. Proviral DNA and viral RNA were both stable for at least 52 weeks when stored at -20 °C, compared to progressive RNA decay over time at +30 °C. DBS storage conditions and duration had a significant effect on HIV-1 RNA amplification. Our results demonstrate that DBS storage at ambient temperature (+30 °C) should not exceed two weeks, with long-term storage at -20 °C or lower. PMID:26147689

  9. Dried blood spot analysis of donepezil in support of a GLP 3-month dose-range finding study in rats.

    PubMed

    Meier-Davis, Susan R; Meng, Min; Yuan, Weiwei; Diehl, Lisa; Arjmand, Fatima M; Lucke, Rebecca M; Huang, Betsy; Wen, Jianye; Shudo, Jutaro; Nagata, Tetsuto

    2012-01-01

    Donepezil hydrochloride is a reversible acetyl cholinesterase inhibitor approved for Alzheimer disease treatment. As an alternate therapy, a donepezil hydrochloride transdermal patch is in development. Recommended nonclinical safety studies include a 3-month Good Laboratory Practice (GLP) dose-range finding (DRF) study prior to conducting the 2-year dermal carcinogenicity study in rats. Demonstration of systemic exposure is necessary to interpret the in vivo data. Previous nonclinical reports supporting oral dosing have utilized liquid chromatography tandem mass spectrometry (LC/MS/MS) to quantify donepezil concentrations in plasma. Smaller species with limited blood volumes do not allow serial sampling to derive the full pharmacokinetic profile from a single animal. Therefore, the option of another analytical method requiring decreased sample volumes is desirable as it would decrease the required number of animals while obtaining the complete profile. The dried blood spot (DBS) technique allows drug level measurement from a few microliters; however, the method is still not widely utilized in GLP studies. Because donepezil plasma levels are known by the oral route, DBS was used to bridge the previous oral data and to support a 13-week GLP DRF study for repeated topical application in rats, comparing oral administration with 4 topical formulations. The DBS method was validated and demonstrated robustness and reproducibility for application to the DRF study. The assay results were comparable to a previously reported plasma LC/MS/MS assay-derived pharmacokinetic profile and provided justification for selection of the topical formulation and dose levels for the subsequent dermal carcinogenicity study. PMID:22705881

  10. Comparison of the Babesia duncani (WA1) IgG detection rates among clinical sera submitted to a reference laboratory for WA1 IgG testing and blood donor specimens from diverse geographic areas of the United States.

    PubMed

    Prince, Harry E; Lapé-Nixon, Mary; Patel, Hemlata; Yeh, Cindy

    2010-11-01

    All reported cases of WA1 babesiosis have occurred in the Pacific coast region of the United States, suggesting that WA1 is limited to this geographic area. However, we detected WA1 IgG in 27% of clinical sera sent to our laboratory for WA1 IgG testing from across the United States over a 2-year period, suggesting that exposure to WA1 or a closely related organism occurs outside Pacific coast states. We sought to determine if this high WA1 IgG detection rate among clinical specimens merely reflects WA1 seroprevalence outside the Pacific region. WA1 IgG, as well as Babesia microti IgG, was measured in 900 blood donor specimens from 9 states. Overall seroprevalence was 2.0% for WA1 and 0.4% for B. microti; regional seroprevalences ranged from 0 to 4% and 0 to 2%, respectively. Additional studies were performed to determine if WA1 IgG reactivity was attributable to polyclonal B-cell activation associated with acute Epstein-Barr virus (EBV) infection; 40 WA1 IgG-positive clinical sera and the 18 WA1 IgG-positive blood donor specimens were all negative for EBV capsid antigen (EBVCA) IgM (a marker of acute EBV infection), and 40 EBVCA IgM-positive sera were all negative for WA1 IgG. These findings indicate that the high WA1 IgG detection rate among clinical specimens does not simply reflect the national WA1 seroprevalence among blood donors or nonspecific reactivity due to acute EBV infection. Rather, the findings suggest that infection with WA1 or a related organism is more common than indicated by the literature and is not limited to Pacific coast states. PMID:20861326

  11. Direct analysis of [6,6-(2)H2]glucose and [U-(13)C6]glucose dry blood spot enrichments by LC-MS/MS.

    PubMed

    Coelho, Margarida; Mendes, Vera M; Lima, Inês S; Martins, Fátima O; Fernandes, Ana B; Macedo, M Paula; Jones, John G; Manadas, Bruno

    2016-06-01

    A liquid chromatography tandem mass spectrometry (LC-MS/MS) using multiple reaction monitoring (MRM) in a triple-quadrupole scan mode was developed and comprehensively validated for the determination of [6,6-(2)H2]glucose and [U-(13)C6]glucose enrichments from dried blood spots (DBS) without prior derivatization. The method is demonstrated with dried blood spots obtained from rats administered with a primed-constant infusion of [U-(13)C6]glucose and an oral glucose load enriched with [6,6-(2)H2]glucose. The sensitivity is sufficient for analysis of the equivalent to <5μL of blood and the overall method was accurate and precise for the determination of DBS isotopic enrichments. PMID:27107853

  12. Development and Application of Zirconia Coated Paper Substrate for High Sensitivity Analysis of Therapeutic Drugs in Dried Blood Spots.

    PubMed

    Zheng, Yajun; Wang, Qian; Wang, Xiaoting; Chen, Ying; Wang, Xuan; Zhang, Xiaoling; Bai, Zongquan; Han, Xiaoxiao; Zhang, Zhiping

    2016-07-19

    Paper spray mass spectrometry has been demonstrated to be promising for direct analysis of therapeutic drugs in dried blood spots (DBS); however, the strong hydrogen bond and van de Waals interactions between paper substrate and analytes containing polar functional groups (e.g., therapeutic drugs) affect greatly the elution behavior and analysis sensitivity of compounds of interest during paper spray. Herein, we developed a one-sided ZrO2 coated paper substrate through a facile vacuum filtration approach using commercial ZrO2 particles as coating material and soluble starch as adhesive agent. Owing to the unique surface properties, as-prepared ZrO2 paper substrate has been shown to have excellent performance for analysis of therapeutic drugs in DBS during paper spray mass spectrometry. In contrast to original cellulose paper substrates, improvements of 43-189-fold in lower limit of quantitation (LLOQ) were obtained for the tested drugs using ZrO2 coated paper for paper spray. In comparing with the previously reported grade SG81 paper and one-sided silica coated paper, the LLOQs of the tested drugs with as-prepared ZrO2 paper decreased 1.5-16.5-fold relative to those from the above two, revealing that ZrO2 coated paper is a good candidate for paper spray in high sensitivity analysis of therapeutic drugs in DBS. PMID:27314839

  13. Measuring thyroxine and thyrotropin simultaneously in a dried blood sample on filter paper, to screen for neonatal hypothroidism

    SciTech Connect

    Nagataki, S.; Ishibashi, K.,; Ohsawa, R.

    1980-05-01

    A highly sensitive radioimmunoassay of thyroxine and thyrotropin for mass screening for neonatal hypothroidism was developed. This assay involves a single disc (3 mm diameter) of dried blood on filter paper. The minimum detectable concentrations are 15 pg/tube (10 )g/L) for thyroxine and 15 nano-int. units/tube (10 milli-int. units/L) for thyrotropin; intra- and interassay CV's are <15% in both assays. The high sensitivity of this method is due to use of labeled thyroxine with high specific activity (3 kCi/g) and of an anti-thyrotropin serum with high affinity (K/sub eq/ = 7.8 x 10 rr L/mol). With this method, 1137 newborns were screened; a follow-up study revealed that only newborns with both high thyrotropin and low thyroxine concentrations had permanent hypothyroidism. It is concluded that this method is sensitive, simple, and reliable and that the recall rate with this method is much lower than that of tests for measuring thyroxine or thyrotropin alone.

  14. THE STABILITY OF MARKERS IN DRIED-BLOOD SPOTS FOR RECOMMENDED NEWBORN SCREENING DISORDERS IN THE UNITED STATES

    PubMed Central

    Adam, BW; Hall, EM; Sternberg, M; Lim, TH; Flores, SR; O’Brien, S; Simms, D; Li, LX; De Jesus, VR; Hannon, WH

    2015-01-01

    Objective We aimed to measure separately the contributions of heat and humidity to changes in levels of 34 markers of inborn disorders in dried-blood-spot (DBS) samples. Design and Methods Paired sets of DBSs were stored at 37°C for predetermined intervals in low-humidity and high-humidity environments. Marker levels of all samples in each complete sample set were measured in a single analytic run. Results During the 30±5 day study, galactose-1-phosphate uridyltransferase and biotinidase lost almost 65% of initial activities in low-humidity storage; most of the degradation in 27 other markers was attributable to adverse effects of high humidity storage; seven markers in DBSs stored at high humidity lost more than 90% of initial levels by the end of the study and four of the seven lost more than 50% of initial levels within the first week of storage. Conclusions Minimizing both humidity and temperature in the DBS transportation and storage environments is essential to maintaining sample integrity. PMID:21963384

  15. Dry Olive Leaf Extract Counteracts L-Thyroxine-Induced Genotoxicity in Human Peripheral Blood Leukocytes In Vitro

    PubMed Central

    Žukovec Topalović, Dijana; Živković, Lada; Čabarkapa, Andrea; Djelić, Ninoslav; Bajić, Vladan; Spremo-Potparević, Biljana

    2015-01-01

    The thyroid hormones change the rate of basal metabolism, modulating the consumption of oxygen and causing production of reactive oxygen species, which leads to the development of oxidative stress and DNA strand breaks. Olive (Olea europaea L.) leaf contains many potentially bioactive compounds, making it one of the most potent natural antioxidants. The objective of this study was to evaluate the genotoxicity of L-thyroxine and to investigate antioxidative and antigenotoxic potential of the standardized oleuropein-rich dry olive leaf extract (DOLE) against hydrogen peroxide and L-thyroxine-induced DNA damage in human peripheral blood leukocytes by using the comet assay. Various concentrations of the extract were tested with both DNA damage inducers, under two different experimental conditions, pretreatment and posttreatment. Results indicate that L-thyroxine exhibited genotoxic effect and that DOLE displayed protective effect against thyroxine-induced genotoxicity. The number of cells with DNA damage, was significantly reduced, in both pretreated and posttreated samples (P < 0.05). Comparing the beneficial effect of all tested concentrations of DOLE, in both experimental protocols, it appears that extract was more effective in reducing DNA damage in the pretreatment, exhibiting protective role against L-thyroxine effect. This feature of DOLE can be explained by its capacity to act as potent free radical scavenger. PMID:25789081

  16. Dry olive leaf extract counteracts L-thyroxine-induced genotoxicity in human peripheral blood leukocytes in vitro.

    PubMed

    Topalović, Dijana Žukovec; Živković, Lada; Čabarkapa, Andrea; Djelić, Ninoslav; Bajić, Vladan; Dekanski, Dragana; Spremo-Potparević, Biljana

    2015-01-01

    The thyroid hormones change the rate of basal metabolism, modulating the consumption of oxygen and causing production of reactive oxygen species, which leads to the development of oxidative stress and DNA strand breaks. Olive (Olea europaea L.) leaf contains many potentially bioactive compounds, making it one of the most potent natural antioxidants. The objective of this study was to evaluate the genotoxicity of L-thyroxine and to investigate antioxidative and antigenotoxic potential of the standardized oleuropein-rich dry olive leaf extract (DOLE) against hydrogen peroxide and L-thyroxine-induced DNA damage in human peripheral blood leukocytes by using the comet assay. Various concentrations of the extract were tested with both DNA damage inducers, under two different experimental conditions, pretreatment and posttreatment. Results indicate that L-thyroxine exhibited genotoxic effect and that DOLE displayed protective effect against thyroxine-induced genotoxicity. The number of cells with DNA damage, was significantly reduced, in both pretreated and posttreated samples (P < 0.05). Comparing the beneficial effect of all tested concentrations of DOLE, in both experimental protocols, it appears that extract was more effective in reducing DNA damage in the pretreatment, exhibiting protective role against L-thyroxine effect. This feature of DOLE can be explained by its capacity to act as potent free radical scavenger. PMID:25789081

  17. Evaluation of nucleic acid stabilization products for ambient temperature shipping and storage of viral RNA and antibody in a dried whole blood format.

    PubMed

    Dauner, Allison L; Gilliland, Theron C; Mitra, Indrani; Pal, Subhamoy; Morrison, Amy C; Hontz, Robert D; Wu, Shuenn-Jue L

    2015-07-01

    Loss of sample integrity during specimen transport can lead to false-negative diagnostic results. In an effort to improve upon the status quo, we used dengue as a model RNA virus to evaluate the stabilization of RNA and antibodies in three commercially available sample stabilization products: Whatman FTA Micro Cards (GE Healthcare Life Sciences, Pittsburgh, PA), DNAstāble Blood tubes (Biomātrica, San Diego, CA), and ViveST tubes (ViveBio, Alpharetta, GA). Both contrived and clinical dengue-positive specimens were stored on these products at ambient temperature or 37°C for up to 1 month. Antibody and viral RNA levels were measured by enzyme-linked immunosorbent assay (ELISA) and quantitative reverse transcription polymerase chain reaction (qRT-PCR) assays, respectively, and compared with frozen unloaded controls. We observed reduced RNA and antibody levels between stabilized contrived samples and frozen controls at our earliest time point, and this was particularly pronounced for the FTA cards. However, despite some time and temperature dependent loss, a 94.6-97.3% agreement was observed between stabilized clinical specimens and their frozen controls for all products. Additional considerations such as cost, sample volume, matrix, and ease of use should inform any decision to incorporate sample stabilization products into a diagnostic testing workflow. We conclude that DNAstāble Blood and ViveST tubes are useful alternatives to traditional filter paper for ambient temperature shipment of clinical specimens for downstream molecular and serological testing. PMID:25940193

  18. Clinical Validation and Implications of Dried Blood Spot Sampling of Carbamazepine, Valproic Acid and Phenytoin in Patients with Epilepsy

    PubMed Central

    Kong, Sing Teang; Lim, Shih-Hui; Lee, Wee Beng; Kumar, Pasikanthi Kishore; Wang, Hwee Yi Stella; Ng, Yan Lam Shannon; Wong, Pei Shieen; Ho, Paul C.

    2014-01-01

    To facilitate therapeutic monitoring of antiepileptic drugs (AEDs) by healthcare professionals for patients with epilepsy (PWE), we applied a GC-MS assay to measure three AEDs: carbamazepine (CBZ), phenytoin (PHT) and valproic acid (VPA) levels concurrently in one dried blood spot (DBS), and validated the DBS-measured levels to their plasma levels. 169 PWE on either mono- or polytherapy of CBZ, PHT or/and VPA were included. One DBS, containing ∼15 µL of blood, was acquired for the simultaneous measurement of the drug levels using GC-MS. Simple Deming regressions were performed to correlate the DBS levels with the plasma levels determined by the conventional immunoturbimetric assay in clinical practice. Statistical analyses of the results were done using MedCalc Version 12.6.1.0 and SPSS 21. DBS concentrations (Cdbs) were well-correlated to the plasma concentrations (Cplasma): r = 0.8381, 0.9305 and 0.8531 for CBZ, PHT and VPA respectively, The conversion formulas from Cdbs to plasma concentrations were [0.89×CdbsCBZ+1.00]µg/mL, [1.11×CdbsPHT−1.00]µg/mL and [0.92×CdbsVPA+12.48]µg/mL respectively. Inclusion of the red blood cells (RBC)/plasma partition ratio (K) and the individual hematocrit levels in the estimation of the theoretical Cplasma from Cdbs of PHT and VPA further improved the identity between the observed and the estimated theoretical Cplasma. Bland-Altman plots indicated that the theoretical and observed Cplasma of PHT and VPA agreed well, and >93.0% of concentrations was within 95% CI (±2SD); and similar agreement (1∶1) was also found between the observed Cdbs and Cplasma of CBZ. As the Cplasma of CBZ, PHT and VPA can be accurately estimated from their Cdbs, DBS can therefore be used for drug monitoring in PWE on any of these AEDs. PMID:25255292

  19. Simultaneous measurement of imatinib, nilotinib and dasatinib in dried blood spot by ultra high performance liquid chromatography tandem mass spectrometry.

    PubMed

    Kralj, Eva; Trontelj, Jurij; Pajič, Tadej; Kristl, Albin

    2012-08-15

    Imatinib, dasatinib and nilotinib are three tyrosine kinase inhibitors currently used to treat Bcr-Abl1 positive chronic myelogenous leukaemia (CML). However, achieving maximum benefit with these drugs may require optimal dosing and adherence to therapy. In those cases, therapeutic drug monitoring (TDM) can be a useful tool in managing patients with CML. The paper presents simple and high throughput method for simultaneous determination of all three TKIs in dried blood spot (DBS) samples from CML patients. DBS samples were prepared by applying 10 μL of spiked whole blood onto an Agilent DBS cards. Whole blood spot was punched out of the card, transferred to a well in a 96-well Captiva ND Lipids filter plate. After the addition of isotopically labelled internal standard, the drug was extracted with 0.1% formic acid in methanol. The collected extract (1 μL) was injected onto a Phenomenex Kinetex 50 mm × 2.1 mm C18 column and eluted with acetonitrile gradient into a triple quadrupole ESI-MS/MS Agilent 6460 operated in positive mode. The total run time was only 2.6 min. The method was validated in terms of linearity, selectivity, specificity, accuracy, precision, absolute and relative matrix effect and stability. The effect of haematocrit (Hct) on the accurate concentration determination was also examined. The method was linear in the range of 50-5000 μg/L for imatinib and nilotinib and in the range of 2.5-250 μg/L for dasatinib, with correlation coefficient values higher than 0.997. Lower limits of quantification (LLOQ) were 50 μg/L for imatinib and nilotinib and 2.5 μg/L for dasatinib. The method proved to be accurate (% bias < 13.2) and precise (CV < 10.3%) on intra- as well as on inter-day basis. Sample matrix (% ME=94.5-106.7) and different Hct values had no significant effect on the accuracy of measured concentrations. Samples proved to be stable whilst stored on DBS cards at room temperature or in the refrigerator; however, at 40 °C the stability of

  20. Quantitative Analysis of Therapeutic Drugs in Dried Blood Spot Samples by Paper Spray Mass Spectrometry: An Avenue to Therapeutic Drug Monitoring

    NASA Astrophysics Data System (ADS)

    Manicke, Nicholas Edward; Abu-Rabie, Paul; Spooner, Neil; Ouyang, Zheng; Cooks, R. Graham

    2011-09-01

    A method is presented for the direct quantitative analysis of therapeutic drugs from dried blood spot samples by mass spectrometry. The method, paper spray mass spectrometry, generates gas phase ions directly from the blood card paper used to store dried blood samples without the need for complex sample preparation and separation; the entire time for preparation and analysis of blood samples is around 30 s. Limits of detection were investigated for a chemically diverse set of some 15 therapeutic drugs; hydrophobic and weakly basic drugs, such as sunitinib, citalopram, and verapamil, were found to be routinely detectable at approximately 1 ng/mL. Samples were prepared by addition of the drug to whole blood. Drug concentrations were measured quantitatively over several orders of magnitude, with accuracies within 10% of the expected value and relative standard deviation (RSD) of around 10% by prespotting an internal standard solution onto the paper prior to application of the blood sample. We have demonstrated that paper spray mass spectrometry can be used to quantitatively measure drug concentrations over the entire therapeutic range for a wide variety of drugs. The high quality analytical data obtained indicate that the technique may be a viable option for therapeutic drug monitoring.

  1. Quantitation of Gamma-Hydroxybutyric Acid in Dried Blood Spots: Feasibility Assessment for Newborn Screening of Succinic Semialdehyde Dehydrogenase (SSADH) Deficiency

    PubMed Central

    Forni, Sabrina; Pearl, Phillip L.; Gibson, K. Michael; Yu, Yuezhou; Sweetman, Lawrence

    2013-01-01

    Objective SSADH deficiency, the most prevalent autosomal recessive disorder of GABA degradation, is characterized by elevated gamma-hydroxybutyric acid (GHB). Neurological outcomes may be improved with early intervention and anticipatory guidance. Morbidity has been compounded by complications, e.g. hypotonia, in undiagnosed infants with otherwise routine childhood illnesses. We report pilot methodology on the feasibility of newborn screening for SSADH deficiency. Method Dried blood spot (DBS) cards from patients affected with SSADH deficiency were compared with 2831 archival DBS cards for gamma-hydroxybutyric acid content. Following extraction with methanol, GHB in DBS was separated and analyzed using ultra high-performance liquid chromatography tandem mass spectrometry. Results Methodology was validated to meet satisfactory accuracy and reproducibility criteria, including intra-day and inter-day validation. Archival refrigerated dried blood spots samples of babies, infants and children (N=2831) were screened for GHB, yielding a mean +/- S.D. of 8 ± 5 nM (99.9 %-tile 63 nM) (Min 0.0 Max 78 nM). The measured mean and median concentrations in blood spots derived from seven SSADH deficient patients were 1182 nM and 699 nM respectively (Min 124, Max 4851nM). Conclusions GHB concentration in all 2831 dried blood spot cards was well below the lowest concentration of affected children. These data provide proof-of-principle for screening methodology to detect SSADH deficiency with applicability to newborn screening and earlier diagnosis. PMID:23742746

  2. Cotinine and trans 3'-hydroxycotinine in dried blood spots as biomarkers of tobacco exposure and nicotine metabolism.

    PubMed

    Murphy, Sharon E; Wickham, Katherine M; Lindgren, Bruce R; Spector, Logan G; Joseph, Anne

    2013-01-01

    Tobacco use is the major preventable cause of premature death in the United States. Second-hand smoke (SHS) exposure also contributes to a number of premature deaths as well as other negative health outcomes. An accurate assessment of tobacco smoke exposure is critical to understanding these disease processes. The plasma concentration of cotinine, the primary metabolite of nicotine, is widely accepted as a quantitative measure of tobacco and SHS exposure. However, it is not always feasible to collect plasma. Dried blood spots (DBS), which are collected routinely from newborns and often from young children for lead screening, provide an alternative sampling method. We have developed a quantitative high throughput liquid chromatography tandem mass spectrometry method for the analysis of cotinine in DBS. The limit of quantitation was 0.3 ng/g (~ 0.2 ng/ml plasma). Cotinine levels in DBS from 83 smokers and 99 non-smokers exposed to SHS were determined. Plasma cotinine concentrations in these subjects ranged from <0.02 to 443 ng/ml. Cotinine was detected in DBS from 157 subjects, and the correlation between cotinine in plasma and DBS was excellent, 0.992 (P<0.001). We also determined the ratio of trans 3'-hydroxycotinine to cotinine, a measure of nicotine metabolism, in DBS from smokers. This ratio in DBS was well correlated with the ratio in plasma, 0.94 (P<0.001). In a small study, we confirmed the feasibility of using extant DBS collected for lead screening to assess SHS exposure in children. PMID:23443235

  3. Chemical composition of solar dried blood and the ruminal content and its effect on performance of Japanese quails

    PubMed Central

    Mishra, Jyotiprabha; Abraham, Robinson J. J; Rao, V. Appa; Rajini, R. Asha; Mishra, B. P.; Sarangi, N. R.

    2015-01-01

    Aim: The aim was to determine the chemical composition of solar dried blood and rumen content (DBRC) and further ascertain the concentration at which DBRC could be included in Japanese quail diets without any adverse effect on its performance. Materials and Methods: Feeding trial on the effect of DBRC on performance of Japanese quails was studied up to 5 weeks. 252 numbers of day old (Nandanam Type III breed) Japanese quails were purchased from Poultry Research Station, Madhavaram and divided into 7 batches (control+ six treatments) each consisting of 36 birds. The DBRC was included at 0%, 5%, 10%, 15%, 20%, 25% and 30% in diets as control, treatment-1 (T1), treatment-2 (T2), treatment-3 (T3), treatment-4 (T4), treatment-5 (T5) and treatment-6 (T6) respectively in a completely randomized design to replace soybean meal in Japanese quail feed. The birds were provided with ad-labidum feed and drinking water ad-libitum during the entire experimental period. Results: The crude protein (CP), crude fiber (CF), ether extract (EE) and ash contents of DBRC were 35.87%, 17.40%, 3.6% and 12.6%, respectively. The amount of essential amino acids and non-essential amino acid content were found to be 12.98 and 4.87 (g/100 g of feed) respectively in DBRC feed. Result showed that all birds fed DBRC diets performed better than the control group. Mortality was unaffected by dietary treatments. There was a significant difference (p<0.01) observed in weight gain in treatment groups compared to the control. Conclusion: Up to 30% DBRC could be incorporated in the diets of Japanese quails without any adverse effects on its performance. PMID:27047002

  4. Development of a Broad-Range 23S rDNA Real-Time PCR Assay for the Detection and Quantification of Pathogenic Bacteria in Human Whole Blood and Plasma Specimens

    PubMed Central

    Gaibani, Paolo; Mariconti, Mara; Bua, Gloria; Bonora, Sonia; Sassera, Davide; Landini, Maria Paola; Mulatto, Patrizia; Novati, Stefano; Bandi, Claudio; Sambri, Vittorio

    2013-01-01

    Molecular methods are important tools in the diagnosis of bloodstream bacterial infections, in particular in patients treated with antimicrobial therapy, due to their quick turn-around time. Here we describe a new broad-range real-time PCR targeting the 23S rDNA gene and capable to detect as low as 10 plasmid copies per reaction of targeted bacterial 23S rDNA gene. Two commercially available DNA extraction kits were evaluated to assess their efficiency for the extraction of plasma and whole blood samples spiked with different amount of either Staphylococcus aureus or Escherichia coli, in order to find the optimal extraction method to be used. Manual QIAmp extraction method with enzyme pre-treatment resulted the most sensitive for detection of bacterial load. Sensitivity of this novel assay ranged between 10 and 103 CFU per PCR reaction for E. coli and S. aureus in human whole blood samples depending on the extraction methods used. Analysis of plasma samples showed a 10- to 100-fold reduction of bacterial 23S rDNA in comparison to the corresponding whole blood specimens, thus indicating that whole blood is the preferential sample type to be used in this real-time PCR protocol. Our results thus show that the 23S rDNA gene represents an optimal target for bacteria quantification in human whole blood. PMID:23586027

  5. Evaluation of Different Cytomegalovirus (CMV) DNA PCR Protocols for Analysis of Dried Blood Spots from Consecutive Cases of Neonates with Congenital CMV Infections▿

    PubMed Central

    Soetens, Oriane; Vauloup-Fellous, Christelle; Foulon, Ina; Dubreuil, Pascal; De Saeger, Ben; Grangeot-Keros, Liliane; Naessens, Anne

    2008-01-01

    Two protocols for the extraction of cytomegalovirus (CMV) DNA and two methods for the amplification of CMV DNA in dried blood spots were evaluated for the retrospective diagnosis of congenital CMV infection. During the period from 1996 to 2006, a urine screening program detected 76 congenitally infected neonates. Stored Guthrie cards with blood from 55 cases and 12 controls were tested. Two spots of dried blood were cut from each card and evaluated in two centers. CMV DNA was extracted from a whole single spot. Center 1 used phenol-chloroform extraction and ethanol precipitation followed by a conventional PCR. Center 2 used the NucliSens easyMAG automated DNA/RNA extraction platform (bioMérieux) followed by a real-time PCR. For evaluation of the extraction method, DNA extracted from each blood spot was evaluated by the amplification method used by the collaborating center. The sensitivities were 66% for center 1 and 73% for center 2. None of the controls were positive. A sensitivity as high as 82% could be obtained by combining the most sensitive extraction method (the phenol-chloroform procedure) with the most sensitive PCR method (real-time PCR). The detection rate was not influenced by the duration of storage of the spots. The sensitivity was higher with blood from congenitally infected cases due to a primary maternal CMV infection, regardless of the protocol used. However, the difference reached significance only for the least-sensitive protocol (P = 0.036). PMID:18199787

  6. Automated high-performance liquid chromatographic method for the determination of guanidinoacetic acid in dried blood spots: a tool for early diagnosis of guanidinoacetate methyltransferase deficiency.

    PubMed

    Carducci, C; Birarelli, M; Santagata, P; Leuzzi, V; Carducci, C; Antonozzi, I

    2001-05-01

    A new automated method for the assay of guanidinoacetic acid (GAA) in dried blood spot (DBS) on filter paper is reported. The method, based on reversed-phase (RP)-HPLC, precolumn derivatisation with benzoin and fluorescence detection, has shown good precision and sensitivity and requires only minimal sample handling. The validity of the method was demonstrated by analysing the neonatal blood spot of a patient affected by guanidinoacetate methyltransferase (GAMT) deficiency. GAA concentration was found to be nearly 12-fold higher than the mean control value. We propose this method as an inexpensive and widely applicable tool for the diagnosis of GAMT deficiency. PMID:11393723

  7. Simultaneous quantitation of hexacosanoyl lysophosphatidylcholine, amino acids, acylcarnitines, and succinylacetone during FIA–ESI–MS/MS analysis of dried blood spot extracts for newborn screening

    PubMed Central

    Haynes, Christopher A.; De Jesús, Víctor R.

    2016-01-01

    Objectives The goal of this study was to include the quantitation of hexacosanoyl lysophosphatidylcholine, a biomarker for X-linked adrenoleukodystrophy and other peroxisomal disorders, in the routine extraction and analysis procedure used to quantitate amino acids, acylcarnitines, and succinylacetone during newborn screening. Criteria for the method included use of a single punch from a dried blood spot, one simple extraction of the punch, no high-performance liquid chromatography, and utilizing tandem mass spectrometry to quantitate the analytes. Design and methods Dried blood spot punches were extracted with a methanolic solution of stable-isotope labeled internal standards, formic acid, and hydrazine, followed by flow injection analysis–electrospray ionization–tandem mass spectrometry. Results Quantitation of amino acids, acylcarnitines, and hexacosanoyl lysophosphatidylcholine using this combined method was similar to results obtained using two separate methods. Conclusions A single dried blood spot punch extracted by a rapid (45 min), simple procedure can be analyzed with high throughput (2 min per sample) to quantitate amino acids, acylcarnitines, succinylacetone, and hexacosanoyl lysophosphatidylcholine. PMID:26432925

  8. Targeted screening for the detection of Pompe disease in patients with unclassified limb-girdle muscular dystrophy or asymptomatic hyperCKemia using dried blood: A Spanish cohort.

    PubMed

    Gutiérrez-Rivas, E; Bautista, J; Vílchez, J J; Muelas, N; Díaz-Manera, J; Illa, I; Martínez-Arroyo, A; Olivé, M; Sanz, I; Arpa, J; Fernández-Torrón, R; López de Munáin, A; Jiménez, L; Solera, J; Lukacs, Z

    2015-07-01

    We aimed to screen for Pompe disease in patients with unclassified limb-girdle muscular dystrophy (LGMD) or asymptomatic hyperCKemia using dried blood spot (DBS) assays. Subsequently, we aimed to calculate the diagnostic delay between initial symptom presentation and the diagnosis. A prospective, multicenter, observational study was conducted in 348 patients: 146 with unclassified LGMD and 202 with asymptomatic or paucisymptomatic hyperCKemia. We quantified levels of acid alpha-glucosidase (GAA) from dried blood spots analyzed fluorometrically. The test was positive in 20 patients, and Pompe disease was confirmed by genetic testing in 16. Undiagnosed Pompe disease was detected in 7.5% of patients with LGMD and in 2.5% of patients with persistent, idiopathic elevation of serum creatine kinase. The c.-32-13 T > G mutation was found most commonly. The diagnostic delay was 15 years on average. In conclusion, DBS tests are useful and reliable screening tools for Pompe disease. We recommend the dried blood spot test to be included in the diagnostic work-up of patients with unclassified myopathies with proximal weakness and/or hyperCKemia of unknown cause and, when positive, to define the diagnosis, it will have to be confirmed by biochemical and/or molecular genetic analysis. PMID:25998610

  9. The Validity of Phosphatidylethanol in Dried Blood Spots of Newborns for the Identification of Prenatal Alcohol Exposure

    PubMed Central

    Bakhireva, Ludmila N.; Leeman, Lawrence; Savich, Renate D.; Cano, Sandra; Gutierrez, Hilda; Savage, Daniel D.; Rayburn, William F.

    2014-01-01

    Background Accurate identification of prenatal alcohol exposure (PAE) in the newborn period offers an opportunity for early identification of children at risk for future neurocognitive problems and the implementation of interventional approaches earlier in life. PAE newborn screening by measuring phosphatidylethanol in dried blood spot (PEth-DBS) cards is feasible, logistically easier, and more cost-efficient compared to other biomarkers. However, the sensitivity and specificity of this method have yet to be established. Methods This prospective cohort study examined validity of PEth-DBS among 28 infants with PAE and 32 controls relative to maternal self-report and other biomarkers. Pregnant women were recruited from a University of New Mexico clinic and followed to early postpartum period. The composite index, which was based on self-reported measures of alcohol use and allowed to classify subjects into PAE and control groups, was the criterion measure used to estimate sensitivity and specificity of PEth-DBS. Results The study included large proportions of patients representing ethnic minorities (7.4% American Indian, 81.7% Hispanic/Latina), low education (54.2%

  10. Orientation of histopathology specimens.

    PubMed

    Burns, A; Adams, J; Endersby, S

    2004-02-01

    We present a simple way of orientating large specimens being sent to the laboratory for histopathological examination by supplementing the pinning of the specimen on a cork board with Polaroid photographs of the specimen and numbered tags. PMID:14706306

  11. Cinemicrographic specimen housing

    NASA Technical Reports Server (NTRS)

    Wilkins, J. R.

    1979-01-01

    Housing used to observe gravitation effects on specimens embedded in support media, such as agar, supports microbial specimens vertically for time-lapsed cinemicrographic studies. Procedure cannot be performed with conventional microscopes which see specimens in horizontal plane only.

  12. NASA Biological Specimen Repository

    NASA Technical Reports Server (NTRS)

    McMonigal, K. A.; Pietrzyk, R. A.; Sams, C. F.; Johnson, M. A.

    2010-01-01

    The NASA Biological Specimen Repository (NBSR) was established in 2006 to collect, process, preserve and distribute spaceflight-related biological specimens from long duration ISS astronauts. This repository provides unique opportunities to study longitudinal changes in human physiology spanning may missions. The NBSR collects blood and urine samples from all participating ISS crewmembers who have provided informed consent. These biological samples are collected once before flight, during flight scheduled on flight days 15, 30, 60, 120 and within 2 weeks of landing. Postflight sessions are conducted 3 and 30 days after landing. The number of in-flight sessions is dependent on the duration of the mission. Specimens are maintained under optimal storage conditions in a manner that will maximize their integrity and viability for future research The repository operates under the authority of the NASA/JSC Committee for the Protection of Human Subjects to support scientific discovery that contributes to our fundamental knowledge in the area of human physiological changes and adaptation to a microgravity environment. The NBSR will institute guidelines for the solicitation, review and sample distribution process through establishment of the NBSR Advisory Board. The Advisory Board will be composed of representatives of all participating space agencies to evaluate each request from investigators for use of the samples. This process will be consistent with ethical principles, protection of crewmember confidentiality, prevailing laws and regulations, intellectual property policies, and consent form language. Operations supporting the NBSR are scheduled to continue until the end of U.S. presence on the ISS. Sample distribution is proposed to begin with selections on investigations beginning in 2017. The availability of the NBSR will contribute to the body of knowledge about the diverse factors of spaceflight on human physiology.

  13. Use of micro samples of finger prick blood dried on filter paper for a quick and simple dipstick dot-EIA for diagnosis of amebic liver abscess (ALA).

    PubMed

    Sharma, M; Ghosh, S; Singal, A K; Anand, B S; Talwar, G P

    1994-01-01

    Filter paper was used as a support to absorb micro samples of finger prick blood for detection of antibodies to Entamoeba histolytica (causative organism of amebiasis) by a rapid dipstick dot-EIA technique; 8 microliters of blood was sufficient to saturate discs (diameter 6 mm) of Whatman paper No. 3. Conditions of elution of blood from the discs were optimized and the best results were obtained when 0.4 ml of buffer was used for elution for 30 minutes at room temperature. The filter paper technique is extremely useful for field use and for diagnosis of amebic liver abscess on a large scale since it does not involve centrifugation of blood and use of sterile vials because blood dried on paper can be stored in polythene bags at room temperature for up to 3 months and for a week at 42 degrees C without significant loss of antibody activity, thereby eliminating the need of refrigeration of samples for storage or transportation to a reference laboratory. Elution of blood from different discs of the same patient was found to be highly reproducible without appreciable loss of sensitivity or specificity. Twenty-four confirmed ALA, 29 non-ALA, and 25 apparently normal healthy controls, with no previous history of amebiasis, were tested. Sensitivity and specificity of the test was found to be 96% and 92%, respectively. PMID:8189328

  14. Urine culture - catheterized specimen

    MedlinePlus

    Culture - urine - catheterized specimen; Urine culture - catheterization; Catheterized urine specimen culture ... urinary tract infections may be found in the culture. This is called a contaminant. You may not ...

  15. Combining rapid diagnostic tests and dried blood spot assays for point-of-care testing of human immunodeficiency virus, hepatitis B and hepatitis C infections in Burkina Faso, West Africa.

    PubMed

    Kania, D; Bekalé, A M; Nagot, N; Mondain, A-M; Ottomani, L; Meda, N; Traoré, M; Ouédraogo, J B; Ducos, J; Van de Perre, P; Tuaillon, E

    2013-12-01

    People screened for human immunodeficiency virus (HIV) using rapid diagnostic tests (RDTs) in Africa remain generally unaware of their status for hepatitis B (HBV) and hepatitis C (HCV) infections. We evaluated a two-step screening strategy in Burkina Faso, using both HIV RDTs and Dried Blood Spot (DBS) assays to confirm an HIV-positive test, and to test for HBV and HCV infections. HIV counselling and point-of-care testing were performed at a voluntary counselling and testing centre with HBV, HCV status and HIV confirmation using DBS specimens, being assessed at a central laboratory. Serological testing on plasma was used as the reference standard assay to control for the performance of DBS assays. Nineteen out of 218 participants included in the study were positive for HIV using RDTs. A fourth-generation HIV ELISA and immunoblot assays on DBS confirmed HIV status. Twenty-four out of 25 participants infected with HBV were found positive for hepatitis B surface antigen (HBsAg) using DBS. One sample with a low HBsAg concentration on plasma was not detected on DBS. Five participants tested positive for HCV antibodies were confirmed positive with an immunoblot assay using DBS specimens. Laboratory results were communicated within 7 days to participants with no loss to follow up of participants between the first and second post-test counselling sessions. In conclusion, DBS collection during HIV point-of-care testing enables screening and confirmation of HBV, HCV and HIV infections. Diagnosis using DBS may assist with implementation of national programmes for HBV, HCV and HIV screening and clinical care in middle- to low-income countries. PMID:23902574

  16. Dried blood spot on-card derivatization: an alternative form of sample handling to overcome the instability of thiorphan in biological matrix.

    PubMed

    Mess, Jean-Nicholas; Taillon, Marie-Pierre; Côté, Cynthia; Garofolo, Fabio

    2012-12-01

    Thiorphan, the active metabolite of racecadotril, can undergo oxidation in biological matrices such as blood and plasma. In bioanalysis, a general approach for the stabilization of such a molecule is to derivatize the thiol group to a more stable thioether, often requiring complex handling procedures at the clinical site. In this research, the concept of dried blood spot (DBS) on-card derivatization was evaluated to stabilize thiorphan. DBS cards were in-house pre-treated with 2-bromo-3'-methoxyacetophenone and left to dry prior to blood spotting. Thiorphan was shown to be effectively derivatized to thiorphan-methoxyacetophenone once applied on the in-house pre-treated cards. Thiorphan-methoxyacetophenone was extracted by soaking a 6 mm DBS punch in methanol containing the internal standard (thiorphan-methoxyacetophenone-D₅). Chromatographic separation was achieved on a Waters XBridge C₁₈ column with a gradient elution of 5 mM NH₄HCO₃ and methanol in 2.5 min and detection by ESI(+)/MS/MS. A linear (weighted 1/x²) relationship was obtained over a concentration range of 5.00-600.00 ng/mL. The assay met regulatory guidelines acceptance criteria for sensitivity, selectivity, precision and accuracy, matrix effect, recovery, dilution integrity and multiple stability evaluations. The DBS on-card derivatization has shown to be an easy and reliable alternative form of sample collection for the quantification of thiorphan. PMID:22511292

  17. Quantification of Sulfatides in Dried Blood and Urine Spots From Metachromatic Leukodystrophy Patients by Liquid Chromatography/Electrospray Tandem Mass Spectrometry

    PubMed Central

    Barcenas, Mariana; Suhr, Teryn R.; Scott, C. Ronald; Turecek, Frantisek; Gelb, Michael H.

    2014-01-01

    Background Treatments are being developed for metachromatic leukodystrophy (MLD), suggesting the need for eventual newborn screening. Previous studies have shown that sulfatide molecular species are increased in the urine of MLD patients compared to samples from non-MLD individuals, but there is no data using dried blood spots (DBS), the most common sample available for newborn screening laboratories. Methods We used ultra-high performance liquid chromatography/tandem mass spectrometry (UHPLC/MS/MS) to quantify sulfatides in DBS and dried urine spots from 14 MLD patients and 50 non-MLD individuals. Results Several sulfatide molecular species were increased in dried urine samples from all MLD samples compared to non-MLD samples. Sulfatides, especially low molecular species, were increased in DBS from MLD patients, but the sulfatide levels were relatively low. There was good separation in sulfatide levels between MLD and non-MLD samples when dried urine spots were used, but not with DBS, because DBS from non-MLD individuals have measurable levels of sulfatides. Conclusion Sulfatide accumulation studies in urine, but not in DBS, emerges as the method of choice if newborn screening is to be proposed for MLD. PMID:24370383

  18. Determination of fatty acid ethyl esters in dried blood spots by LC-MS/MS as markers for ethanol intake: application in a drinking study.

    PubMed

    Luginbühl, Marc; Schröck, Alexandra; König, Stefan; Schürch, Stefan; Weinmann, Wolfgang

    2016-05-01

    The forensic utility of fatty acid ethyl esters (FAEEs) in dried blood spots (DBS) as short-term confirmatory markers for ethanol intake was examined. An LC-MS/MS method for the determination of FAEEs in DBS was developed and validated to investigate FAEE formation and elimination in a drinking study, whereby eight subjects ingested 0.66-0.84 g/kg alcohol to reach blood alcohol concentrations (BAC) of 0.8 g/kg. Blood was taken every 1.5-2 h, BAC was determined, and dried blood spots were prepared, with 50 μL of blood, for the determination of FAEEs. Lower limits of quantitation (LLOQ) were between 15 and 37 ng/mL for the four major FAEEs. Validation data are presented in detail. In the drinking study, ethyl palmitate and ethyl oleate proved to be the two most suitable markers for FAEE determination. Maximum FAEE concentrations were reached in samples taken 2 or 4 h after the start of drinking. The following mean peak concentrations (c̅ max) were reached: ethyl myristate 14 ± 4 ng/mL, ethyl palmitate 144 ± 35 ng/mL, ethyl oleate 125 ± 55 ng/mL, ethyl stearate 71 ± 21 ng/mL, total FAEEs 344 ± 91 ng/mL. Detectability of FAEEs was found to be on the same time scale as BAC. In liquid blood samples containing ethanol, FAEE concentrations increase post-sampling. This study shows that the use of DBS fixation prevents additional FAEE formation in blood samples containing ethanol. Positive FAEE results obtained by DBS analysis can be used as evidence for the presence of ethanol in the original blood sample. Graphical Abstract Time courses for fatty acid ethyl ester (FAEE) concentrations in DBS and ethanol concentrations for subject 1 over a period of 7 h. Ethanol ingestion occured during the first hour of the time course. PMID:26968564

  19. Viral Load Detection Using Dried Blood Spots in a Cohort of HIV-1-Infected Children in Uganda: Correlations with Clinical and Immunological Criteria for Treatment Failure

    PubMed Central

    Lundin, Rebecca; Petrara, Maria Raffaella; Penazzato, Martina; Massavon, William; Kizito, Susan; Nabachwa, Sandra Monica; Nannyonga Musoke, Maria; Namisi, Charles; Morelli, Erika; Bilardi, Davide; Mazza, Antonio; Zanchetta, Marisa; Giaquinto, Carlo; De Rossi, Anita

    2014-01-01

    Correlations between clinical/immunological treatment failure and viral load (VL) detected by dried blood spot (DBS) sampling were explored in HIV-1-infected children in Uganda. Of 104 children on combined antiretroviral treatment (cART), 12.5% experienced clinical and/or immunological failure, while 28.8%, 44.2%, and 26.9% had VLs of <1,000, 1,000 to 5,000, and >5,000 copies/ml, respectively. Clinical/immunological failure poorly predicted virological failure. PMID:24789197

  20. Viral load detection using dried blood spots in a cohort of HIV-1-infected children in Uganda: correlations with clinical and immunological criteria for treatment failure.

    PubMed

    Costenaro, Paola; Lundin, Rebecca; Petrara, Maria Raffaella; Penazzato, Martina; Massavon, William; Kizito, Susan; Nabachwa, Sandra Monica; Nannyonga Musoke, Maria; Namisi, Charles; Morelli, Erika; Bilardi, Davide; Mazza, Antonio; Zanchetta, Marisa; Giaquinto, Carlo; De Rossi, Anita

    2014-07-01

    Correlations between clinical/immunological treatment failure and viral load (VL) detected by dried blood spot (DBS) sampling were explored in HIV-1-infected children in Uganda. Of 104 children on combined antiretroviral treatment (cART), 12.5% experienced clinical and/or immunological failure, while 28.8%, 44.2%, and 26.9% had VLs of <1,000, 1,000 to 5,000, and >5,000 copies/ml, respectively. Clinical/immunological failure poorly predicted virological failure. PMID:24789197

  1. A Novel Dried Blood Spot-LCMS Method for the Quantification of Methotrexate Polyglutamates as a Potential Marker for Methotrexate Use in Children

    PubMed Central

    Hawwa, Ahmed F.; AlBawab, AbdelQader; Rooney, Madeleine; Wedderburn, Lucy R.; Beresford, Michael W.; McElnay, James C.

    2014-01-01

    Objective Development and validation of a selective and sensitive LCMS method for the determination of methotrexate polyglutamates in dried blood spots (DBS). Methods DBS samples [spiked or patient samples] were prepared by applying blood to Guthrie cards which was then dried at room temperature. The method utilised 6-mm disks punched from the DBS samples (equivalent to approximately 12 µl of whole blood). The simple treatment procedure was based on protein precipitation using perchloric acid followed by solid phase extraction using MAX cartridges. The extracted sample was chromatographed using a reversed phase system involving an Atlantis T3-C18 column (3 µm, 2.1×150 mm) preceded by Atlantis guard column of matching chemistry. Analytes were subjected to LCMS analysis using positive electrospray ionization. Key Results The method was linear over the range 5–400 nmol/L. The limits of detection and quantification were 1.6 and 5 nmol/L for individual polyglutamates and 1.5 and 4.5 nmol/L for total polyglutamates, respectively. The method has been applied successfully to the determination of DBS finger-prick samples from 47 paediatric patients and results confirmed with concentrations measured in matched RBC samples using conventional HPLC-UV technique. Conclusions and Clinical Relevance The methodology has a potential for application in a range of clinical studies (e.g. pharmacokinetic evaluations or medication adherence assessment) since it is minimally invasive and easy to perform, potentially allowing parents to take blood samples at home. The feasibility of using DBS sampling can be of major value for future clinical trials or clinical care in paediatric rheumatology. PMID:24587116

  2. NASA Biological Specimen Repository

    NASA Technical Reports Server (NTRS)

    Pietrzyk, Robert; McMonigal, K. A.; Sams, C. F.; Johnson, M. A.

    2009-01-01

    The NASA Biological Specimen Repository (NBSR) has been established to collect, process, annotate, store, and distribute specimens under the authority of the NASA/JSC Committee for the Protection of Human Subjects. The International Space Station (ISS) provides a platform to investigate the effects of microgravity on human physiology prior to lunar and exploration class missions. The NBSR is a secure controlled storage facility that is used to maintain biological specimens over extended periods of time, under well-controlled conditions, for future use in approved human spaceflight-related research protocols. The repository supports the Human Research Program, which is charged with identifying and investigating physiological changes that occur during human spaceflight, and developing and implementing effective countermeasures when necessary. The storage of crewmember samples from many different ISS flights in a single repository will be a valuable resource with which researchers can validate clinical hypotheses, study space-flight related changes, and investigate physiological markers All samples collected require written informed consent from each long duration crewmember. The NBSR collects blood and urine samples from all participating long duration ISS crewmembers. These biological samples are collected pre-flight at approximately 45 days prior to launch, during flight on flight days 15, 30, 60 120 and within 2 weeks of landing. Postflight sessions are conducted 3 and 30 days following landing. The number of inflight sessions is dependent on the duration of the mission. Operations began in 2007 and as of October 2009, 23 USOS crewmembers have completed or agreed to participate in this project. As currently planned, these human biological samples will be collected from crewmembers covering multiple ISS missions until the end of U.S. presence on the ISS or 2017. The NBSR will establish guidelines for sample distribution that are consistent with ethical principles

  3. Development and validation of a dried blood spot assay for the quantification of ribavirin using liquid chromatography coupled to mass spectrometry

    PubMed Central

    Jimmerson, Leah C.; Zheng, Jia-Hua; Bushman, Lane R.; MacBrayne, Christine E.; Anderson, Peter L.; Kiser, Jennifer J.

    2014-01-01

    Efficient, inexpensive and sensitive assays for the measurement of drugs are of interest for pharmacokinetic and pharmacodynamics (PK-PD) analysis. Dried blood spots (DBS) are a unique bioanaltyical matrix with the potential to fulfill this interest for the measurement of numerous analytes. Here we describe the development and validation of a reversed-phase high performance liquid chromatographic (LC), tandem mass spectrometry (MS/MS) assay for the determination of ribavirin (RBV) in DBS. A 3mm punch from spotted and dried whole blood was extracted in methanol utilizing isotopically labeled internal standard for LC-MS/MS analysis. Validation was performed over a range of 0.05 μg/mL to 10.0 μg/mL and the method was shown to be precise (coefficient of variation ≤ 15%) and accurate (within ±15% of control). These acceptance criteria were met for hematocrit ranges of 20-54%, for center versus edge punches and for spot volumes from 10-60 μL. RBV was stable for up to 140 days at room temperature and −20°C as well as for three freeze/thaw cycles. Correlation of RBV in DBS versus in plasma yielded r2 ≥ 0.98 demonstrating that DBS can be used as an alternative to plasma for PK-PD studies in human subjects. PMID:24291608

  4. Dietary supplementation with dried chicory root triggers changes in the blood serum proteins engaged in the clotting process and the innate immune response in growing pigs.

    PubMed

    Lepczynski, A; Herosimczyk, A; Ozgo, M; Skomial, J; Taciak, M; Barszcz, M; Berezecka, N

    2015-02-01

    The aim of the study was to characterize the systemic immune and metabolic alterations in the blood serum of growing pigs in response to a dietary supplementation with 4% of dried chicory roots. This was achieved by examining the influence of the experimental diet on serum protein changes especially these related with immunology and lipid metabolism. Serum proteins with the isoelectric point ranging from pH 3.0 to 10.0 were separated using high resolution two-dimensional electrophoresis. As a result, we found that experimental diet triggered significant changes in 37 protein spots. Of these, 14 were up-regulated, whereas 23 showed down-regulation. Of 37 significantly altered protein spots, 24 were successfully identified, representing 14 distinct gene products. Implementation of the dried chicory roots into the diet of growing pigs caused a significant down-regulation of apolipoprotein C-II complement component C6, C-reactive protein, CD14 antigen, C4b binding protein α and β chains, and fibrinogen. Piglets fed experimental diet had similar IgA, IgG and IgM concentrations, although the level of IgM tended to be lower compared to the control group. It is concluded that diet supplemented with 4% of dried chicory root may exert anti-inflammatory properties and affect lipid metabolism in growing pigs. PMID:25716964

  5. Specimen collection and preservation

    USGS Publications Warehouse

    Franson, J. Christian

    1987-01-01

    Specimens, as discussed in this handbook, have but a single purpose--to provide information leading to the diagnosis of a cause of disease or death. A specimen may be an intact carcass, various tissues removed from carcasses, or parasites. In any event, the specimen should be as fresh and undamaged as possible.

  6. Short communication: Reference limits for blood analytes in Holstein late-pregnant heifers and dry cows: Effects of parity, days relative to calving, and season.

    PubMed

    Brscic, M; Cozzi, G; Lora, I; Stefani, A L; Contiero, B; Ravarotto, L; Gottardo, F

    2015-11-01

    Reference limits for metabolic profiles in Holstein late-pregnant heifers and dry cows were determined considering the effects of parity, days relative to calving, and season. Blood samples were collected from 104 pregnant heifers and 186 dry cows (68 primiparous and 118 pluriparous) from 60 to 10 d before the expected calving date in 31 dairy farms in northeastern Italy. Sampling was performed during summer (182 samples) and the following winter (108 samples). All the animals were judged as clinically healthy at a veterinary visit before sampling. Outliers were removed from data of each blood analyte, and variables that were not normally distributed were log transformed. A mixed model was used to test the fixed effects of parity (late-pregnant heifers, primiparous or pluriparous dry cows), class of days relative to calving (60-41 d, 40-21 d, 20-10 d), season (summer or winter), and the interactions between parity and class of days relative to calving and between parity and season, with farm as random effect. Single general reference limits and 95% confidence intervals were generated for analytes that did not vary according to fixed effects. Whenever a fixed effect included in the model significantly affected a given analyte, specific reference limits and 95% confidence intervals were generated for each of its levels. Albumin, urea, triglycerides, alanine aminotransferase, aspartate aminotransferase, creatinine kinase, conjugated bilirubin, calcium, phosphorus, magnesium, potassium, chloride, zinc, copper, and iron concentrations were not influenced by any of the fixed effects. Total protein, globulins, creatinine, glucose, alkaline phosphatase, gamma glutamyltransferase, lactate dehydrogenase, and sodium plasma concentrations were affected by parity. The class of days relative to calving had a significant effect on the concentrations of total protein, globulins, fatty acids, cholesterol, total bilirubin, and sodium. Season affected plasma concentrations of

  7. Use of Dried Blood Spots for Estimating Children?s Exposures to Heavy Metals in Epidemiological Research

    EPA Science Inventory

    Background: Children’s exposures to arsenic (As), lead (Pb), mercury (Hg), and cadmium (Cd) are of particular concern in early-life. Exposures to heavy metals are traditionally measured in whole venous blood, which is costly and invasive. As an alternative we describe a met...

  8. Analysis of polychlorinated biphenyls and organochlorine pesticides in archived dried blood spots and its application to track temporal trends of environmental chemicals in newborns

    PubMed Central

    Ma, Wan-Li; Gao, Chongjing; Bell, Erin M.; Druschel, Charlotte M.; Caggana, Michele; Aldous, Kenneth M.; Buck Louis, Germaine M.; Kannan, Kurunthachalam

    2014-01-01

    Dried blood spots (DBS) collected from infants shortly after birth for the newborn screening program (NSP) in the United States are valuable resources for the assessment of exposure to environmental chemicals in newborns. The NSP was debuted as a public health program in the United States in the 1960s; and the DBS samples collected over a period of time can be used in tracking temporal trends in exposure to environmental chemicals by newborns. In this study, polychlorinated biphenyls (PCBs) and organochlorine pesticides (OCPs) were measured in DBS samples collected from newborns in Upstate New York from 1997 to 2011 by gas chromatography-high resolution mass spectrometry (GC-HRMS). Twelve PCBs and two OCPs were found in DBS samples at a detection rate above 50% (n = 51). The mean whole blood concentration of ΣPCBs (sum of 12 congeners) over the 15-year period was 1.06 ng/mL, followed by p,p′-DDE (0.421 ng/mL) and HCB (0.065 ng/mL). The measured concentrations of PCBs and p,p′-DDE in infants’ blood were comparable to those reported in cord blood, suggesting maternal/trans-placental transfer of these compounds from mothers to fetuses. The concentrations of ΣPCBs and p,p′-DDE in blood samples of infants decreased significantly between 1997 and 2001, and no significant reduction was found thereafter. This observation is consistent with the trends reported for these chemicals in other human tissues in the United States. PMID:24968082

  9. Comparability of HbA1c and lipids measured with dried blood spot versus venous samples: a systematic review and meta-analysis

    PubMed Central

    2014-01-01

    Background Levels of haemoglobin A1c (HbA1c) and blood lipids are important determinants of risk in patients with diabetes. Standard analysis methods based upon venous blood samples can be logistically challenging in resource-poor settings where much of the diabetes epidemic is occurring. Dried blood spots (DBS) provide a simple alternative method for sample collection but the comparability of data from analyses based on DBS is not well established. Methods We conducted a systematic review and meta-analysis to define the association of findings for HbA1c and blood lipids for analyses based upon standard methods compared to DBS. The Cochrane, Embase and Medline databases were searched for relevant reports and summary regression lines were estimated. Results 705 abstracts were found by the initial electronic search with 6 further reports identified by manual review of the full papers. 16 studies provided data for one or more outcomes of interest. There was a close agreement between the results for HbA1c assays based on venous and DBS samples (DBS = 0.9858venous + 0.3809), except for assays based upon affinity chromatography. Significant adjustment was required for assays of total cholesterol (DBS = 0.6807venous + 1.151) but results for triglycerides (DBS = 0.9557venous + 0.1427) were directly comparable. Conclusions For HbA1c and selected blood lipids, assays based on DBS samples are clearly associated with assays based on standard venous samples. There are, however, significant uncertainties about the nature of these associations and there is a need for standardisation of the sample collection, transportation, storage and analysis methods before the technique can be considered mainstream. This should be a research priority because better elucidation of metabolic risks in resource poor settings, where venous sampling is infeasible, will be key to addressing the global epidemic of cardiovascular diseases. PMID:25045323

  10. Rapid quantification of conjugated and unconjugated bile acids and C27 precursors in dried blood spots and small volumes of serum.

    PubMed

    Janzen, N; Sander, S; Terhardt, M; Das, A M; Sass, J O; Kraetzner, R; Rosewich, H; Rosevich, H; Peter, M; Sander, J

    2010-06-01

    The aim of the study was to develop a method for fast and reliable diagnosis of peroxisomal diseases and to facilitate differential diagnosis of cholestatic hepatopathy. For the quantification of bile acids and their conjugates as well as C(27) precursors di- and trihydroxycholestanoic acid (DHCA, THCA), in small pediatric blood samples we combined HPLC separation on a reverse-phase C18 column with ESI-MS/MS analysis in the negative ion mode. Analysis was done with good precision (CV 3,7%-11.1%) and sufficient sensitivity (LOQ: 11-91 nmol/L) without derivatization. Complete analysis of 17 free and conjugated bile acids from dried blood spots and 10 microL serum samples, respectively, was performed within 12 min. Measurement of conjugated primary bile acids plus DHCA and THCA as well as ursodeoxycholic acid was done in 4.5 min. In blood spots of healthy newborns, conjugated primary bile acids were found in the range of 0.01 to 2.01 micromol/L. Concentrations of C(27) precursors were below the detection limit in normal controls. DHCA and THCA were specifically elevated in cases of peroxysomal defects and one Zellweger patient. PMID:20093478

  11. Application of an intracellular assay for determination of tenofovir-diphosphate and emtricitabine-triphosphate from erythrocytes using dried blood spots.

    PubMed

    Zheng, Jia-Hua; Rower, Caitlin; McAllister, Kevin; Castillo-Mancilla, Jose; Klein, Brandon; Meditz, Amie; Guida, L Anthony; Kiser, Jennifer J; Bushman, Lane R; Anderson, Peter L

    2016-04-15

    This communication describes the application of an existing intracellular methodology to the quantitation of tenofovir-diphosphate (TFV-DP) and emtricitabine-triphosphate (FTC-TP) from erythrocytes using dried blood spots (DBS). Concentrations were determined from a 3mm DBS punch extracted into a 70:30 methanol:water solution (lysed cellular matrix). This extraction solution was then subjected to a previously validated analytical procedure for lysed cellular matrix. Experiments for DBS validation used replicate samples from study participants to demonstrate acceptable reproducibility with spot volumes ranging from 10-50 μL and punch location either from the edge or center of the spot. Analysis of paired DBS with purified red blood cells showed that a 3mm DBS punch contained an average of 11.9 million cells for the observed hematocrit range of the participants (35-50%). Numerous stability tests were completed showing that whole blood in an EDTA vacutainer could sit for 24h at room temperature prior to spotting, and DBS could remain at room temperature for up to five days including shipment at ambient using 2-days delivery. DBS stability in storage was acceptable up to 18 months at -20°C or -80°C and DBS could undergo 4 Freeze/Thaw cycles. The described method was applied to HIV prophylaxis studies, demonstrating powerful associations with HIV acquisition through its ability to discriminate gradients of adherence. PMID:26829517

  12. LC-MS/MS method for simultaneous determination on a dried blood spot of multiple analytes relevant for treatment monitoring in patients with tyrosinemia type I.

    PubMed

    la Marca, Giancarlo; Malvagia, Sabrina; Materazzi, Serena; Della Bona, Maria Luisa; Boenzi, Sara; Martinelli, Diego; Dionisi-Vici, Carlo

    2012-01-17

    Tyrosinemia type 1 is caused by deficiency of fumarylacetoacetate hydrolase. The enzymatic defect impairs the conversion of fumarylacetoacetate to fumarate, causing accumulation of succinylacetone which induces severe liver and kidney dysfunction along with mutagenic changes and hepatocellular carcinoma. Treatment is based on nitisinone (NTBC), an enzymatic inhibitor which suppresses succinylacetone production. NTBC, which has dramatically changed the disease course improving liver and kidney functions and reducing risk of liver cancer, causes a side effect of the increase of tyrosine levels. Treatment is therefore based on the combination of NTBC with a protein-restricted diet to prevent the potential toxicity of excessive tyrosine accumulation. Long-term therapy requires a careful monitoring in blood of NTBC levels along with other disease biomarkers, which include succinylacetone, and a selected panel of circulating aminoacids. We have developed a straightforward and fast MS/MS method for the simultaneous determination of NTBC, succinylacetone, tyrosine, phenylalanine, and methionine on a dried blood spot requiring a 2 min run. A single assay suitable for quantitative evaluation of all biochemical markers is of great advance over conventional methods, especially in pediatric patients, since it reduces laboratory costs and blood sampling, is less invasive and particularly suitable for pediatric patients, and allows easier storage and shipping. PMID:22148291

  13. Dihydropteridine reductase activity in dried blood spots: effects of aging and senile dementia of the Alzheimer type.

    PubMed Central

    Jeeps, C M; Silcox, A; Lloyd, B; Clayton, B E

    1986-01-01

    Dihydropteridine reductase (EC 1.6.99.7) (DHPR) activity was measured in blood spots from 50 neonates, 52 healthy adults aged 30-62 years, and 21 elderly controls aged 67-97 years, as well as 32 demented patients of whom 25 had senile dementia of the Alzheimer type. Enzyme activity was stable for seven days at 4 degrees C and for at least 14 days at -20 degrees C. No important difference was found between the DHPR activity of venous and capillary blood. DHPR activity was considerably lower in the healthy adult group compared with neonates and the elderly group, and there was no sex difference at any age. The erythrocyte DHPR activity of patients with senile dementia of the Alzheimer type was similar to that of elderly controls. This result differs from that previously reported for leucocytes. PMID:3950042

  14. Quantification of phosphatidylethanol 16:0/18:1, 18:1/18:1, and 16:0/16:0 in venous blood and venous and capillary dried blood spots from patients in alcohol withdrawal and control volunteers.

    PubMed

    Kummer, Natalie; Ingels, Ann-Sofie; Wille, Sarah M R; Hanak, Catherine; Verbanck, Paul; Lambert, Willy E E; Samyn, Nele; Stove, Christophe P

    2016-01-01

    Phosphatidylethanol species (PEths) are promising biomarkers of alcohol consumption. Here, we report on the set-up, validation, and application of a novel UHPLC-ESI-MS/MS method for the quantification of PEth 16:0/18:1, PEth 18:1/18:1, and PEth 16:0/16:0 in whole blood (30 μL) and in venous (V, 30 μL) or capillary (C, 3 punches (3 mm)) dried blood spots (DBS). The methods were linear from 10 (LLOQ) to 2000 ng/mL for PEth 16:0/18:1, from 10 (LLOQ) to 1940 ng/mL for PEth 18:1/18:1, and from 19 (LLOQ) to 3872 ng/mL for PEth 16:0/16:0. Extraction efficiencies were higher than 55% (RSD < 18%) and matrix effects compensated for by IS were between 77 and 125% (RSD < 10%). Accuracy, repeatability, and intermediate precision fulfilled acceptance criteria (bias and RSD below 13%). Validity of the procedure for determination of PEth 16:0/18:1 in blood was demonstrated by the successful participation in a proficiency test. The quantification of PEths in C-DBS was not significantly influenced by the hematocrit, punch localization, or spot volume. The stability of PEths in V-DBS stored at room temperature was demonstrated up to 6 months. The method was applied to authentic samples (whole blood, V-DBS, and C-DBS) from 50 inpatients in alcohol withdrawal and 50 control volunteers. Applying a cut-off value to detect inpatients at 221 ng/mL for PEth 16:0/18:1 provided no false positive results and a good sensitivity (86%). Comparison of quantitative results (Bland-Altman plot, Passing-Bablok regression, and Wilcoxon signed rank test) revealed that V-DBS and C-DBS were valid alternatives to venous blood for the detection of alcohol consumption. PMID:26597914

  15. A novel rapid genotyping technique for Collie eye anomaly: SYBR Green-based real-time polymerase chain reaction method applicable to blood and saliva specimens on Flinders Technology Associates filter paper.

    PubMed

    Chang, Hye-Sook; Mizukami, Keijiro; Yabuki, Akira; Hossain, Mohammad A; Rahman, Mohammad M; Uddin, Mohammad M; Arai, Toshiro; Yamato, Osamu

    2010-09-01

    Collie eye anomaly (CEA) is a canine inherited ocular disease that shows a wide variety of manifestations and severity of clinical lesions. Recently, a CEA-associated mutation was reported, and a DNA test that uses conventional polymerase chain reaction (PCR) has now become available. The objective of the current study was to develop a novel rapid genotyping technique by using SYBR Green-based real-time PCR for future large-scale surveys as a key part in the strategy to eradicate CEA by selective breeding. First, a SYBR Green-based real-time PCR assay for genotyping of CEA was developed and evaluated by using purified DNA samples from normal, carrier, and affected Border Collies in which genotypes had previously been determined by conventional PCR. This real-time PCR assay demonstrated appropriate amplifications in all genotypes, and the results were consistent with those of conventional PCR. Second, the availability of Flinders Technology Associates filter paper (FTA card) as DNA templates for the real-time PCR assay was evaluated by using blood and saliva specimens to determine suitability for CEA screening. DNA-containing solution prepared from a disc of blood- or saliva-spotted FTA cards was available directly as templates for the real-time PCR assay when the volume of solution was 2.5% of the PCR mixture. In conclusion, SYBR Green-based real-time PCR combined with FTA cards is a rapid genotyping technique for CEA that can markedly shorten the overall time required for genotyping as well as simplify the sample preparation. Therefore, this newly developed technique suits large-scale screening in breeding populations of Collie-related breeds. PMID:20807925

  16. OR Specimen Labeling.

    PubMed

    Zervakis Brent, Mary Ann

    2016-02-01

    Mislabeled surgical specimens jeopardize patient safety and quality care. The purpose of this project was to determine whether labeling surgical specimens with two patient identifiers would result in an 80% reduction in specimen labeling errors within six months and a 100% reduction in errors within 12 months. Our failure mode effects analysis found that the lack of two patient identifiers per label was the most unsafe step in our specimen handling process. We piloted and implemented a new process in the OR using the Plan-Do-Check-Act conceptual framework. The audit process included collecting data and making direct observations to determine the sustainability of the process change; however, the leadership team halted the direct observation audit after four months. The total number of surgical specimen labeling errors was reduced by only 60% within six months and 62% within 12 months; therefore, the goal of the project was not met. However, OR specimen labeling errors were reduced. PMID:26849982

  17. Dry Mouth

    MedlinePlus

    ... of this page please turn Javascript on. Dry Mouth What Is Dry Mouth? Dry mouth is the feeling that there is ... when a person has dry mouth. How Dry Mouth Feels Dry mouth can be uncomfortable. Some people ...

  18. Effect of Ensiled Mulberry Leaves and Sun-Dried Mulberry Fruit Pomace on Finishing Steer Growth Performance, Blood Biochemical Parameters, and Carcass Characteristics

    PubMed Central

    Zhou, Zhenming; Zhou, Bo; Ren, Liping; Meng, Qingxiang

    2014-01-01

    Fifty-one Simmental crossbred steers (357.0±16.5 kg) were used to compare a standard total mix ration (TMR) with variants on animal performance, ruminal fermentation, blood biochemical parameters, and carcass characteristics. Corn grain and cotton seed meal were partially replaced by ensiled mulberry leaves (EML) or sun-dried mulberry fruit pomace (SMFP). Experimental diets had similar amounts of crude protein (CP), acid detergent fiber (ADF), and metabolizable energy (ME). Animals were divided into three groups: control group (CONT), 8% EML group, and 6.3% SMFP group. Performance, including average daily weight gain (ADG), and dry matter intake (DMI), was measured. Blood and rumen samples were collected at the end of the experiment (16 weeks). There were no differences in final body weight (P = 0.743), ADG (P = 0.425), DMI (P = 0.642), or ADG/DMI (P = 0.236) between the groups. There were no differences (P = 0.2024) in rumen pH values; ammonia N was lower (P = 0.0076) in SMFP than in the EML and CONT groups. There were differences in the concentrations of total and individual volatile fatty acids, while no differences were determined in blood biochemical parameters (i.e., plasma glucose, urea concentrations, triglycerides, total protein, insulin, IgG, alanine transaminase, and aspartate aminotransferase, P ≥ 0.098). No differences were observed in carcass characteristics (P ≥ 0.513), tenderness (P = 0.844), adipose and lean color values (P ≥ 0.149), and chemical composition (P ≥ 0.400); however, intramuscular fat was lower in the EML and SMFP groups compared to the CONT animals (P = 0.034). In conclusion, diets supplemented with these two mulberry products in an isocaloric and isonitrogenous manner have similar effects to corn grain and cotton seed meals on steer performance, blood biochemical parameters and carcass characteristics, with the exception of ruminal VFA concentrations and lower intramuscular fat content. PMID

  19. Effect of ensiled mulberry leaves and sun-dried mulberry fruit pomace on finishing steer growth performance, blood biochemical parameters, and carcass characteristics.

    PubMed

    Zhou, Zhenming; Zhou, Bo; Ren, Liping; Meng, Qingxiang

    2014-01-01

    Fifty-one Simmental crossbred steers (357.0 ± 16.5 kg) were used to compare a standard total mix ration (TMR) with variants on animal performance, ruminal fermentation, blood biochemical parameters, and carcass characteristics. Corn grain and cotton seed meal were partially replaced by ensiled mulberry leaves (EML) or sun-dried mulberry fruit pomace (SMFP). Experimental diets had similar amounts of crude protein (CP), acid detergent fiber (ADF), and metabolizable energy (ME). Animals were divided into three groups: control group (CONT), 8% EML group, and 6.3% SMFP group. Performance, including average daily weight gain (ADG), and dry matter intake (DMI), was measured. Blood and rumen samples were collected at the end of the experiment (16 weeks). There were no differences in final body weight (P = 0.743), ADG (P = 0.425), DMI (P = 0.642), or ADG/DMI (P = 0.236) between the groups. There were no differences (P = 0.2024) in rumen pH values; ammonia N was lower (P = 0.0076) in SMFP than in the EML and CONT groups. There were differences in the concentrations of total and individual volatile fatty acids, while no differences were determined in blood biochemical parameters (i.e., plasma glucose, urea concentrations, triglycerides, total protein, insulin, IgG, alanine transaminase, and aspartate aminotransferase, P ≥ 0.098). No differences were observed in carcass characteristics (P ≥ 0.513), tenderness (P = 0.844), adipose and lean color values (P ≥ 0.149), and chemical composition (P ≥ 0.400); however, intramuscular fat was lower in the EML and SMFP groups compared to the CONT animals (P = 0.034). In conclusion, diets supplemented with these two mulberry products in an isocaloric and isonitrogenous manner have similar effects to corn grain and cotton seed meals on steer performance, blood biochemical parameters and carcass characteristics, with the exception of ruminal VFA concentrations and lower intramuscular fat content. PMID:24427304

  20. LC-MS/MS bioanalysis of loratadine (Claritin) in dried blood spot (DBS) samples collected by subjects in a clinical research study.

    PubMed

    Li, Wenkui; Doherty, John; Moench, Paul; Flarakos, Jimmy; Tse, Francis L S

    2015-03-01

    A high-performance liquid chromatography-tandem mass spectrometric (LC-MS/MS) method has been developed and validated for the quantitative analysis of loratadine, an H1 histamine antagonist, in human dried blood spot (DBS) samples following a single self-administered 10 or 20mg oral dose. The samples were produced by spotting approximately 30μl of whole blood onto PE-226 cards. Two 3-mm discs were cut from the DBS samples and extracted using aqueous methanol containing the internal standard. After transfer and drying of the resulting sample extract, the reconstituted residues were chromatographed using a Waters XSelect C18 column and isocratic elution for MS/MS detection. The possible impact due to hematocrit, volume of blood sample spotted, storage temperature, and humidity, on the accuracy of measured DBS results were investigated. The results showed that only spotted blood volume might have an impact; a small volume (10μl) tended to give a larger negative bias in the measured value than the large volume ones (≥20μl). The current method was fully validated over a dynamic range of 0.200-20.0ng/ml with correlation coefficients (r(2)) for three validation batches equal to or better than 0.990. The intra-day accuracy and precision at the LLOQ were -11.5 to 0.0% bias and 6.4 to 8.9% CV, respectively. For the other QC samples (0.600, 3.00, 10.0 and 15.0ng/ml), the precision ranged from 4.2 to 9.8% CV and from 6.3 to 8.1% CV, respectively, in the intra-day and inter-day evaluations; the accuracy ranged from -1.7 to 10.0% and 2.7 to 5.3% bias, respectively, in the intra-day and inter-day batches. Loratadine is stable in the DBS samples for at least 271 days at ambient temperature in a desiccator, for at least 24h at 60°C and under 80% relative humidity, followed by re-conditioning at ambient temperature in a desiccator. The current methodology has been applied to determine the loratadine levels in DBS samples collected by subjects in a clinical research study to

  1. Early life lead exposure causes gender-specific changes in the DNA methylation profile of DNA extracted from dried blood spots

    PubMed Central

    Sen, Arko; Heredia, Nicole; Senut, Marie-Claude; Hess, Matthew; Land, Susan; Qu, Wen; Hollacher, Kurt; Dereski, Mary O; Ruden, Douglas M

    2015-01-01

    Aims In this paper, we tested the hypothesis that early life lead (Pb) exposure associated DNA methylation (5mC) changes are dependent on the sex of the child and can serve as biomarkers for Pb exposure. Methods In this pilot study, we measured the 5mC profiles of DNA extracted from dried blood spots (DBS) in a cohort of 43 children (25 males and 18 females; ages from 3 months to 5 years) from Detroit. Result & Discussion We found that the effect of Pb-exposure on the 5-mC profiles can be separated into three subtypes: affected methylation loci which are conserved irrespective of the sex of the child (conserved); affected methylation loci unique to males (male-specific); and affected methylation loci unique to females (female-specific). PMID:26077427

  2. A subsized fatigue specimen

    NASA Technical Reports Server (NTRS)

    Jeelani, S.; Natarajan, R.; Reddy, G. R.

    1986-01-01

    A subsized fatigue specimen has been designed to overcome the difficulty of machining a full-sized specimen from cast superalloy components. A finite element analysis confirmed that the stress was maximum at the gauge section for any given set of clamping and tensile loads, and that the stresses developed due to clamping forces were negligible compared with those due to tensile or compressive loads. Fatigue data generated using subsized specimens of AISI 4130 steel, 2024-T4 aluminum alloy and 6Al-4V titanium alloy compared well with those available in the literature for full-sized specimens.

  3. Free thyroxine values in dried blood spots on filter paper in newborns are related to both gestational age and birth body weight.

    PubMed

    Pacchiarotti, A; Bartalena, L; Chiovato, L; Falcone, M; Buratti, L; Ciampi, M; Giusti, L F; Grasso, L; Fenzi, G F; Martino, E

    1988-01-01

    The results of free thyroxine (FT4) measurements in dried blood spots on filter paper in 744 euthyroid newborns (616 at term, 128 preterm), 10 newborns with congenital hypothyroidism and 4 euthyroid newborns with congenital TBG deficiency are reported. FT4 was measured by column adsorption chromatography of free hormone followed by radioimmunoassay in the eluate. FT4 values averaged 24 +/- 0.2 pmol/L (mean +/- SE) in euthyroid newborns, 23.0 +/- 0.9 pmol/L in euthyroid newborns with TBG deficiency (p = NS), and 5.7 +/- 0.4 pmol/L in hypothyroid newborns (p less than 0.001 vs both groups). Total T4 (TT4) values in newborns with TBG deficiency were not different from those in hypothyroid newborns, but were significantly lower than those in euthyroid newborns without TBG abnormalities. FT4 values were higher in full-term newborns than in preterm newborns (25.2 +/- 0.3 vs 21.2 +/- 0.5 pmol/L, p less than 0.001). In both full-term and preterm newborns FT4 values in dried blood spots increased with birth body weight (bbw), virtually plateauing when bbw was greater than 2,500 g. The cut-off values established on the basis of the bbw (8.0 and 13.1 pmol/L for a bbw of less than or equal to 2,500 g and greater than 2,500 g, respectively) showed higher specificity and predictive value of positive results than the cut-off values based on the gestational age. In any case, the sensitivity, specificity and predictive values of FT4 determinations proved to be higher than those of TT4 and TSH measurements.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3139742

  4. Extraction of single-copy nuclear DNA from forensic specimens with a variety of postmortem histories.

    PubMed

    Evison, M P; Smillie, D M; Chamberlain, A T

    1997-11-01

    Specimens of human bone, teeth and dried blood spots from 3 months to 91 years old, with a variety of postmortem histories, were used in a comparative study of recovery of single-copy nuclear DNA sequences from forensic material. Sequences of the amelogenin and HLA-DPB1 genes were chosen for their value in sexing and identification. Sequences of the mitochondrial non-coding region V were also amplified to compare the recovery of mitochondrial and single-copy nuclear DNA. A variation of the silica method for DNA extraction was refined for application to the forensic specimens in this sample. Single-copy nuclear DNA was amplified from 100% of recent postoperative bone specimens (n = 6), 80% of forensic teeth and bone specimens (n = 10), 78% of recently extracted teeth (n = 18), 78% of exhumed bone up to 91 years old (n = 37) and 69% of 15 year old bone specimens fixed in 10% formalin (n = 20). Amelogenin sexing was correct in 85% of cases (n = 74) in which the sex of the donor had been recorded. There was no correlation between the age of the specimen and the extent of DNA preservation. PMID:9397544

  5. 46 CFR 4.06-20 - Specimen collection requirements.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... provide a specimen of their urine according to 46 CFR part 16 and 49 CFR part 40. (2) Specimen collection and shipping kits used to conduct drug testing must be used according to 49 CFR part 40. ... involved in the SMI must provide a specimen of their breath, blood, or saliva to the marine employer...

  6. 46 CFR 4.06-20 - Specimen collection requirements.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... provide a specimen of their urine according to 46 CFR part 16 and 49 CFR part 40. (2) Specimen collection and shipping kits used to conduct drug testing must be used according to 49 CFR part 40. ... involved in the SMI must provide a specimen of their breath, blood, or saliva to the marine employer...

  7. 46 CFR 4.06-20 - Specimen collection requirements.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... provide a specimen of their urine according to 46 CFR part 16 and 49 CFR part 40. (2) Specimen collection and shipping kits used to conduct drug testing must be used according to 49 CFR part 40. ... involved in the SMI must provide a specimen of their breath, blood, or saliva to the marine employer...

  8. 46 CFR 4.06-20 - Specimen collection requirements.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... provide a specimen of their urine according to 46 CFR part 16 and 49 CFR part 40. (2) Specimen collection and shipping kits used to conduct drug testing must be used according to 49 CFR part 40. ... involved in the SMI must provide a specimen of their breath, blood, or saliva to the marine employer...

  9. Development and validation of an enantioselective LC-MS/MS method for the analysis of the anthelmintic drug praziquantel and its main metabolite in human plasma, blood and dried blood spots.

    PubMed

    Meister, Isabel; Leonidova, Anna; Kovač, Jana; Duthaler, Urs; Keiser, Jennifer; Huwyler, Jörg

    2016-01-25

    Praziquantel (PZQ) is the treatment of choice against various trematode and cestode infections. To study the pharmacokinetics of PZQ in patients infected with the liver fluke Opisthorchis viverrini, we developed and validated an enantioselective liquid chromatography coupled to tandem mass spectrometry method for the analysis of R - and S -PZQ and its R -trans-4-OH-PZQ metabolite in human plasma, blood and dried blood spots (DBS). The analytes were detected in the positive mode using selected reaction monitoring (R- and S-PZQ: m/z 312.2 → 202.2; R-trans -4-OH-PZQ: m/z 328.0 → 202.0). Prior to the chiral separation with a cellulose tris(3-chloro-4-methylphenylcarbamate) column, the analytes were purified from matrix contaminants and concentrated on a C-18 trapping column. The analytical range for each PZQ enantiomer was 0.01-2.5 μg/mL, and 0.1-25 μg/mL for the metabolite. The method met the requirements regarding precision (± 15%, ± 20% at the lower limit of quantification-LLOQ), intra- and inter-assay accuracy (85-115%, 80-120% at LLOQ), and linearity (R(2) ≥ 0.998). The analytes were stable in stock solutions as well as in plasma, blood and DBS. For DBS, the influences of hematocrit and blood spot size were considered as minor. Our validation results show that the method presented here is precise, accurate and selective, and can be used for pharmacokinetic studies. Moreover, the enantioselective separation was achieved with a run time of 11.5 min and a simple sample processing method. PMID:26517852

  10. Temporal trends of polybrominated diphenyl ethers (PBDEs) in the blood of newborns from New York State during 1997 through 2011: analysis of dried blood spots from the newborn screening program.

    PubMed

    Ma, Wan-Li; Yun, Sehun; Bell, Erin M; Druschel, Charlotte M; Caggana, Michele; Aldous, Kenneth M; Buck Louis, Germaine M; Kannan, Kurunthachalam

    2013-07-16

    Polybrominated diphenyl ethers (PBDEs) are ubiquitous environmental pollutants, and on a global basis, North American populations are exposed to the highest doses of PBDEs. In response to the exponential increase in human exposure to PBDEs during the late 1990s, some PBDE formulations were phased out from production in the early 2000s. The effectiveness of the phase-out of commercial penta-BDE and octa-BDE mixtures in 2004 in the U.S. on human exposure levels is not known. Dried blood spots (DBSs), collected for the newborn screening program (NSP) in the U.S., are a valuable resource for the elucidation of trends in exposure to environmental pollutants in newborns. In this study, seven PBDE congeners were determined by gas chromatography-high resolution mass spectrometry (GC-HRMS) in archived DBS samples (in total, 51 blood spot composites from 1224 newborns) collected from newborns in New York State (NYS) from 1997 to 2011. The most frequently detected PBDE congener was BDE-47, with a detection rate (DR) of 86%, followed by BDE-99 (DR: 45%) and BDE-100 (DR: 43%). The mean concentrations determined during 1997 through 2011 in the whole blood of newborns were 0.128, 0.040, and 0.012 ng/mL for BDE-47, -99, and -100, respectively. A significant correlation was found among the concentrations of three major congeners (p < 0.001). PBDE concentrations were similar during 1997 through 2002 and, thereafter, decreased significantly, which was similar to the trends observed for perfluorinated compounds (PFCs) in DBS samples. Occurrence of PBDEs in the whole blood of newborns confirms that these compounds do cross the placental barrier. PMID:23755886

  11. Performance of Roche CAP/CTM HIV-1 qualitative test version 2.0 using dried blood spots for early infant diagnosis.

    PubMed

    Gueye, Sokhna Bousso; Diop-Ndiaye, Halimatou; Diallo, Mamadou Malick; Ly, Omar; Sow-Ndoye, Aissatou; Diagne-Gueye, Ndèye Diabou; Kébé-Fall, Khady; Diop, Fatou; Gaye-Diallo, Aïssatou; Belec, Laurent; Mboup, Souleymane; Touré-Kane, Coumba

    2016-03-01

    In the context of early infant diagnosis (EID) decentralization in sub-Saharan Africa, dried blood spot (DBS) is now widely used for HIV proviral DNA detection in resource-limited settings. A new version of CAP/CTM (version 2) has been introduced, recently by Roche Diagnosis as a new real-time PCR assay to replace previous technologies on qualitative detection of HIV-1 DNA using whole blood and DBS samples. The objective of this study was to evaluate CAP/CTM version 2 compared to CAP/CTM version 1 and Amplicor on DBS. A total of 261 DBS were collected from children aged 4 weeks to 17 months born from HIV-seropositive mothers and tested by the three techniques. CAP/CTM version 2 showed 100% of agreement with Amplicor including 74 positive results and 187 negative results. CAP/CTM version 2 versus CAP/CTM version 1 as well as CAP/CTM version 1 versus Amplicor showed two discordant results giving a sensitivity of 98.6%, specificity of 99.5%, positive predictive value of 98.6% and negative predictive value of 99.5%. The concordance was 99.12% (95% of confidence interval) giving a Kappa coefficient of 0.97 (p<0.001). These findings confirmed the expected good performance of CAP/CTM version 2 for HIV-1 EID. PMID:26706730

  12. Reliability of DNA methylation measures from dried blood spots and mononuclear cells using the HumanMethylation450k BeadArray

    PubMed Central

    Dugué, Pierre-Antoine; English, Dallas R.; MacInnis, Robert J.; Jung, Chol-Hee; Bassett, Julie K.; FitzGerald, Liesel M.; Wong, Ee Ming; Joo, Jihoon E.; Hopper, John L.; Southey, Melissa C.; Giles, Graham G.; Milne, Roger L.

    2016-01-01

    The reliability of methylation measures from the widely used HumanMethylation450 (HM450K) microarray has not been assessed for DNA from dried blood spots (DBS) or peripheral blood mononuclear cells (PBMC), nor for combined data from different studies. Repeated HM450K methylation measures in DNA from DBS and PBMC samples were available from participants in six case-control studies nested within the Melbourne Collaborative Cohort Study. Reliability was assessed for individual CpGs by calculating the intraclass correlation coefficient (ICC) based on technical replicates (samples repeated in a single study; 126 PBMC, 136 DBS) and study duplicates (samples repeated across studies; 280 PBMC, 769 DBS) using mixed-effects models. Reliability based on technical replicates was moderate for PBMC (median ICC = 0.42), but lower for DBS (median ICC = 0.20). Study duplicates gave lower ICCs than technical replicates. CpGs that were either highly methylated or unmethylated generally had lower ICCs, which appeared to be mostly related to their lower variability. The ICCs for global methylation measures were high, typically greater than 0.70. The reliability of methylation measures determined by the HM450K microarray is wide-ranging and depends primarily on the variability in methylation at individual CpG sites. The power of association studies is low for a substantial proportion of CpGs in the HM450K assay. PMID:27457678

  13. Comparison of active dry yeast (Saccharomyces cerevisiae) and yeast culture for growth performance, carcass traits, meat quality and blood indexes in finishing bulls.

    PubMed

    Geng, Chun-Yin; Ren, Li-Ping; Zhou, Zhen-Ming; Chang, Ying; Meng, Qing-Xiang

    2016-08-01

    This study was conducted to compare the effect of active dry yeasts (ADY) and yeast cultures (YC), two typical products of yeast preparations, on growth performance, carcass characteristics, meat quality and blood indexes in finishing bulls fed a high-concentrate diet. Forty-five finishing bulls (mean body weight (BW) ± standard deviation: 505 ± 29 kg BW) were allocated to three groups of 15 bulls and assigned randomly to one of three diets which were CON diet (basal diet), ADY diet (basal diet + Levucell SC) and YC diet (basal diet + Diamond V XP), respectively. After 98 days of trial, all bulls were slaughtered. The result showed that ADY rather than YC improved growth performance and carcass traits of bulls compared to CON. Moreover, both ADY and YC improved beef tenderness and changed blood indexes related to fat metabolism. In conclusion, ADY had more pronounced effect on growth performance of bulls fed high-concentrate diet, and both ADY and YC improved the beef quality by intensive fat metabolism. PMID:26472702

  14. 37 CFR 2.59 - Filing substitute specimen(s).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2010-07-01 2010-07-01 false Filing substitute specimen(s..., DEPARTMENT OF COMMERCE RULES OF PRACTICE IN TRADEMARK CASES Drawing § 2.59 Filing substitute specimen(s). (a... specimen(s), the applicant must: (1) For an amendment to allege use under § 2.76, verify by affidavit...

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

    PubMed

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

    2014-01-01

    Recently, research into alternative sample materials, such as oral fluid or filter-dried blood has been intensified, in order to facilitate cost-effective and animal-friendly sampling of individuals or groups of pigs for diagnostic purposes. The objective of this study was to compare the sensitivity of porcine reproductive and respiratory syndrome virus (PRRSV)-RNA detection by reverse transcription quantitative real-time PCR (RT-qPCR) in serum, FTA filter-dried blood and oral fluid sampled from individual pigs. Ten PRRSV negative pigs were injected with an EU-type PRRSV live vaccine. Blood and oral fluid samples were taken from each pig before, and 4, 7, 14 and 21 days after vaccination. All samples were then analyzed by PRRSV RT-qPCR. In serum, eight often pigs tested RT-qPCR positive at different time points post infection. Absolute quantification showed low serum PRRSV-RNA loads in most samples. In comparison to serum, sensitivity of PRRSV-RNA detection was strongly reduced in matched FTA filter-dried blood and in oral fluid from the same pigs. These results indicate that with low PRRSV-RNA loads the diagnostic sensitivity of PRRSV-RNA detection by RT-qPCR achieved with serum is currently unmatched by either FTA filter-dried blood or oral fluid. PMID:24881272

  16. The use of dried blood spots for quantification of 15 antipsychotics and 7 metabolites with ultra-high performance liquid chromatography - tandem mass spectrometry.

    PubMed

    Patteet, Lisbeth; Maudens, Kristof E; Stove, Christophe P; Lambert, Willy E; Morrens, Manuel; Sabbe, Bernard; Neels, Hugo

    2015-06-01

    Therapeutic drug monitoring of antipsychotics is important in optimizing individual therapy. In psychiatric populations, classical venous blood sampling is experienced as frightening. Interest in alternative techniques, like dried blood spots (DBS), has consequently increased. A fast and easy to perform DBS method for quantification of 16 antipsychotics (amisulpride, aripiprazole, asenapine, bromperidol, clozapine, haloperidol, iloperidone, levosulpiride, lurasidone, olanzapine, paliperidone, pipamperone, quetiapine, risperidone, sertindole and zuclopenthixol) and 8 metabolites was developed. DBS were prepared using 25 μL of whole blood and extraction of complete spots was performed using methanol: methyl-t-butyl-ether (4:1). After evaporation, the extract was reconstituted in the mobile phase and 10 μL were injected on an ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Separation using a C18 column and gradient elution with a flow rate of 0.5 mL/min resulted in a 6-min run-time. Ionization was performed in positive mode and a dynamic MRM method was applied. Median recovery was 66.4 % (range 28.7-84.5%). Accuracy was within the acceptance criteria, except for pipamperone (LLOQ and low concentration) and lurasidone (low concentration). Imprecision was only aberrant for lurasidone at low and medium concentration. All compounds were stable during 1 month at room temperature, 4 °C and -18 °C. Lurasidone was unstable when the extract was stored for 12 h on the autosampler. Absolute matrix effects (ME) (median 66.1%) were compensated by the use of deuterated IS (median 98.8%). The DBS method was successfully applied on 25-μL capillary DBS from patients and proved to be a reliable alternative for quantification of all antipsychotics except for olanzapine and N-desmethylolanzapine. PMID:25132670

  17. Comparison of the quantification of acetaminophen in plasma, cerebrospinal fluid and dried blood spots using high-performance liquid chromatography-tandem mass spectrometry

    PubMed Central

    Taylor, Rachel R.; Hoffman, Keith L.; Schniedewind, Björn; Clavijo, Claudia; Galinkin, Jeffrey L.; Christians, Uwe

    2013-01-01

    Acetaminophen (paracetamol, N-(4-hydroxyphenyl) acetamide) is one of the most commonly prescribed drugs for the management of pain in children. Quantification of acetaminophen in pre-term and term neonates and small children requires the availability of highly sensitive assays in small volume blood samples. We developed and validated an LC-MS/MS assay for the quantification of acetaminophen in human plasma, cerebro-spinal fluid (CSF) and dried blood spots (DBS). Reconstitution in water (DBS only) and addition of a protein precipitation solution containing the deuterated internal standard were the only manual steps. Extracted samples were analyzed on a Kinetex 2.6 μm PFP column using an acetonitrile/formic acid gradient. The analytes were detected in the positive multiple reaction mode. Alternatively, DBS were automatically processed using direct desorption in a sample card and preparation (SCAP) robotic autosampler in combination with online extraction. The range of reliable response in plasma and CSF was 3.05-20,000 ng/ml (r2 > 0.99) and 27.4-20,000 ng/ml (r2 > 0.99) for DBS (manual extraction and automated direct desorption). Inter-day accuracy was always within 85-115% and inter-day precision for plasma, CSF and manually extracted DBS were less than 15%. Deming regression analysis comparing 167 matching pairs of plasma and DBS samples showed a correlation coefficient of 0.98. Bland Altman analysis indicated a 26.6% positive bias in DBS, most likely reflecting the blood: plasma distribution ratio of acetaminophen. DBS are a valid matrix for acetaminophen pharmacokinetic studies. PMID:23670126

  18. Personalized monitoring of therapeutic salicylic acid in dried blood spots using a three-layer setup and desorption electrospray ionization mass spectrometry.

    PubMed

    Siebenhaar, Markus; Küllmer, Kai; Fernandes, Nuno Miguel de Barros; Hüllen, Volker; Hopf, Carsten

    2015-09-01

    Desorption electrospray ionization (DESI) mass spectrometry is an emerging technology for direct therapeutic drug monitoring in dried blood spots (DBS). Current DBS methods require manual application of small molecules as internal standards for absolute drug quantification. With industrial standardization in mind, we superseded the manual addition of standard and built a three-layer setup for robust quantification of salicylic acid directly from DBS. We combined a dioctyl sodium sulfosuccinate weave facilitating sample spreading with a cellulose layer for addition of isotope-labeled salicylic acid as internal standard and a filter paper for analysis of the standard-containing sample by DESI-MS. Using this setup, we developed a quantification method for salicylic acid from whole blood with a validated linear curve range from 10 to 2000 mg/L, a relative standard deviation (RSD%) ≤14%, and determination coefficients of 0.997. The limit of detection (LOD) was 8 mg/L and the lower limit of quantification (LLOQ) was 10 mg/L. Recovery rates in method verification by LC-MS/MS were 97 to 101% for blinded samples. Most importantly, a study in healthy volunteers after administration of a single dose of Aspirin provides evidence to suggest that the three-layer setup may enable individual pharmacokinetic and endpoint testing following blood collection by finger pricking by patients at home. Taken together, our data suggests that DBS-based quantification of drugs by DESI-MS on pre-manufactured three-layer cartridges may be a promising approach for future near-patient therapeutic drug monitoring. PMID:26168972

  19. Plastinated Knee Specimens: A Novel Educational Tool

    PubMed Central

    Neha; Lalwani, Sanjeev; Dhingra, Renu

    2013-01-01

    Introduction: During the routine dissection of knee joints in an anatomy dissection hall, it was observed that the specimens had deteriorated overtime, due to their repeated handling and the use of high concentrations of formalin for their fixation. In order to stop their further deterioration, we decided to plastinate these specimens. Thus, the present study was undertaken to prepare plastinated knee specimens from old embalmed cadavers and to compare them with freshly fixed, plastinated specimens. Objectives: 1. To plastinate old embalmed and fresh formalin fixed knee regions. 2. To demonstrate the extra and the intracapsular structures around both the plastinated knee regions. 3. To compare their morphological features in terms of their colours, dilatation and flexibility. Methods: A total of 15 knee joint specimens from among fresh formalin (5-8%) fixed (group I) and old embalmed bodies (group II) were collected, washed and plastinated by using the standard S-10 silicon technique and they were compared for their colours, dilatation and flexibility. Results: All the plastinated specimens showed an accurate reproduction of the tissue details that were comparable to those of the natural unfixed specimens. A comparison among the two groups showed a significant difference in terms of the colour, dilatation and the flexibility of the specimens. The plastinated knee joint specimens from group I were of good quality, with negligible shrinkage, more flexibility and well preserved morphologies. Conclusion: Plastinated knee specimens can serve as an excellent educational tool for the undergraduate and postgraduate students of anatomy, radiology and orthopaedics, as they are dry, odourless and nontoxic, with a good structural preservation and a higher instructional value. The fresh knee regions, when they were plastinated, were aesthetically superior in terms of their colours, dilatation and flexibility, thus making them ideal for teaching and hands-on experiences. PMID

  20. Multiplex Assay of Second-Line Anti-Tuberculosis Drugs in Dried Blood Spots Using Ultra-Performance Liquid Chromatography-Tandem Mass Spectrometry

    PubMed Central

    Lee, Kyunghoon; Jun, Sun-Hee; Han, Minje; Song, Sang Hoon; Park, Jong Sun; Lee, Jae Ho; Park, Kyoung Un

    2016-01-01

    As dried blood spots (DBSs) have various advantages over conventional venous blood sampling, some assays for detection of one or two anti-tuberculosis (TB) drugs in DBSs have been developed. However, there are no assays currently available for the simultaneous measurement of three or more anti-TB drugs in DBSs. In this study, we developed and evaluated a multiplex method for detecting nine anti-TB drugs including streptomycin, kanamycin, clarithromycin, cycloserine, moxifloxacin, levofloxacin, para-aminosalicylic acid, prothionamide, and linezolid in DBSs by using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Seventy-nine patient samples of DBS were analyzed on the UPLC-MS/MS system. All drug concentrations were determined within 4 min, and assay performance was evaluated. All drugs were clearly separated without ion suppression. Within-run and between-run precisions were 1.7-13.0% and 5.7-17.0%, respectively, at concentrations representing low and high levels for the nine drugs. Lower limits of detection and quantification were 0.06-0.6 and 0.5-5.0 µg/mL, respectively. Linearity was acceptable at five level concentrations for each drug. Correlations between drug concentrations in plasma and DBSs by using Passing-Bablock regression and Pearson's rho (ρ, 0.798-0.989) were acceptable. In conclusion, we developed a multiplex assay to measure nine second-line anti-TB drugs in DBSs successfully. This assay provided convenient and rapid drug quantification and could have applications in drug monitoring during treatment. PMID:27374716

  1. Multiplex Assay of Second-Line Anti-Tuberculosis Drugs in Dried Blood Spots Using Ultra-Performance Liquid Chromatography-Tandem Mass Spectrometry.

    PubMed

    Lee, Kyunghoon; Jun, Sun Hee; Han, Minje; Song, Sang Hoon; Park, Jong Sun; Lee, Jae Ho; Park, Kyoung Un; Song, Junghan

    2016-09-01

    As dried blood spots (DBSs) have various advantages over conventional venous blood sampling, some assays for detection of one or two anti-tuberculosis (TB) drugs in DBSs have been developed. However, there are no assays currently available for the simultaneous measurement of three or more anti-TB drugs in DBSs. In this study, we developed and evaluated a multiplex method for detecting nine anti-TB drugs including streptomycin, kanamycin, clarithromycin, cycloserine, moxifloxacin, levofloxacin, para-aminosalicylic acid, prothionamide, and linezolid in DBSs by using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Seventy-nine patient samples of DBS were analyzed on the UPLC-MS/MS system. All drug concentrations were determined within 4 min, and assay performance was evaluated. All drugs were clearly separated without ion suppression. Within-run and between-run precisions were 1.7-13.0% and 5.7-17.0%, respectively, at concentrations representing low and high levels for the nine drugs. Lower limits of detection and quantification were 0.06-0.6 and 0.5-5.0 μg/mL, respectively. Linearity was acceptable at five level concentrations for each drug. Correlations between drug concentrations in plasma and DBSs by using Passing-Bablock regression and Pearson's rho (ρ 0.798-0.989) were acceptable. In conclusion, we developed a multiplex assay to measure nine second-line anti-TB drugs in DBSs successfully. This assay provided convenient and rapid drug quantification and could have applications in drug monitoring during treatment. PMID:27374716

  2. Validation and Application of a Dried Blood Spot Assay for Biofilm-Active Antibiotics Commonly Used for Treatment of Prosthetic Implant Infections.

    PubMed

    Knippenberg, Ben; Page-Sharp, Madhu; Salman, Sam; Clark, Ben; Dyer, John; Batty, Kevin T; Davis, Timothy M E; Manning, Laurens

    2016-08-01

    Dried blood spot (DBS) antibiotic assays can facilitate pharmacokinetic (PK)/pharmacodynamic (PD) studies in situations where venous blood sampling is logistically difficult. We sought to develop, validate, and apply a DBS assay for rifampin (RIF), fusidic acid (FUS), and ciprofloxacin (CIP). These antibiotics are considered active against organisms in biofilms and are therefore commonly used for the treatment of infections associated with prosthetic implants. A liquid chromatography-mass spectroscopy DBS assay was developed and validated, including red cell partitioning and thermal stability for each drug and the rifampin metabolite desacetyl rifampin (Des-RIF). Plasma and DBS concentrations in 10 healthy adults were compared, and the concentration-time profiles were incorporated into population PK models. The limits of quantification for RIF, Des-RIF, CIP, and FUS in DBS were 15 μg/liter, 14 μg/liter, 25 μg/liter, and 153 μg/liter, respectively. Adjusting for hematocrit, red cell partitioning, and relative recovery, DBS-predicted plasma concentrations were comparable to measured plasma concentrations for each antibiotic (r > 0.95; P < 0.0001), and Bland-Altman plots showed no significant bias. The final population PK estimates of clearance, volume of distribution, and time above threshold MICs for measured and DBS-predicted plasma concentrations were comparable. These drugs were stable in DBSs for at least 10 days at room temperature and 1 month at 4°C. The present DBS antibiotic assays are robust and can be used as surrogates for plasma concentrations to provide valid PK and PK/PD data in a variety of clinical situations, including therapeutic drug monitoring or studies of implant infections. PMID:27270283

  3. Application of dried blood spots to determine vitamin D status in a large nutritional study with unsupervised sampling: the Food4Me project.

    PubMed

    Hoeller, Ulrich; Baur, Manuela; Roos, Franz F; Brennan, Lorraine; Daniel, Hannelore; Fallaize, Rosalind; Forster, Hannah; Gibney, Eileen R; Gibney, Mike; Godlewska, Magdalena; Hartwig, Kai; Kolossa, Silvia; Lambrinou, Christina P; Livingstone, Katherine M; Lovegrove, Julie A; Macready, Anna L; Manios, Yannis; Marsaux, Cyril F M; Martinez, J Alfredo; Celis-Morales, Carlos; Moschonis, George; Navas-Carretero, Santiago; O'Donovan, Clare B; San-Cristobal, Rodrigo; Saris, Wim H M; Surwiłło, Agnieszka; Traczyk, Iwona; Tsirigoti, Lydia; Walsh, Marianne C; Woolhead, Clara; Mathers, John C; Weber, Peter

    2016-01-28

    An efficient and robust method to measure vitamin D (25-hydroxy vitamin D3 (25(OH)D3) and 25-hydroxy vitamin D2 in dried blood spots (DBS) has been developed and applied in the pan-European multi-centre, internet-based, personalised nutrition intervention study Food4Me. The method includes calibration with blood containing endogenous 25(OH)D3, spotted as DBS and corrected for haematocrit content. The methodology was validated following international standards. The performance characteristics did not reach those of the current gold standard liquid chromatography-MS/MS in plasma for all parameters, but were found to be very suitable for status-level determination under field conditions. DBS sample quality was very high, and 3778 measurements of 25(OH)D3 were obtained from 1465 participants. The study centre and the season within the study centre were very good predictors of 25(OH)D3 levels (P<0·001 for each case). Seasonal effects were modelled by fitting a sine function with a minimum 25(OH)D3 level on 20 January and a maximum on 21 July. The seasonal amplitude varied from centre to centre. The largest difference between winter and summer levels was found in Germany and the smallest in Poland. The model was cross-validated to determine the consistency of the predictions and the performance of the DBS method. The Pearson's correlation between the measured values and the predicted values was r 0·65, and the sd of their differences was 21·2 nmol/l. This includes the analytical variation and the biological variation within subjects. Overall, DBS obtained by unsupervised sampling of the participants at home was a viable methodology for obtaining vitamin D status information in a large nutritional study. PMID:26548417

  4. Pharmacokinetic Study of Praziquantel Enantiomers and Its Main Metabolite R-trans-4-OH-PZQ in Plasma, Blood and Dried Blood Spots in Opisthorchis viverrini-Infected Patients

    PubMed Central

    Meister, Isabel; Kovac, Jana; Duthaler, Urs; Odermatt, Peter; Huwyler, Jörg; Vanobberghen, Fiona; Sayasone, Somphou; Keiser, Jennifer

    2016-01-01

    Background Praziquantel (PZQ) is the treatment of choice for infections with the liver fluke Opisthorchis viverrini, a major health problem in Southeast Asia. However, pharmacokinetic (PK) studies investigating the disposition of PZQ enantiomers (R- and S-PZQ) and its main metabolite, R-trans-4-OH-PZQ, in diseased patients are lacking. The implementation of a dried blood spot (DBS) sampling technique would ease the performance of PK studies in remote areas without clinical facilities. The aim of the present study is to provide data on the disposition of PZQ enantiomers and R-trans-4-OH-PZQ in opisthorchiasis patients and to validate the use of DBS compared to plasma and blood sampling. Methodology/Principal Findings PZQ was administered to nine O. viverrini-infected patients at 3 oral doses of 25 mg/kg in 4 h intervals. Plasma, blood and DBS were simultaneously collected at selected time points from 0 to 24 h post-treatment. PK parameters were determined using non-compartmental analysis. Drug concentrations and areas under the curve (AUC0–24h) measured in the 3 matrices were compared using Bland-Altman analysis. We observed plasma AUC0–24hs of 1.1, 9.0 and 188.7 μg/ml*h and half-lives of 1.1, 3.3 and 6.4 h for R-PZQ, S-PZQ and R-trans-4-OH, respectively. Maximal plasma concentrations (Cmax) of 0.2, 0.9 and 13.9 μg/ml for R-PZQ, S-PQZ and R-trans-4-OH peaked at 7 h for PZQ enantiomers and at 8.7 h for the metabolite. Individual drug concentration measurements and patient AUC0–24hs displayed ratios of blood or DBS versus plasma between 79–94% for R- and S-PZQ, and between 108–122% for R-trans-4-OH. Conclusions/Significance Pharmacodynamic (PD) in vitro studies on PZQ enantiomers and R-trans-4-OH-PZQ are necessary to be able to correlate PK parameters with efficacy. DBS appears to be a valid alternative to conventional venous sampling for PK studies in PZQ-treated patients. PMID:27152952

  5. Controlled environment specimen transfer.

    PubMed

    Damsgaard, Christian D; Zandbergen, Henny; W Hansen, Thomas; Chorkendorff, Ib; B Wagner, Jakob

    2014-08-01

    Specimen transfer under controlled environment conditions, such as temperature, pressure, and gas composition, is necessary to conduct successive complementary in situ characterization of materials sensitive to ambient conditions. The in situ transfer concept is introduced by linking an environmental transmission electron microscope to an in situ X-ray diffractometer through a dedicated transmission electron microscope specimen transfer holder, capable of sealing the specimen in a gaseous environment at elevated temperatures. Two catalyst material systems have been investigated; Cu/ZnO/Al2O3 catalyst for methanol synthesis and a Co/Al2O3 catalyst for Fischer-Tropsch synthesis. Both systems are sensitive to ambient atmosphere as they will oxidize after relatively short air exposure. The Cu/ZnO/Al2O3 catalyst, was reduced in the in situ X-ray diffractometer set-up, and subsequently, successfully transferred in a reactive environment to the environmental transmission electron microscope where further analysis on the local scale were conducted. The Co/Al2O3 catalyst was reduced in the environmental microscope and successfully kept reduced outside the microscope in a reactive environment. The in situ transfer holder facilitates complimentary in situ experiments of the same specimen without changing the specimen state during transfer. PMID:24824787

  6. A simple method for the analysis by MS/MS of underivatized amino acids on dry blood spots from newborn screening.

    PubMed

    Wang, Chunyan; Zhang, Wenyan; Song, Fengrui; Liu, Zhiqiang; Liu, Shuying

    2012-05-01

    The analysis by electrospray-ionization tandem mass spectrometry of amino acids with butyl esterification and isotopically labeled internal standard is routine in newborn screening laboratories worldwide. In the present study, we established a direct analysis method of higher accuracy that uses a non-deuterated internal standard. The automatic sampler and the pump of an LC apparatus were used to inject sample and mobile phase to MS, but no LC column was needed. The dry blood spot (DBS) material was prepared at levels of low, medium and high concentration; the running time was 1 min. In parallel to the new procedure, we applied the established method to analyze nine amino acids on DBS of healthy newborns and phenylketonuria newborns. The newly proposed method of product ion confirmation scan along with multiple reaction monitoring resulted in a very accurate identification of each amino acid. Our innovative protocol had high sensitivity and specificity in the analysis of cases of suspected metabolic diseases. PMID:21509487

  7. Validation of biomarkers of CVD risk from dried blood spots in community-based research: Methodologies and study-specific serum equivalencies

    PubMed Central

    Samuelsson, Laura B.; Hall, Martica H.; McLean, Shakir; Porter, James H.; Berkman, Lisa; Marino, Miguel; Sembajwe, Grace; McDade, Thomas W.; Buxton, Orfeu M.

    2016-01-01

    Objective Dried blood spot (DBS) methodology offers significant advantages over venipuncture in vulnerable populations or large-scale studies, including reduced participant burden and higher response rates. Uncertainty about validity of cardiovascular risk biomarkers remains a barrier to wide-scale use. We determined the validity of DBS-derived biomarkers of CVD risk versus gold-standard assessments, and study-specific, serum-equivalency values for clinical relevance of DBS-derived values. Methods Concurrent venipuncture serum and DBS samples (n=150 adults) were assayed in CLIA-certified and DBS laboratories, respectively. Time controls of DBS standard samples were assayed single-blind along with test samples. Linear regression analyses evaluated DBS-to-serum equivalency values; agreement and bias were assessed via Bland-Altman plots. Results Linear regressions of venipuncture values on DBS-to-serum equivalencies provided R2 values for TC, HDL-C, CRP of 0.484, 0.118, 0.666, respectively. Bland-Altman plots revealed minimal systematic bias between DBS-to-serum and venipuncture values; precision worsened at higher mean values of CRP. Time controls reveal little degradation or change in analyte values for HDL-C and CRP over 30 weeks. Conclusions DBS-assessed biomarkers represent a valid alternative to venipuncture assessments. Large studies using DBS should include study-specific serum-equivalency determinations to optimize individual-level sensitivity, viability of detecting intervention effects, and generalizability in community-level, primary prevention interventions. PMID:26652683

  8. Preparation of Chitosan and Water-Soluble Chitosan Microspheres via Spray-Drying Method to Lower Blood Lipids in Rats Fed with High-Fat Diets

    PubMed Central

    Tao, Yi; Zhang, Hong-Liang; Hu, Yin-Ming; Wan, Shuo; Su, Zheng-Quan

    2013-01-01

    This experiment aimed to investigate the effects of the chitosan (CTS) and water-soluble chitosan (WSC) microspheres on plasma lipids in male Sprague-Dawley rats fed with high-fat diets. CTS microspheres and WSC microspheres were prepared by the spray-drying technique. Scanning electron microscopy (SEM) micrographs showed that the microspheres were nearly spherical in shape. The mean size of CTS microspheres was 4.07 μm (varying from 1.50 to 7.21 μm) and of WSC microspheres was 2.00 μm (varying from 0.85 to 3.58 μm). The rats were classified into eight groups (n = 8) and were fed with high-fat diets for two weeks to establish the hyperlipidemic condition and were then treated with CTS microspheres and WSC microspheres, CTS and WSC for four weeks. The results showed that CTS and WSC microspheres reduced blood lipids and plasma viscosity and increased the serum superoxide dismutase (SOD) levels significantly. This study is the first report of the lipid-lowering effects of CTS and WSC microspheres. CTS and WSC microspheres were found to be more effective in improving hyperlipidemia in rats than common CTS and WSC. PMID:23429200

  9. Profiling of acylcarnitines and sterols from dried blood or plasma spot by atmospheric pressure thermal desorption chemical ionization (APTDCI) tandem mass spectrometry.

    PubMed

    Corso, Gaetano; D'Apolito, Oceania; Garofalo, Daniela; Paglia, Giuseppe; Dello Russo, Antonio

    2011-11-01

    Free carnitine and acylcarnitines play an important role in the metabolism of fatty acids. Sterols are structural lipids found in the membranes of many eukaryotic cells, and they also have functional roles such as the regulation of membrane permeability and fluidity, activity of membrane-bound enzymes and signals transduction. Abnormal profiles of these compounds in biological fluids may be useful markers of metabolic changes. In this review, we describe the subset of the lipidome represented by acylcarnitines and sterols, and we summarize how these compounds have been analyzed in the past. Over the last 50years, lipid mass spectrometry (MS) has evolved to become one of the most useful techniques for metabolic analysis. Today, the introduction of new ambient ionization techniques coupled to MS (AMS), which are characterized by the direct desorbing/ionizing of molecules from solid samples, is generating new possibilities for in situ analysis. Recently, we developed an AMS approach called APTDCI to desorb/ionize using a heated gas flow and an electrical discharge to directly analyze sterols and indirectly investigate acylcarnitines in dried blood or plasma spot samples. Here, we also describe the APTDCI method and some of its clinical applications, and we underline the common complications and issues that remain to be resolved. PMID:21683155

  10. Preparation of Chitosan and Water-Soluble Chitosan Microspheres via Spray-Drying Method to Lower Blood Lipids in Rats Fed with High-Fat Diets.

    PubMed

    Tao, Yi; Zhang, Hong-Liang; Hu, Yin-Ming; Wan, Shuo; Su, Zheng-Quan

    2013-01-01

    This experiment aimed to investigate the effects of the chitosan (CTS) and water-soluble chitosan (WSC) microspheres on plasma lipids in male Sprague-Dawley rats fed with high-fat diets. CTS microspheres and WSC microspheres were prepared by the spray-drying technique. Scanning electron microscopy (SEM) micrographs showed that the microspheres were nearly spherical in shape. The mean size of CTS microspheres was 4.07 μm (varying from 1.50 to 7.21 μm) and of WSC microspheres was 2.00 μm (varying from 0.85 to 3.58 μm). The rats were classified into eight groups (n = 8) and were fed with high-fat diets for two weeks to establish the hyperlipidemic condition and were then treated with CTS microspheres and WSC microspheres, CTS and WSC for four weeks. The results showed that CTS and WSC microspheres reduced blood lipids and plasma viscosity and increased the serum superoxide dismutase (SOD) levels significantly. This study is the first report of the lipid-lowering effects of CTS and WSC microspheres. CTS and WSC microspheres were found to be more effective in improving hyperlipidemia in rats than common CTS and WSC. PMID:23429200

  11. Development of a One-Step Probe Based Molecular Assay for Rapid Immunodiagnosis of Infection with M. tuberculosis Using Dried Blood Spots

    PubMed Central

    Blauenfeldt, Thomas; Heyckendorf, Jan; Graff Jensen, Sidse; Lange, Christoph; Drabe, Camilla; Hermansen, Thomas S.; de Thurah, Lena; Lillebaek, Troels; Eugen-Olsen, Jesper; Seersholm, Niels; Hoff, Søren; Bonde, Jesper; Ruhwald, Morten

    2014-01-01

    Background Antigen specific release of IP-10 is the most promising alternative marker to IFN-γ for infection with M. tuberculosis. Compared to Interferon-γ release assays (IGRA), IP-10 is released in high levels enabling novel approaches such as field friendly dried blood spots (DBS) and molecular detection. Aim To develop a robust IP-10 based molecular assay for the diagnosis of infection with M. tubercuolsis from whole blood and DBS. Method We developed a one-step probe based multiplex RT-qPCR assay for detecting IP-10 and IFN-γ mRNA expression from whole blood and DBS samples. The assay was validated and applied for the diagnosis of M. tuberculosis infection in DBS samples from 43 patients with confirmed TB, 13 patients with latent TB and 96 presumed uninfected controls. In parallel, IP-10 and INF-γ levels were measured in Quantiferon (QFT-TB) plasma supernatants. Results IP-10 mRNA upregulation was detectable at 4 hours after stimulation (6 fold upregulation) peaking at 8 hours (108 fold upregulation). IFN-γ expression occurred in concert but levels were lower (peak 6.7 fold upregulation). IP-10 gene expression level was significantly higher in patients with tuberculosis (median 31.2, IQR 10.7–67.0) and persons with latent tuberculosis infection (LTBI) (41.2, IQR 9.8–64.9) compared to healthy controls (1.6, IQR 1.1–2.4; p<0.0001). The IP-10 mRNA and protein based tests had comparable diagnostic accuracy to QFT-TB, sensitivity (85% and 88% vs 85%) and specificity (96% and 96% vs 97%, p = ns.). Conclusion We developed a rapid, robust and accurate molecular immunodiagnostic test for M. tuberculosis infection. By combining DBS based sample acquisition, mail or currier based sample transport with centralized molecular detection, this immunodiagnostic test concept can reduce the local technological requirements everywhere and make it possible to offer highly accurate immunodiagnostic tests in low resource settings. PMID:25184553

  12. Genomics and museum specimens.

    PubMed

    Nachman, Michael W

    2013-12-01

    Nearly 25 years ago, Allan Wilson and colleagues isolated DNA sequences from museum specimens of kangaroo rats (Dipodomys panamintinus) and compared these sequences with those from freshly collected animals (Thomas et al. 1990). The museum specimens had been collected up to 78 years earlier, so the two samples provided a direct temporal comparison of patterns of genetic variation. This was not the first time DNA sequences had been isolated from preserved material, but it was the first time it had been carried out with a population sample. Population geneticists often try to make inferences about the influence of historical processes such as selection, drift, mutation and migration on patterns of genetic variation in the present. The work of Wilson and colleagues was important in part because it suggested a way in which population geneticists could actually study genetic change in natural populations through time, much the same way that experimentalists can do with artificial populations in the laboratory. Indeed, the work of Thomas et al. (1990) spawned dozens of studies in which museum specimens were used to compare historical and present-day genetic diversity (reviewed in Wandeler et al. 2007). All of these studies, however, were limited by the same fundamental problem: old DNA is degraded into short fragments. As a consequence, these studies mostly involved PCR amplification of short templates, usually short stretches of mitochondrial DNA or microsatellites. In this issue, Bi et al. (2013) report a breakthrough that should open the door to studies of genomic variation in museum specimens. They used target enrichment (exon capture) and next-generation (Illumina) sequencing to compare patterns of genetic variation in historic and present-day population samples of alpine chipmunks (Tamias alpinus) (Fig. 1). The historic samples came from specimens collected in 1915, so the temporal span of this comparison is nearly 100 years. PMID:24138088

  13. 37 CFR 2.59 - Filing substitute specimen(s).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2013-07-01 2013-07-01 false Filing substitute specimen(s). 2.59 Section 2.59 Patents, Trademarks, and Copyrights UNITED STATES PATENT AND TRADEMARK OFFICE, DEPARTMENT OF COMMERCE RULES OF PRACTICE IN TRADEMARK CASES Drawing § 2.59 Filing substitute specimen(s)....

  14. 37 CFR 2.59 - Filing substitute specimen(s).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2011-07-01 2011-07-01 false Filing substitute specimen(s). 2.59 Section 2.59 Patents, Trademarks, and Copyrights UNITED STATES PATENT AND TRADEMARK OFFICE, DEPARTMENT OF COMMERCE RULES OF PRACTICE IN TRADEMARK CASES Drawing § 2.59 Filing substitute specimen(s)....

  15. 37 CFR 2.59 - Filing substitute specimen(s).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2012-07-01 2012-07-01 false Filing substitute specimen(s). 2.59 Section 2.59 Patents, Trademarks, and Copyrights UNITED STATES PATENT AND TRADEMARK OFFICE, DEPARTMENT OF COMMERCE RULES OF PRACTICE IN TRADEMARK CASES Drawing § 2.59 Filing substitute specimen(s)....

  16. Selective screening for lysosomal storage diseases with dried blood spots collected on filter paper in 4,700 high-risk colombian subjects.

    PubMed

    Uribe, Alfredo; Giugliani, Roberto

    2013-01-01

    Lysosomal storage disorders (LSDs) are a very heterogeneous group of hereditary disorders. The diagnostic process usually involves complex sampling, processing, testing, and validation procedures, performed by specialized laboratories only, which causes great limitations in reaching a diagnosis for patients affected by these diseases.There are few studies about LSDs in Colombia. The diagnostic limitations often make medical practitioners disregard the possibility of these disorders while diagnosing their patients. The current study documents the results of a 7-year screening in high-risk patients, aimed to detect LSDs using dried blood spots (DBS) collected on filter paper, with a micromethodology that facilitates diagnosis even with a large number of samples.The activities of α-galactosidase A, α glucosidase, α-L-iduronidase, arylsulfatase B, β-galactosidase, β-glucosidase, total hexosaminidase, iduronate sulfatase, and chitotriosidase were analyzed in high-risk patients for lysosomal disease. The catalytic activity was evaluated with fluorometric micromethods using artificial substrates marked with 4-methylumbelliferone.The reference values for a control population were established for the enzymes listed above, and 242 patients were found to have an enzyme deficiency, guiding to the following diagnoses: Fabry disease (n = 31), Pompe disease (n = 16), Hurler Syndrome (n = 15), Maroteaux-Lamy Syndrome (n = 34), GM1 Gangliosidosis (n = 10), Morquio B (n = 1), Gaucher disease (n = 101), Sandhoff disease (n = 1), Mucolipidosis (n = 2), and Hunter Syndrome (n = 31). In conclusion, this protocol provides a comprehensive diagnostic approach which could be carried out in Colombia and made it available to medical services spread around the country, enabling the identification of a large number of patients affected by LSDs, which could potentially benefit from the therapeutic tools already available for many of these diseases. PMID:23609959

  17. Evaluation of Mutual Drug-Drug Interaction within Geneva Cocktail for Cytochrome P450 Phenotyping using Innovative Dried Blood Sampling Method.

    PubMed

    Bosilkovska, Marija; Samer, Caroline; Déglon, Julien; Thomas, Aurélien; Walder, Bernhard; Desmeules, Jules; Daali, Youssef

    2016-09-01

    Cytochrome P450 (CYP) activity can be assessed using a 'cocktail' phenotyping approach. Recently, we have developed a cocktail (Geneva cocktail) which combines the use of low-dose probes with a low-invasiveness dried blood spots (DBS) sampling technique and a single analytical method for the phenotyping of six major CYP isoforms. We have previously demonstrated that modulation of CYP activity after pre-treatment with CYP inhibitors/inducer could be reliably predicted using Geneva cocktail. To further validate this cocktail, in this study, we have verified whether probe drugs contained in the latter cause mutual drug-drug interactions. In a randomized, four-way, Latin-square crossover study, 30 healthy volunteers received low-dose caffeine, flurbiprofen, omeprazole, dextromethorphan and midazolam (a previously validated combination with no mutual drug-drug interactions); fexofenadine alone; bupropion alone; or all seven drugs simultaneously (Geneva cocktail). Pharmacokinetic profiles of the probe drugs and their metabolites were determined in DBS samples using both conventional micropipette sampling and new microfluidic device allowing for self-sampling. The 90% confidence intervals for the geometric mean ratios of AUC metabolite/AUC probe for CYP probes administered alone or within Geneva cocktail fell within the 0.8-1.25 bioequivalence range indicating the absence of pharmacokinetic interaction. The same result was observed for the chosen phenotyping indices, that is metabolic ratios at 2 hr (CYP1A2, CYP3A) or 3 hr (CYP2B6, CYP2C9, CYP2C19, CYP2D6) post-cocktail administration. DBS sampling could successfully be performed using a new microfluidic device. In conclusion, Geneva cocktail combined with an innovative DBS sampling device can be used routinely as a test for simultaneous CYP phenotyping. PMID:27009433

  18. Validation of Biomarkers of CVD Risk from Dried Blood Spots in Community-Based Research: Methodologies and Study-Specific Serum Equivalencies.

    PubMed

    Samuelsson, Laura B; Hall, Martica H; McLean, Shakir; Porter, James H; Berkman, Lisa; Marino, Miguel; Sembajwe, Grace; McDade, Thomas W; Buxton, Orfeu M

    2015-01-01

    Dried blood spot (DBS) methodology offers significant advantages over venipuncture in studies of vulnerable populations or large-scale studies, including reduced participant burden and higher response rates. Uncertainty about the validity of cardiovascular disease (CVD) risk biomarkers remains a barrier to wide-scale use. We determined the validity of DBS-derived biomarkers of CVD risk versus gold-standard assessments, and study-specific, serum-equivalency values for clinical relevance of DBS-derived values. Concurrent venipuncture serum and DBS samples (n = 150 adults) were assayed in Clinical Laboratory Improvement Amendments-certified and DBS laboratories, respectively. Time controls of DBS standard samples were assayed single-blind along with test samples. Linear regression analyses evaluated DBS-to-serum equivalency values; agreement and bias were assessed via Bland-Altman plots. Linear regressions of venipuncture values on DBS-to-serum equivalencies provided R(2) values for total cholesterol, high-density lipoprotein cholesterol (HDL-C), and C-reactive protein (CRP) of 0.484, 0.118, and 0.666, respectively. Bland-Altman plots revealed minimal systematic bias between DBS-to-serum and venipuncture values; precision worsened at higher mean values of CRP. Time controls revealed little degradation or change in analyte values for HDL-C and CRP over 30 weeks. We concluded that DBS-assessed biomarkers represent a valid alternative to venipuncture assessments. Large studies using DBS should include study-specific serum-equivalency determinations to optimize individual-level sensitivity, the viability of detecting intervention effects, and generalizability in community-level primary prevention interventions. PMID:26652683

  19. Development and Evaluation of an Affordable Real-Time Qualitative Assay for Determining HIV-1 Virological Failure in Plasma and Dried Blood Spots

    PubMed Central

    Kliphuis, Aletta; Bronze, Michelle; Wallis, Carole L.; Kityo, Cissy; Balinda, Sheila; Stevens, Wendy; Spieker, Nicole; de Oliveira, Tulio; Rinke de Wit, Tobias F.; Schuurman, Rob

    2013-01-01

    Virological failure (VF) has been identified as the earliest, most predictive determinant of HIV-1 antiretroviral treatment (ART) failure. Due to the high cost and complexity of virological monitoring, VF assays are rarely performed in resource-limited settings (RLS). Rather, ART failure is determined by clinical monitoring and to a large extent immunological monitoring. This paper describes the development and evaluation of a low-cost, dried blood spot (DBS)-compatible qualitative assay to determine VF, in accordance with current WHO guideline recommendations for therapy switching in RLS. The assay described here is an internally controlled qualitative real-time PCR targeting the conserved long terminal repeat domain of HIV-1. This assay was applied to HIV-1 subtypes A to H and further evaluated on HIV-1 clinical plasma samples from South Africa (n = 191) and Tanzania (n = 42). Field evaluation was performed in Uganda using local clinical plasma samples (n = 176). Furthermore, assay performance was evaluated for DBS. This assay is able to identify VF for all major HIV-1 group M subtypes with equal specificity and has a lower detection limit of 1.00E+03 copies/ml for plasma samples and 5.00E+03 copies/ml for DBS. Comparative testing yielded accurate VF determination for therapy switching in 89% to 96% of samples compared to gold standards. The assay is robust and flexible, allowing for “open platform” applications and producing results comparable to those of commercial assays. Assay design enables application in laboratories that can accommodate real-time PCR equipment, allowing decentralization of testing to some extent. Compatibility with DBS extends access of sampling and thus access to this test to remote settings. PMID:23596235

  20. Application of a Liquid Extraction Based Sealing Surface Sampling Probe for Mass Spectrometric Analysis of Dried Blood Spots and Mouse Whole-Body Thin Tissue Sections

    SciTech Connect

    Van Berkel, Gary J; Kertesz, Vilmos

    2009-01-01

    The utility of a liquid extraction based sealing surface sampling probe (SSSP) for the direct mass spectrometric analysis of targeted drugs and metabolites in dried blood spots (DBSs) and whole mouse thin tissue sections was demonstrated. The accuracy and precision for the quantitative analysis of a minimum of 50 ng/mL sitamaquine or acetaminophen in DBSs on paper were well within the required 15% dictated by internationally recognized acceptance criteria for assay validations. Analysis of whole-body mouse thin tissue sections from animals dosed with propranolol, adhered to an adhesive tape substrate, provided semi-quantitative information for propranolol and its hydroxyproranolol glucuronide metabolite within specific organs of the tissue. The relative abundances recorded for the two compounds in the brain, lung, kidney and liver were in nominal agreement with previously reported amounts based on analysis using a liquid microjunction surface sampling probe (LMJ-SSP), and whole-body autoradiography (WBA) and HPLC-MS analysis. The ability to sample and analyze from tape-adhered tissue samples, which are generally employed in WBA analysis, presents the possibility of consecutive WBA and SSSP-MS analysis of the same tissue section. This would facilitate assignment, and possibly quantitation, of the different molecular forms of total drug related material detected in the WBA analysis. The flexibility to sample larger or smaller spot sizes, alternative probe sealing mechanisms, and a reduction in internal volumes and associated sample carryover issues will be among the first simple improvements necessary to make the SSSP-MS method a practical DBS and/or thin tissue section analysis tool or to expand its use to other surface sampling applications.

  1. A stepped wedge cluster randomized control trial of dried blood spot testing to improve the uptake of hepatitis C antibody testing within UK prisons

    PubMed Central

    Whitaker, Rhiannon; Perrett, Stephanie; Zou, Lu; Hickman, Matthew; Lyons, Marion

    2015-01-01

    Background: The prevalence of hepatitis C (HCV) is elevated within prison populations, yet diagnosis in prisons remains low. Dried blood spot testing (DBST) is a simple procedure for the detection of HCV antibodies; its impact on testing in the prison context is unknown. Methods: We carried out a stepped-wedge cluster-randomized control trial of DBST for HCV among prisoners within five male prisons and one female prison. Each prison was a separate cluster. The order in which the intervention (training in use of DBST for HCV testing and logistic support) was introduced was randomized across clusters. The outcome measure was the HCV testing rate by prison. Imputation analysis was carried out to account for missing data. Planned and actual intervention times differed in some prisons; data were thus analysed by intention to treat (ITT) and by observed step times. Results: There was insufficient evidence of an effect of the intervention on testing rate using either the ITT intervention time (OR: 0.84; 95% CI: 0.68–1.03; P = 0.088) or using the actual intervention time (OR: 0.86; 95% CI: 0.71–1.06; P = 0.153). This was confirmed by the pooled results of five imputed data sets. Conclusions: DBST as a stand-alone intervention was insufficient to increase HCV diagnosis within the UK prison setting. Factors such as staff training and allocation of staff time for regular clinics are key to improving service delivery. We demonstrate that prisons can conduct rigorous studies of new interventions, but data collection can be problematic. Trial registration: International Standard Randomized Controlled Trial Number Register (ISRCTN number ISRCTN05628482). PMID:25061233

  2. Dried Blood Spot Testing for Seven Steroids Using Liquid Chromatography-Tandem Mass Spectrometry With Reference Interval Determination in the Korean Population

    PubMed Central

    Kim, Borahm; Lee, Mi-Na; Park, Hyung-Doo; Kim, Jong Won; Chang, Yun Sil; Park, Won Soon

    2015-01-01

    Background Conventional screening for congenital adrenal hyperplasia (CAH) using immunoassays generates a large number of false-positive results. A more specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has been introduced to minimize unnecessary follow-ups. However, because of limited data on its use in the Korean population, LC-MS/MS has not yet been incorporated into newborn screening programs in this region. The present study aims to develop and validate an LC-MS/MS method for the simultaneous determination of seven steroids in dried blood spots (DBS) for CAH screening, and to define age-specific reference intervals in the Korean population. Methods We developed and validated an LC-MS/MS method to determine the reference intervals of cortisol, 17-hydroxyprogesterone, 11-deoxycortisol, 21-deoxycortisol, androstenedione, corticosterone, and 11-deoxycorticosterone simultaneously in 453 DBS samples. The samples were from Korean subjects stratified by age group (78 full-term neonates, 76 premature neonates, 89 children, and 100 adults). Results The accuracy, precision, matrix effects, and extraction recovery were satisfactory for all the steroids at three concentrations; values of intra- and inter-day precision coefficients of variance, bias, and recovery were 0.7-7.7%, -1.5-9.8%, and 49.3-97.5%, respectively. The linearity range was 1-100 ng/mL for cortisol and 0.5-50 ng/mL for other steroids (R2>0.99). The reference intervals were in agreement with the previous reports. Conclusions This LC-MS/MS method and the reference intervals validated in the Korean population can be successfully applied to analyze seven steroids in DBS for the diagnosis of CAH. PMID:26354345

  3. Reference intervals of α-glycosidase, β-glycosidase, and α-galactosidase in dried blood spot in a Turkish newborn population.

    PubMed

    Aldemir, Ozan; Ergun, Pelin; Güneş, Sezgin; Köroğlu, Ozge Altun; Yalaz, Mehmet; Kültürsay, Nilgün; Coker, Mahmut; Sözmen, Eser Y

    2013-09-01

    Inherited lysosomal storage diseases (LSDs) are rare, and diagnosis is often delayed for 7-10 years. Since the therapies have become available for a limited number of LSDs, (Fabry, Gaucher, Pompe, and MPS-1), early diagnosis of treatable LSDs can be lifesaving or ameliorating and allows timely treatment before irreversible damage occurs. Recently, the use of dried blood spot test (DBS) for newborn screening of LSDs has been proposed for newborn screening tests. They are noninvasive, sensitive, and specific assays with the further advantage of a fast turnaround time compared to measurement in leukocyte and/or fibroblast culture. We aimed to determine the reference intervals for lysosomal enzyme activities of newborn babies in our population and to investigate the effect of gestational week on enzyme activity. One hundred thirty healthy newborn babies (70 girls, 60 boys) were included into the study. α-Glycosidase, β-glycosidase, and α-galactosidase activities in DBS samples of newborns were determined fluorometrically. Reference intervals were calculated using Dixon's rule and percentiles of 2.5-97.5. Cutoff limits (5 %) for α-glycosidase, β-glycosidase, and α-galactosidase activities were 0.57, 0.92, and 2.18, respectively. α-Galactosidase activity was higher in girls compared to boys (p < 0.05). Interestingly, α-glycosidase and β-glycosidase activities of newborns who were delivered before 38 weeks were significantly lower than those who were delivered at 39-40 weeks. Conclusion It is of utmost importance to define the reference intervals for lysosomal enzyme activities as well as cutoff limits for newborn babies with regard to gestational age and sex. More studies to clarify the reason for the change in enzyme activity by gestational week will be required. PMID:23661235

  4. Incident Infection and Resistance Mutation Analysis of Dried Blood Spots Collected in a Field Study of HIV Risk Groups, 2007-2010

    PubMed Central

    Wei, Xierong; Smith, Amanda J.; Forrest, David W.; Cardenas, Gabriel A.; Beck, Dano W.; LaLota, Marlene; Metsch, Lisa R.; Sionean, Catlainn; Owen, S. Michele; Johnson, Jeffrey A.

    2016-01-01

    Objective To assess the utility of cost-effective dried blood spot (DBS) field sampling for incidence and drug resistance surveillance of persons at high risk for HIV infection. Methods We evaluated DBS collected in 2007–2010 in non-clinical settings by finger-stick from HIV-positive heterosexuals at increased risk of HIV infection (n = 124), men who have sex with men (MSM, n = 110), and persons who inject drugs (PWID, n = 58). Relative proportions of recent-infection findings among risk groups were assessed at avidity index (AI) cutoffs of ≤25%, ≤30%, and ≤35%, corresponding to an infection mean duration of recency (MDR) of 220.6, 250.4, and 278.3 days, respectively. Drug resistance mutation prevalence was compared among the risk groups and avidity indices. Results HIV antibody avidity testing of all self-reported ARV-naïve persons (n = 186) resulted in 9.7%, 11.3% and 14.0% with findings within the 221, 250, and 278-day MDRs, respectively. The proportion of ARV-naïve MSM, heterosexuals, and PWID reporting only one risk category who had findings below the suggested 30% AI was 23.1%, 6.9% and 3.6% (p<0.001), respectively. MSM had the highest prevalence of drug resistance and the only cases of transmitted multi-class resistance. Among the ARV-experienced, MSM had disproportionately more recent-infection results than did heterosexuals and PWID. Conclusions The disproportionately higher recent-infection findings for MSM as compared to PWID and heterosexuals increased as the MDR window increased. Unreported ARV use might explain greater recent-infection findings and drug resistance in this MSM population. DBS demonstrated utility in expanded HIV testing; however, optimal field handling is key to accurate recent-infection estimates. PMID:27415433

  5. AB108. The appliance of Bio-Plex immunoassay using dried blood spots for mucopolysaccharidosis IVA newborn screening in Taiwan—a pilot study

    PubMed Central

    Lin, Chia-Hui; Chuang, Chih-Kuang; Lin, Hsiang-Yu; Wang, Tuen-Jen; Tsai, Chia-Chen; Lin, Shuan-Pei

    2015-01-01

    Background Mucopolysaccharidosis (MPS) IVA is an autosomal recessive lysosomal storage disorder caused by the deficiency of N-acetylgalactosamine-6-sulfatase (GALNS) resulting in excessive lysosomal storage of keratan sulfate. This excessive storage causes a systemic skeletal dysplasia, short stature, and joint abnormalities. Treatments for MPS IVA are available. Better outcomes are associated with early treatment, which highlights a need for newborn screening for MPS IVA. Methods We have conducted a newborn screening pilot program for MPS IVA since December 1, 2013. Screening involved measuring the quantity of GALNS in dried blood spots on filter paper (DBFP) from newborns using a Bio-Plex immunoassay. The amounts of fluorescence sorting detected by YAG laser with wavelengths of 532 (exciting) and 580 nm (emission) is proportional to the quantity of GALNS protein. Results More than 5,657 neonates have been analyzed, in those, 132 newborns had GALNS quantification less than the cut-off value (48.64 ρg/mL) at the first screening test. Most of them (n=124) were exclusive and only eight had been recalled for a second DBFP collection and GALNS quantity rechecked. The reference values were 48.64-552.4 ρg/mL. For the confirmed MPS IV patients without enzyme replacement therapy (n=11), the GALNS quantities were far less than 5% of the normal population, and ranged from 0.00 to 4.02 ρg/mL. The GALNS quantities of the carriers (n=2) were significantly reduced comparing with those of the normal values. Conclusions The Bio-Plex immunoassay has the potential to be adopted for newborn screening of MPS IVA. This method is reliable, sensitive, validated, simple, and cost-effective in measuring GALNS enzyme in DBFP.

  6. Anaerobic specimen transport device.

    PubMed Central

    Wilkins, T D; Jimenez-Ulate, F

    1975-01-01

    A device is described and evaluated for the anaerobic transport of clinical specimens. The device limits the amount of oxygen entering with the sample to a maximum of 2%, which is rapidly removed by reacting with hydrogen in the presence of a palladium catalyst. The viability on swabs of 12 species of anaerobes, four strains of facultative anaerobes and a strain of Pseudomonas aeruginosa, was maintained during the length of the tests (24 or 48 h). The results demonstrated that this device protected even the more oxygen-sensitive clinical anaerobes from death due to oxygen exposure. This device can be used for swabs as well as for anaerobic collection and liquid and solid specimens. Images PMID:1104656

  7. Biaxial Creep Specimen Fabrication

    SciTech Connect

    JL Bump; RF Luther

    2006-02-09

    This report documents the results of the weld development and abbreviated weld qualification efforts performed by Pacific Northwest National Laboratory (PNNL) for refractory metal and superalloy biaxial creep specimens. Biaxial creep specimens were to be assembled, electron beam welded, laser-seal welded, and pressurized at PNNL for both in-pile (JOYO reactor, O-arai, Japan) and out-of-pile creep testing. The objective of this test campaign was to evaluate the creep behavior of primary cladding and structural alloys under consideration for the Prometheus space reactor. PNNL successfully developed electron beam weld parameters for six of these materials prior to the termination of the Naval Reactors program effort to deliver a space reactor for Project Prometheus. These materials were FS-85, ASTAR-811C, T-111, Alloy 617, Haynes 230, and Nirnonic PE16. Early termination of the NR space program precluded the development of laser welding parameters for post-pressurization seal weldments.

  8. Maternal Screening for Hypothyroidism and Thyroiditis Using Filter Paper Specimens

    PubMed Central

    Foley, T.P.; Henry, J.J.; Hofman, L.F.; Sanfilippo, J.S.; Naylor, E.W.

    2013-01-01

    Abstract Background and Objective Hypothyroidism and autoimmune thyroiditis are more prevalent than previously considered in women during pregnancy and the postpartum, and are associated with adverse effects on the mother and her fetus. We determined the efficacy and accuracy of screening women for primary hypothyroidism and autoimmune thyroiditis by testing TSH and two thyroid antibodies (TAb): thyroperoxidase antibodies (TPOAb) and thyroglobulin antibodies (TgAb), in eluates of filter paper specimens collected during early pregnancy and the postpartum. Methods We enrolled 494 first-trimester pregnant women with no exclusion criteria into a prospective study to detect primary hypothyroidism and autoimmune thyroiditis. Finger stick blood was applied to filter paper, dried in room air, eluted, and promptly tested for TSH and TAb. A total of 178 of the pregnant women (36%) were tested in the early postpartum. Women with abnormal results had confirmatory serum tests. Results It was found that 91 pregnant women (18.4%) and 43 postpartum women (24.2%) had abnormal TSH values (>4.0 mU/L) and/or positive TAb; 140 pregnant women (28.3%) had TSH values >2.5 mU/L. All subjects with TSH values >4.0 mU/L tested positive for TAb. Eighteen women (3.6%) who tested normal during pregnancy tested abnormal in the postpartum. Conclusions This study confirms that TSH and TPOAb measured in eluates of blood-spotted filter paper specimens are excellent screening tests to detect primary hypothyroidism and autoimmune thyroiditis in pregnant and postpartum women. Results are very comparable to serum data in this population published in the literature. PMID:24025107

  9. Multiaxial graphite test specimen

    SciTech Connect

    1988-09-01

    A multiaxial test program is to be conducted by Oak Ridge National Laboratory (ORNL) on the core component graphite. The objectives of the tests are to obtain failure data under uniaxial and biaxial states of stress in order to construct a failure surface in a two-dimensional stress space. These data will be used in verifying the accuracy of the maximum stress failure theory being proposed for use in designing the core graphite components. Tubular specimens are proposed to be used and are either loaded axially and/or subjected to internal pressure. This report includes a study on three specimen configurations. The conclusions of that study indicate that an elliptical transition geometry procedures the smallest discontinuity effects. Several loading combustions were studied using the elliptical transition specimen. The primary purpose is to establish the location of the highest stress state and its relation to the gage section for all of the loading conditions. The tension/internal pres sure loading condition (1:1) indicated that the high stress area is just outside the gage section but still should be acceptable. 5 refs., 18 figs.

  10. Radioimmunoassay of ''free thyroxin'' in dried blood spots on filter paper - preliminary observations on the effective differentiation of subjects with congenital hypothyroidism from those with subnormal thyroxin-binding globulin and normal subjects

    SciTech Connect

    Mizuta, H.; Miyai, K.; Ichihara, K.; Amino, N.; Harada, T.; Nose, O.; Tanizawa, O.

    1982-03-01

    In this sensitive, simple method for measuring ''free thyroxin'' (FT/sub 4/) in eluates of dried blood spots on filter paper by use of a radioimmunoassay kit (Amerlex Free T/sub 4/ RIA), the measurable range of FT/sub 4/ is 1.8 to 57 ng/L (equivalent to the concentration in serum), or 7 to 237 fg/tube. The mean coefficients of variation for within assay-within spots, within assay-between spots, and between assays were 5.3%, 5.0%, and 6.2%, respectively. FT/sub 4/ in blood spotted on filter paper is stable for at least a month when dried and kept at either -20/sup 0/C, 4/sup 0/C, room temperature (about 25/sup 0/C), or 37/sup 0/C. The results for FT/sub 4/ in dried blood spots correlated closely with the free-T/sub 4/ concentration in serum (r = 0.99). The method can be used to differentiate cases of primary and secondary hypothyroidism from normal subjects and those with subnormal thyroxin-binding globulin. This method may be useful in screening for congenital hypothyroidism, because sample-retesting is not necessary.

  11. Short Communication: Effect of heat stress during the dry period on gene expression in mammary tissue and peripheral blood mononuclear cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Heat stress (HT) during the dry period compromises mammary gland development, decreases future milk production, and impairs immune status of dairy cows. Our objective was to evaluate the effect of cooling HT cows during the dry period on gene expression of the mammary gland and lymphocytes. Cows wer...

  12. Dry hair

    MedlinePlus

    ... or using harsh soaps or alcohols Excessive blow-drying Dry air Menkes kinky hair syndrome Malnutrition Underactive ... or twice a week Add conditioners Avoid blow drying and harsh styling products

  13. Dry hair

    MedlinePlus

    Some causes of dry hair are: Anorexia nervosa Excessive hair washing, or using harsh soaps or alcohols Excessive blow-drying Dry air Menkes kinky hair syndrome Malnutrition Underactive parathyroid ( ...

  14. Method for thinning specimen

    DOEpatents

    Follstaedt, David M.; Moran, Michael P.

    2005-03-15

    A method for thinning (such as in grinding and polishing) a material surface using an instrument means for moving an article with a discontinuous surface with an abrasive material dispersed between the material surface and the discontinuous surface where the discontinuous surface of the moving article provides an efficient means for maintaining contact of the abrasive with the material surface. When used to dimple specimens for microscopy analysis, a wheel with a surface that has been modified to produce a uniform or random discontinuous surface significantly improves the speed of the dimpling process without loss of quality of finish.

  15. The effects of a 6-month resistance training and dried plum consumption intervention on strength, body composition, blood markers of bone turnover, and inflammation in breast cancer survivors.

    PubMed

    Simonavice, Emily; Liu, Pei-Yang; Ilich, Jasminka Z; Kim, Jeong-Su; Arjmandi, Bahram; Panton, Lynn B

    2014-06-01

    The purpose of this study was to examine the effects of resistance training (RT) and dried plum (DP) consumption on strength, body composition, blood markers of bone, and inflammation in breast cancer survivors (BCS). Twenty-three BCS (RT, n = 12; RT+DP, n = 11), aged 64 ± 7 years, were evaluated at baseline and after 6 months of intervention on the following: muscular strength (chest press and leg extension) via 1-repetition maximums (1RMs); body composition, specifically bone mineral density (BMD) by dual energy X-ray absorptiometry; biochemical markers of bone turnover (bone-specific alkaline phosphatase (BAP), tartrate resistant acid phosphatase (TRAP-5b)); and inflammation (C-reactive protein (CRP)). Target RT prescription was 2 days/week of 10 exercises, including 2 sets of 8-12 repetitions at ∼60%-80% of 1RM. RT+DP also consumed 90 g of DP daily. There were no baseline differences between groups or any group-by-time interactions for any of the variables. BCS increased upper (p < 0.05) (RT: 64 ± 14 to 80 ± 17 kg; RT+DP: 72 ± 23 to 91 ± 20 kg) and lower (p < 0.05) (RT: 69 ± 20 to 87 ± 28 kg; RT+DP: 78 ± 19 to 100 ± 21 kg) body strength. Body composition and BMD improvements were not observed. TRAP-5b decreased in the RT group (p < 0.05) (4.55 ± 1.57 to 4.04 ± 1.63 U/L) and the RT+DP group (p = 0.07) (5.10 ± 2.75 to 4.27 ± 2.03 U/L). Changes in BAP and CRP were not observed. RT was effective for improving biochemical markers of bone turnover and muscular strength in BCS. A longer and higher intensity intervention may be needed to reveal the true effects of RT and DP on body composition and biochemical markers of inflammation. PMID:24869977

  16. Sensitive determination of prohibited drugs in dried blood spots (DBS) for doping controls by means of a benchtop quadrupole/Orbitrap mass spectrometer.

    PubMed

    Thomas, Andreas; Geyer, Hans; Schänzer, Wilhelm; Crone, Catharina; Kellmann, Markus; Moehring, Thomas; Thevis, Mario

    2012-05-01

    In the present study, a new type of mass spectrometer combining a quadrupole mass filter, a higher collision dissociation (HCD) cell and an Orbitrap detector, was evaluated for the analysis of dried blood spots (DBS) in doping controls. DBS analysis is characterized by the necessity to detect prohibited compounds in sub-nanogram-per-milliliter levels with high identification capacity. After extraction of DBS with an organic solvent and liquid chromatographic separation (using a regular C18-RP-analytical UHPLC-column) of target analytes, mass spectrometry is performed with a high-resolution full scan in positive and negative mode by means of electrospray ionisation. Single-product ion mass spectra are acquired using the data-dependent analysis mode (employing an inclusion list) for previously selected precursors of known prohibited compounds with fixed retention time ranges. Besides, a sensitive screening in a targeted approach, non-targeted analysis for retrospective data evaluation is thus possible. The chosen experimental design enables the determination of various drugs from different classes with one generic sample preparation which is shown for 26 selected model compounds (Δ(9)-tetrahydrocannabinol (THC), tetrahydrocannabinol-9-carboxylic acid (THC-COOH), methylhexaneamine, methylphenidate, cocaine, nikethamide, 3,4-methylenedioxyamphetamine, N-methyl-3,4-methylenedioxyamphetamine, strychnine, mesocarb, salbutamol, formoterol, clenbuterol, metandienone, stanozolol, bisoprolol, propranolol, metoprolol, anastrazole, clomiphene, exemestane, dexamethasone, budesonide, selective androgen receptor modulator (SARM) S4 (andarine), SARM S1, hydrochlorothiazide). Generally, only qualitative result interpretation was focussed upon, but for target analytes with deuterium-labelled internal standards (salbutamol, clenbuterol, cocaine, dexamethasone, THC-COOH and THC) quantitative analysis was also possible. Especially the most challenging analytes, THC and its carboxy

  17. Surveillance of Transmitted HIV Drug Resistance Using Matched Plasma and Dried Blood Spot Specimens From Voluntary Counseling and Testing Sites in Ho Chi Minh City, Vietnam, 2007–2008

    PubMed Central

    Hien, Bui Thu; Wagar, Nick; Tram, Tran Hong; Giang, Le Truong; Yang, Chunfu; Wolfe, Mitchell I.; Hien, Nguyen Tran; Tuan, Nguyen Anh

    2012-01-01

    During 2007–2008, surveillance of transmitted human immunodeficiency virus (HIV) drug resistance (TDR) was performed following World Health Organization guidance among clients with newly diagnosed HIV infection attending voluntary counseling and testing (VCT) sites in Ho Chi Minh City (HCMC), Vietnam. Moderate (5%–15%) TDR to nonnucleoside reverse-transcriptase inhibitors (NNRTIs) was observed among VCT clients aged 18–21 years. Follow-up surveillance of TDR in HCMC and other geographic regions of Vietnam is warranted. Data generated will guide the national HIV drug resistance surveillance strategy and support selection of current and future first-line antiretroviral therapy and HIV prevention programs. PMID:22544201

  18. Effects of using ground redberry juniper and dried distillers grains with solubles in lamb feedlot diets: growth, blood serum, fecal, and wool characteristics.

    PubMed

    Whitney, T R; Lupton, C J; Muir, J P; Adams, R P; Stewart, W C

    2014-03-01

    Effects of using ground redberry juniper and dried distillers grains with solubles (DDGS) in Rambouillet lamb (n = 45) feedlot diets on growth, blood serum, fecal, and wool characteristics were evaluated. In a randomized design study with 2 feeding periods (Period 1 = 64% concentrate diet, 35 d; Period 2 = 85% concentrate diet, 56 d), lambs were individually fed 5 isonitrogenous diets: a control diet (CNTL) that contained oat hay but not DDGS or juniper or DDGS-based diets in which 0 (0JUN), 33 (33JUN), 66 (66JUN), or 100% (100JUN) of the oat hay was replaced by juniper. During Period 1, lambs fed CNTL had greater (P < 0.05) DMI and ADG and tended to have greater (P < 0.10) G:F than lambs fed 0JUN or lambs fed DDGS-based diets. Lamb DMI, ADG, and G:F quadratically increased (P < 0.008) as juniper increased in the DDGS-based diets. During Period 2, lambs fed CNTL had greater (P < 0.05) DMI than lambs fed 0JUN or lambs fed DDGS-based diets, but ADG was similar (P > 0.41). Compared to 0JUN, lambs fed CNTL had similar (P = 0.12) G:F and tended to have less G:F (P = 0.07) than lambs fed DDGS-based diets. Among lambs fed DDGS-based diets, DMI was similar (P > 0.19), ADG increased linearly (P = 0.03), and G:F tended to decrease quadratically (P = 0.06) as juniper increased in the diet. Serum IGF-1, serum urea N (SUN), and fecal N were greater (P < 0.05) and serum Ca and P and fecal P were similar (P > 0.13) for lambs fed CNTL vs. lambs fed DDGS-based diets (CNTL). Within lambs fed DDGS-based diets, SUN increased quadratically (P = 0.01) and fecal N increased linearly (P = 0.004), which can partially be attributed to increased dietary urea and condensed tannin intake. Most wool characteristics were not affected, but wool growth per kilogram of BW decreased quadratically (P = 0.04) as percentage of juniper increased in the DDGS-based diets. When evaluating the entire 91-d feeding trial, results indicated that replacing all of the ground oat hay with ground juniper leaves

  19. Dry Mouth

    MedlinePlus

    Dry mouth is the feeling that there is not enough saliva in your mouth. Everyone has a dry mouth once in a while - if they are nervous, ... or under stress. But if you have a dry mouth all or most of the time, it can ...

  20. Dry Mouth

    MedlinePlus

    Dry mouth is the feeling that there is not enough saliva in your mouth. Everyone has a dry mouth once in a while - if they are nervous, ... under stress. But if you have a dry mouth all or most of the time, it can ...

  1. Typing of Plasmodium falciparum DNA from 2 years old Giemsa-stained dried blood spots using nested polymerase chain reaction assay.

    PubMed

    Kumar, D; Dhiman, S; Rabha, B; Goswami, D; Yadav, K; Deka, M; Veer, V; Baruah, I

    2016-01-01

    A panel of 129 Giemsa-stained thick blood spots (TBS) confirmed for Plasmodium falciparum infection having different levels of parasite density were collected from a malaria endemic area. DNA was extracted and nested polymerase chain reaction (PCR) assay was performed to amplify P. falciparum DNA. Nested PCR assay successfully amplified P. falciparum DNA at a very low parasitaemia of ~10 parasites/μl of blood. Current PCR assay is very simple and can be used retrospectively to monitor the invasion and prevalence of different Plasmodium species in endemic areas. PMID:27080775

  2. Pilot environmental specimen bank program

    SciTech Connect

    Wise, S.A.; Zeisler, R.

    1984-10-01

    The concept of an environmental specimen bank for archiving of biological and environmental samples for retrospective analysis has been recognized recently as an important component of systematic environmental monitoring. A pilot program was designed to evaluate the feasibility of a national program by providing actual working experience in all aspects of specimen banking, that is, in specimen collection, processing, storage, and analysis. Four types of environmental specimens, which represent environmental accumulators, were selected for inclusion in the National Bureau of Standards pilot program: human soft tissue (liver), a marine accumulator (marine mussels, Mytilus edulis), a food accumulator and the air pollutant accumulator have not been selected. Attention is focused on the experience gained in the pilot program with the human liver specimens. 32 references, 2 figures, 1 table.

  3. Chromosome aberrations induced in human lymphocytes by U-235 fission neutrons: I. Irradiation of human blood samples in the "dry cell" of the TRIGA Mark II nuclear reactor.

    PubMed

    Fajgelj, A; Lakoski, A; Horvat, D; Remec, I; Skrk, J; Stegnar, P

    1991-11-01

    A set-up for irradiation of biological samples in the TRIGA Mark II research reactor in Ljubljana is described. Threshold activation detectors were used for characterisation of the neutron flux, and the accompanying gamma dose was measured by TLDs. Human peripheral blood samples were irradiated "in vitro" and biological effects evaluated according to the unstable chromosomal aberrations induced. Biological effects of two types of cultivation of irradiated blood samples, the first immediately after irradiation and the second after 96 h storage, were studied. A significant difference in the incidence of chromosomal aberrations between these two types of samples was obtained, while our dose-response curve fitting coefficients alpha 1 = (7.71 +/- 0.09) x 10(-2) Gy-1 (immediate cultivation) and alpha 2 = (11.03 +/- 0.08) x 10(-2) Gy-1 (96 h delayed cultivation) are in both cases lower than could be found in the literature. PMID:1962281

  4. Influence of thermal conditioning media on Charpy specimen test temperature

    SciTech Connect

    Nanstad, R.K.; Swain, R.L.; Berggren, R.G.

    1989-01-01

    The Charpy V-notch (CVN) impact test is used extensively for determining the toughness of structural materials. Research programs in many technologies concerned with structural integrity perform such testing to obtain Charpy energy vs temperature curves. American Society for Testing and Materials Method E 23 includes rather strict requirements regarding determination and control of specimen test temperature. It specifies minimum soaking times dependent on the use of liquids or gases as the medium for thermally conditioning the specimen. The method also requires that impact of the specimen occur within 5 s removal from the conditioning medium. It does not, however, provide guidance regarding choice of conditioning media. This investigation was primarily conducted to investigate the changes in specimen temperature which occur when water is used for thermal conditioning. A standard CVN impact specimen of low-alloy steel was instrumented with surface-mounted and embedded thermocouples. Dependent on the media used, the specimen was heated or cooled to selected temperatures in the range {minus}100 to 100{degree}C using cold nitrogen gas, heated air, acetone and dry ice, methanol and dry ice, heated oil, or heated water. After temperature stabilization, the specimen was removed from the conditioning medium while the temperatures were recorded four times per second from all thermocouples using a data acquisition system and a computer. The results show that evaporative cooling causes significant changes in the specimen temperatures when water is used for conditioning. Conditioning in the other media did not result in such significant changes. The results demonstrate that, even within the guidelines of E 23, significant test temperature changes can occur which may substantially affect the Charpy impact test results if water is used for temperature conditioning. 7 refs., 11 figs.

  5. Dry socket

    MedlinePlus

    ... care for the dry socket at home: Take pain medicine and antibiotics as directed Apply a cold pack to the outside of your jaw Carefully rinse the dry socket as directed by your dentist If taking antibiotics, avoid smoking or using tobacco and alcohol

  6. Manufacturing of Plutonium Tensile Specimens

    SciTech Connect

    Knapp, Cameron M

    2012-08-01

    Details workflow conducted to manufacture high density alpha Plutonium tensile specimens to support Los Alamos National Laboratory's science campaigns. Introduces topics including the metallurgical challenge of Plutonium and the use of high performance super-computing to drive design. Addresses the utilization of Abaqus finite element analysis, programmable computer numerical controlled (CNC) machining, as well as glove box ergonomics and safety in order to design a process that will yield high quality Plutonium tensile specimens.

  7. Ultra-high performance liquid chromatography tandem mass spectrometric method for the determination of tamoxifen, N-desmethyltamoxifen, 4-hydroxytamoxifen and endoxifen in dried blood spots--development, validation and clinical application during breast cancer adjuvant therapy.

    PubMed

    Antunes, Marina Venzon; Raymundo, Suziane; de Oliveira, Vanessa; Staudt, Dilana Elisabeth; Gössling, Gustavo; Peteffi, Giovana Piva; Biazús, Jorge Villanova; Cavalheiro, José Antônio; Tre-Hardy, Marie; Capron, Arnaud; Haufroid, Vincent; Wallemacq, Pierre; Schwartsmann, Gilberto; Linden, Rafael

    2015-01-01

    A LC-MSMS method for the simultaneous determination of tamoxifen, N-desmethyltamoxifen, 4-hydroxytamoxifen and endoxifen in dried blood spots samples was developed and validated. The method employs an ultrasound-assisted liquid extraction and a reversed phase separation in an Acquity(®) C18 column (150×2.1 mm, 1.7 µm). Mobile phase was a mixture of formic acid 0.1% (v/v) pH 2.7 and acetonitrile (gradient from 60:40 to 50:50, v/v). Total analytical run time was 8 min. Precision assays showed CV % lower than 10.75% and accuracy in the range 94.5 to 110.3%. Mean analytes recoveries from DBS ranged from 40% to 92%. The method was successfully applied to 91 paired clinical DBS and plasma samples. Dried blood spots concentrations were highly correlated to plasma, with rs>0.83 (P<0.01). Median estimated plasma concentrations after hematocrit and partition factor adjustment were: TAM 123.3 ng mL(-1); NDT 267.9 ng mL(-1), EDF 10.0 ng mL(-1) and HTF 1.3 ng mL(-1,) representing in average 98 to 104% of the actually measured concentrations. The DBS method was able to identify 96% of patients with plasma EDF concentrations below the clinical threshold related to better prognosis (5.9 ng mL(-1)). The procedure has adequate analytical performance and can be an efficient tool to optimize adjuvant breast cancer treatment, especially in resource limited settings. PMID:25476377

  8. Collection & Processing of Vertebrate Specimens for Arbovirus Studies.

    ERIC Educational Resources Information Center

    Sudia, W. Daniel; And Others

    Described are techniques used by the National Communicable Disease Center in obtaining blood and tissues from man and other vertebrates for arbovirus isolation and antibody studies. Also included are techniques for capturing and handling vertebrates; banding and marking; restraining and bleeding; storing of specimens to preserve antibody and…

  9. Effects of feeding a spray-dried multivalent polyclonal antibody preparation on feedlot performance, feeding behavior, carcass characteristics, rumenitis, and blood gas profile of Brangus and Nellore yearling bulls.

    PubMed

    Millen, D D; Pacheco, R D L; DiLorenzo, N; Martins, C L; Marino, C T; Bastos, J P S T; Mariani, T M; Barducci, R S; Sarti, L M N; DiCostanzo, A; Rodrigues, P H M; Arrigoni, M D B

    2015-09-01

    The objective of this study was to evaluate the effects of replacing monensin (MON) with a spray-dried multivalent polyclonal antibody preparation (PAP) against several ruminal microorganisms on feedlot performance, carcass characteristics, feeding behavior, blood gas profile, and the rumenitis incidence of Brangus and Nellore yearling bulls. The study was designed as a completely randomized design with a 2 × 2 factorial arrangement, replicated 6 times (4 bulls per pen and a total of 24 pens), in which bulls ( = 48) of each biotype were fed diets containing either MON fed at 300 mg/d or PAP fed at 3 g/d. No significant feed additive main effects were observed for ADG ( = 0.27), G:F ( = 0.28), HCW ( = 0.99), or dressing percentage ( = 0.80). However, bulls receiving PAP had greater DMI ( = 0.02) and larger ( = 0.02) final LM area as well as greater ( < 0.01) blood concentrations of bicarbonate and base excess in the extracellular fluid than bulls receiving MON. Brangus bulls had greater ( < 0.01) ADG and DMI expressed in kilograms, final BW, heavier HCW, and larger initial and final LM area than Nellore bulls. However, Nellore bulls had greater daily DMI fluctuation ( < 0.01), expressed as a percentage, and greater incidence of rumenitis ( = 0.05) than Brangus bulls. In addition, Brangus bulls had greater ( < 0.01) DMI per meal and also presented lower ( < 0.01) DM and NDF rumination rates when compared with Nellore bulls. Significant interactions ( < 0.05) between biotype and feed additive were observed for SFA, unsaturated fatty acids (UFA), MUFA, and PUFA concentrations in adipose tissues. When Nellore bulls were fed PAP, fat had greater ( < 0.05) SFA and PUFA contents but less ( < 0.01) UFA and MUFA than Nellore bulls receiving MON. For Brangus bulls, MON led to greater ( < 0.05) SFA and PUFA and less ( < 0.05) UFA and MUFA than Brangus bulls fed PAP. Feeding a spray-dried PAP led to similar feedlot performance compared with that when feeding MON. Spray-dried

  10. Feasibility of Using the Mosquito Blood Meal for Rapid and Efficient Human and Animal Virus Surveillance and Discovery.

    PubMed

    Yang, Yu; Garver, Lindsey S; Bingham, Karen M; Hang, Jun; Jochim, Ryan C; Davidson, Silas A; Richardson, Jason H; Jarman, Richard G

    2015-12-01

    Mosquito blood meals taken from humans and animals potentially represent a useful source of blood for the detection of blood-borne pathogens. In this feasibility study, Anopheles stephensi mosquitoes were fed with blood meals spiked with dengue virus type 2 (DENV-2) and harvested at serial time points. These mosquitoes are not competent vectors, and the virus is not expected to replicate. Ingested blood was spotted on Whatman FTA cards and stored at room temperature. Mosquito abdomens were removed and stored at -80°C. Control blood meal aliquots were stored in vials or applied onto FTA cards. After 4 weeks of storage, the samples were extracted using beadbeating and QIAamp Viral RNA kit (Qiagen Sciences, Germantown, MD). Recovered viral RNA was analyzed by DENV-2 TaqMan RT-PCR assay and next-generation sequencing (NGS). Overall viral RNA recovery efficiency was 15% from the directly applied dried blood spots and approximately 20% or higher for dried blood spots made by blotting mosquito midgut on FTA cards. Viral RNA in mosquito-ingested blood decreases over time, but remains detectable 24 hours after blood feeding. The viral sequences in FTA-stored specimens can be maintained at room temperature. The strategy has the potential utility in expedited zoonotic virus discovery and blood-borne pathogen surveillance. PMID:26416112

  11. Uric acid - blood

    MedlinePlus

    Uric acid is a chemical created when the body breaks down substances called purines. Purines are found in some ... dried beans and peas, and beer. Most uric acid dissolves in blood and travels to the kidneys. ...

  12. Performance of an Early Infant Diagnostic Test, AmpliSens DNA-HIV-FRT, Using Dried Blood Spots Collected from Children Born to Human Immunodeficiency Virus-Infected Mothers in Ukraine.

    PubMed

    Chang, Joy; Tarasova, Tetyana; Shanmugam, Vedapuri; Azarskova, Marianna; Nguyen, Shon; Hurlston, Mackenzie; Sabatier, Jennifer; Zhang, Guoqing; Osmanov, Saladin; Ellenberger, Dennis; Yang, Chunfu; Vitek, Charles; Liulchuk, Maria; Nizova, Natalya

    2015-12-01

    An accurate accessible test for early infant diagnosis (EID) is crucial for identifying HIV-infected infants and linking them to treatment. To improve EID services in Ukraine, dried blood spot (DBS) samples obtained from 237 HIV-exposed children (≤18 months of age) in six regions in Ukraine in 2012 to 2013 were tested with the AmpliSens DNA-HIV-FRT assay, the Roche COBAS AmpliPrep/COBAS TaqMan (CAP/CTM) HIV-1 Qual test, and the Abbott RealTime HIV-1 Qualitative assay. In comparison with the paired whole-blood results generated from AmpliSens testing at the oblast HIV reference laboratories in Ukraine, the sensitivity was 0.99 (95% confidence interval [CI], 0.95 to 1.00) for the AmpliSens and Roche CAP/CTM Qual assays and 0.96 (95% CI, 0.90 to 0.98) for the Abbott Qualitative assay. The specificity was 1.00 (95% CI, 0.97 to 1.00) for the AmpliSens and Abbott Qualitative assays and 0.99 (95% CI, 0.96 to 1.00) for the Roche CAP/CTM Qual assay. McNemar analysis indicated that the proportions of positive results for the tests were not significantly different (P > 0.05). Cohen's kappa (0.97 to 0.99) indicated almost perfect agreement among the three tests. These results indicated that the AmpliSens DBS and whole-blood tests performed equally well and were comparable to the two commercially available EID tests. More importantly, the performance characteristics of the AmpliSens DBS test meets the World Health Organization EID test requirements; implementing AmpliSens DBS testing might improve EID services in resource-limited settings. PMID:26447114

  13. century drying

    NASA Astrophysics Data System (ADS)

    Cook, Benjamin I.; Smerdon, Jason E.; Seager, Richard; Coats, Sloan

    2014-11-01

    Global warming is expected to increase the frequency and intensity of droughts in the twenty-first century, but the relative contributions from changes in moisture supply (precipitation) versus evaporative demand (potential evapotranspiration; PET) have not been comprehensively assessed. Using output from a suite of general circulation model (GCM) simulations from phase 5 of the Coupled Model Intercomparison Project, projected twenty-first century drying and wetting trends are investigated using two offline indices of surface moisture balance: the Palmer Drought Severity Index (PDSI) and the Standardized Precipitation Evapotranspiration Index (SPEI). PDSI and SPEI projections using precipitation and Penman-Monteith based PET changes from the GCMs generally agree, showing robust cross-model drying in western North America, Central America, the Mediterranean, southern Africa, and the Amazon and robust wetting occurring in the Northern Hemisphere high latitudes and east Africa (PDSI only). The SPEI is more sensitive to PET changes than the PDSI, especially in arid regions such as the Sahara and Middle East. Regional drying and wetting patterns largely mirror the spatially heterogeneous response of precipitation in the models, although drying in the PDSI and SPEI calculations extends beyond the regions of reduced precipitation. This expansion of drying areas is attributed to globally widespread increases in PET, caused by increases in surface net radiation and the vapor pressure deficit. Increased PET not only intensifies drying in areas where precipitation is already reduced, it also drives areas into drought that would otherwise experience little drying or even wetting from precipitation trends alone. This PET amplification effect is largest in the Northern Hemisphere mid-latitudes, and is especially pronounced in western North America, Europe, and southeast China. Compared to PDSI projections using precipitation changes only, the projections incorporating both

  14. Positron lifetime studies in thermoplastic polyimide test specimens

    NASA Technical Reports Server (NTRS)

    Singh, J. J.; Stclair, T. L.; Holt, W. H.; Mock, W., Jr.

    1982-01-01

    Positron lifetime measurements were made in two thermoplastic polyimide materials recently developed at Langley. The long component lifetime values in polyimidesulfone samples are 847 + or - 81 Ps (dry) and 764 + or - 91 Ps (saturated). The corresponding values in LARC thermoplastic imides are 1080 + or - 139 Ps (dry) and 711 + or - 96 Ps (saturated). Clearly, the presence of moisture has greater effect on positron lifetime in LARC thermoplastic imides than in the case of polyimidesulfones. This result is consistent with the photomicrographic observations made on frozen water saturated specimens of these materials.

  15. Dry cell battery poisoning

    MedlinePlus

    Batteries - dry cell ... Acidic dry cell batteries contain: Manganese dioxide Ammonium chloride Alkaline dry cell batteries contain: Sodium hydroxide Potassium hydroxide Lithium dioxide dry cell batteries ...

  16. Electrothermal fracturing of tensile specimens

    NASA Technical Reports Server (NTRS)

    Blinn, H. O.; Hanks, J. G.; Perkins, H. P.

    1970-01-01

    Pulling device consisting of structural tube, connecting rod, spring-loaded nuts, loading rod, heating element, and three bulkheads fractures tensile specimens. Alternate heating and cooling increases tensile loading by increments until fracturing occurs. Load cell or strain gage, applied to pulling rod, determines forces applied.

  17. An interlaminar tension strength specimen

    NASA Technical Reports Server (NTRS)

    Jackson, Wade C.; Martin, Roderick H.

    1992-01-01

    This paper describes a technique to determine interlaminar tension strength, sigma(sub 3c) of a fiber reinforced composite material using a curved beam. The specimen was a unidirectional curved beam, bent 90 degrees, with straight arms. Attached to each arm was a hinged loading mechanism which was held by the grips of a tensile testing machine. Geometry effects of the specimen, including the effects of loading arm length, inner radius, thickness, and width, were studied. The data sets fell into two categories: low strength corresponding to a macroscopic flaw related failure and high strength corresponding to a microscopic flaw related failure. From the data available, the loading arm length had no effect on sigma(sub 3c). The inner radius was not expected to have a significant effect on sigma(sub 3c), but this conclusion could not be confirmed because of differences in laminate quality for each curve geometry. The thicker specimens had the lowest value of sigma(sub 3c) because of poor laminate quality. Width was found to affect the value of sigma(sub 3c) only slightly. The wider specimens generally had a slightly lower strength since more material was under high stress, and hence, had a larger probability of containing a significant flaw.

  18. An Interlaminar Tensile Strength Specimen

    NASA Technical Reports Server (NTRS)

    Martin, Roderick H.; Jackson, Wade C.

    1993-01-01

    This paper describes a technique to determine interlaminar tensile strength, sigma(sub 3c), of a fiber reinforced composite material using a curved beam. The specimen was a unidirectional curved beam, bent 90 deg, with straight arms. Attached to each arm was a hinged loading mechanism that was held by the grips of a tension testing machine. Geometry effects of the specimen, including the effects of loading arm length, inner radius, thickness, and width, were studied. The data sets fell into two categories: low strength corresponding to a macroscopic flaw related failure and high strength corresponding to a microscopic flaw related failure. From the data available, the specimen width and loading arm length had little effect on sigma(sub 3c). The inner radius was not expected to have a significant effect on sigma(sub 3c), but this conclusion could not be confirmed because of differences in laminate quality for each curve geometry. The thicker specimens had the lowest value of sigma(sub 3c) because of poor laminate quality.

  19. 10 CFR 26.165 - Testing split specimens and retesting single specimens.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... single specimens. (a) Testing split specimens. (1) If a specimen has been split into Bottle A and Bottle... required, of the specimen in Bottle A. (2) If a specimen was initially tested at a licensee testing... laboratory shall perform initial and confirmatory testing, if required, of the specimen in Bottle A. (3)...

  20. 10 CFR 26.165 - Testing split specimens and retesting single specimens.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... single specimens. (a) Testing split specimens. (1) If a specimen has been split into Bottle A and Bottle... required, of the specimen in Bottle A. (2) If a specimen was initially tested at a licensee testing... laboratory shall perform initial and confirmatory testing, if required, of the specimen in Bottle A. (3)...

  1. Baseline Test Specimen Machining Report

    SciTech Connect

    mark Carroll

    2009-08-01

    The Next Generation Nuclear Plant (NGNP) Project is tasked with selecting a high temperature gas reactor technology that will be capable of generating electricity and supplying large amounts of process heat. The NGNP is presently being designed as a helium-cooled high temperature gas reactor (HTGR) with a large graphite core. The graphite baseline characterization project is conducting the research and development (R&D) activities deemed necessary to fully qualify nuclear-grade graphite for use in the NGNP reactor. Establishing nonirradiated thermomechanical and thermophysical properties by characterizing lot-to-lot and billet-to-billet variations (for probabilistic baseline data needs) through extensive data collection and statistical analysis is one of the major fundamental objectives of the project. The reactor core will be made up of stacks of graphite moderator blocks. In order to gain a more comprehensive understanding of the varying characteristics in a wide range of suitable graphites, any of which can be classified as “nuclear grade,” an experimental program has been initiated to develop an extensive database of the baseline characteristics of numerous candidate graphites. Various factors known to affect the properties of graphite will be investigated, including specimen size, spatial location within a graphite billet, specimen orientation within a billet (either parallel to [P] or transverse to [T] the long axis of the as-produced billet), and billet-to-billet variations within a lot or across different production lots. Because each data point is based on a certain position within a given billet of graphite, particular attention must be paid to the traceability of each specimen and its spatial location and orientation within each billet. The evaluation of these properties is discussed in the Graphite Technology Development Plan (Windes et. al, 2007). One of the key components in the evaluation of these graphite types will be mechanical testing on

  2. Colorful drying.

    PubMed

    Lakio, Satu; Heinämäki, Jyrki; Yliruusi, Jouko

    2010-03-01

    Drying is one of the standard unit operations in the pharmaceutical industry and it is important to become aware of the circumstances that dominate during the process. The purpose of this study was to test microcapsulated thermochromic pigments as heat indicators in a fluid bed drying process. The indicator powders were manually granulated with alpha-lactose monohydrate resulting in three particle-size groups. Also, pellets were coated with the indicator powders. The granules and pellets were fluidized in fluid bed dryer to observe the progress of the heat flow in the material and to study the heat indicator properties of the indicator materials. A tristimulus colorimeter was used to measure CIELAB color values. Color indicator for heat detection can be utilized to test if the heat-sensitive API would go through physical changes during the pharmaceutical drying process. Both the prepared granules and pellets can be used as heat indicator in fluid bed drying process. The colored heat indicators give an opportunity to learn new aspects of the process at real time and could be exploded, for example, for scaling-up studies. PMID:20039220

  3. Dry Eye

    MedlinePlus

    ... surgery, called punctal cautery, is recommended to permanently close the drainage holes. The procedure helps keep the limited volume of tears on the eye for a longer period of time. In some patients with dry eye, supplements or dietary sources (such as tuna fish) of omega-3 fatty ...

  4. Standard Specimen Reference Set: Breast — EDRN Public Portal

    Cancer.gov

    The primary objective of this study is to assemble a well-characterized set of blood specimens to test biomarkers that, in conjunction with mammography, can detect and discriminate breast cancer. These samples will be divided to provide “sets” of specimens that can be tested in a number of different laboratories. Since tests will be performed on the same sets of samples, the data will be directly comparable and decisions regarding which biomarker or set of biomarkers have value in breast cancer detection can be made. These sets will reside at a National Cancer Institute facility at Frederick, MD.

  5. Appendix A: Specimen 72275 documentation

    NASA Technical Reports Server (NTRS)

    Marvin, U. B.

    1974-01-01

    The friability of the matrix of specimen 72275 caused numerous fragments and an abundance of fines to break away from the main mass during transport from the moon and handling in the lunar receiving laboratory. Samples 72275,1 to 72275,14 were labeled during PET examination. Samples 72275,1, 4, 6, 7, 8, and 9 were placed in storage, and the remainder were distributed.

  6. Eccentric loading of microtensile specimens

    NASA Technical Reports Server (NTRS)

    Trapp, Mark A.

    2004-01-01

    Ceramic materials have a lower density than most metals and are capable of performing at extremely high temperatures. The utility of these materials is obvious; however, the fracture strength of brittle materials is not easily predicted and often varies greatly. Characteristically, brittle materials lack ductility and do not yield as other materials. Ceramics materials are naturally populated with microscopic cracks due to fabrication techniques. Upon application of a load, stress concentration occurs at the root of these cracks and fracture will eventually occur at some not easily predicted strength. In order to use ceramics in any application some design methodology must exist from which a component can be placed into service. This design methodology is CARES/LIFE (Ceramics Analysis and Reliability Evaluation of Structures) which has been developed and refined at NASA over the last several decades. The CARES/LIFE computer program predicts the probability of failure of a ceramic component over its service life. CARES combines finite element results from a commercial FE (finite element) package such as ANSYS and experimental results to compute the abovementioned probability of failure. Over the course of several tests CARES has had great success in predicting the life of various ceramic components and has been used throughout industry. The latest challenge is to verify that CARES is valid for MEMS (Micro-Electro Mechanical Systems). To investigate a series of microtensile specimens were fractured in the laboratory. From this data, material parameters were determined and used to predict a distribution of strength for other specimens that exhibit a known stress concentration. If the prediction matches the experimental results then these parameters can be applied to a desired component outside of the laboratory. During testing nearly half of the tensile Specimens fractured at a location that was not expected and hence not captured in the FE model. It has been my duty

  7. Compound Charpy specimens by adhesive joining

    NASA Astrophysics Data System (ADS)

    Ghoneim, M. M.; Hammad, F. H.; Pachur, D.; Britz, L.

    1992-03-01

    Compound (reconstituted) Charpy specimens were manufactured by an adhesive joining method in which each half of a previously tested specimen formed the central section of a new testpiece. 29 adhesives were screened to select the most suitable. Compound specimens were precracked and used as minature fracture mechanics specimens and tested in both 3-point static bending and impact. The results are in good agreement with those of conventional specimens. Recommendations for the most appropriate commercial adhesive for hot cell operations are given.

  8. In Vitro Fracture Testing of Submicron Diameter Collagen Fibril Specimens

    PubMed Central

    Shen, Zhilei Liu; Dodge, Mohammad Reza; Kahn, Harold; Ballarini, Roberto; Eppell, Steven J.

    2010-01-01

    Mechanical testing of collagenous tissues at different length scales will provide improved understanding of the mechanical behavior of structures such as skin, tendon, and bone, and also guide the development of multiscale mechanical models. Using a microelectromechanical-systems (MEMS) platform, stress-strain response curves up to failure of type I collagen fibril specimens isolated from the dermis of sea cucumbers were obtained in vitro. A majority of the fibril specimens showed brittle fracture. Some displayed linear behavior up to failure, while others displayed some nonlinearity. The fibril specimens showed an elastic modulus of 470 ± 410 MPa, a fracture strength of 230 ± 160 MPa, and a fracture strain of 80% ± 44%. The fibril specimens displayed significantly lower elastic modulus in vitro than previously measured in air. Fracture strength/strain obtained in vitro and in air are both significantly larger than those obtained in vacuo, indicating that the difference arises from the lack of intrafibrillar water molecules produced by vacuum drying. Furthermore, fracture strength/strain of fibril specimens were different from those reported for collagenous tissues of higher hierarchical levels, indicating the importance of obtaining these properties at the fibrillar level for multiscale modeling. PMID:20858445

  9. In vitro fracture testing of submicron diameter collagen fibril specimens.

    PubMed

    Shen, Zhilei Liu; Dodge, Mohammad Reza; Kahn, Harold; Ballarini, Roberto; Eppell, Steven J

    2010-09-22

    Mechanical testing of collagenous tissues at different length scales will provide improved understanding of the mechanical behavior of structures such as skin, tendon, and bone, and also guide the development of multiscale mechanical models. Using a microelectromechanical-systems (MEMS) platform, stress-strain response curves up to failure of type I collagen fibril specimens isolated from the dermis of sea cucumbers were obtained in vitro. A majority of the fibril specimens showed brittle fracture. Some displayed linear behavior up to failure, while others displayed some nonlinearity. The fibril specimens showed an elastic modulus of 470 ± 410 MPa, a fracture strength of 230 ± 160 MPa, and a fracture strain of 80% ± 44%. The fibril specimens displayed significantly lower elastic modulus in vitro than previously measured in air. Fracture strength/strain obtained in vitro and in air are both significantly larger than those obtained in vacuo, indicating that the difference arises from the lack of intrafibrillar water molecules produced by vacuum drying. Furthermore, fracture strength/strain of fibril specimens were different from those reported for collagenous tissues of higher hierarchical levels, indicating the importance of obtaining these properties at the fibrillar level for multiscale modeling. PMID:20858445

  10. [Use of dried blood spots in early diagnosis of HIV-1 infection in children born to HIV-infected mothers as part of the prevention of mother-to-child transmission in Benin].

    PubMed

    Tchiakpe, E; Hounto-Ogouyemi, A; Diop Ndiaye, H; Diouara, A A M; Aïssi, A K; Keke, R K; Kpangon, A A; Lafia, B; Métadokou, D; Bouraïma, B; Anthony, D; Hounsinou, A; Alao, M J; Azondekon, A; Ahouidi, A D; Bei, A K; Mbengue, M A S; Touré Kane, C; Zannou, D M

    2016-08-01

    The goal of this study was to evaluate using the molecular diagnosis, infection transmission rate of HIV in children born to HIV-1 positive mothers as part of the prevention of mother-to-child transmission (PMTCT) in Benin. The sample consisted of 524 dried blood spots (DBS) of children born to HIV-1 positive mothers, from 30 sites (PMTCT) taken between October 2009 and June 2010. The diagnosis of HIV-1 was performed by the qualitative detection of viral nucleic acids (RNA and DNA) in DBS on filter paper using the Abbott RealTime(®) HIV-1 Qualitative assay. We found that 51 DBS were positive (9.7%) and 473 were negative (90.3%). The failure rate of PMTCT among 420 mothers who received antiretroviral prophylaxis was 6.7% (28/420). This failure rate was significantly higher among children born to infected mothers on antiretroviral monotherapy than on triple therapy (HAART). The results of our study enrich the data in the literature on highly active antiretroviral chemoprophylaxis to reduce the transmission of HIV-1 from mother to child. PMID:27385037

  11. Assessing the impact of a nurse-delivered home dried blood spot service on uptake of testing for household contacts of hepatitis B-infected pregnant women across two London trusts.

    PubMed

    Keel, P; Edwards, G; Flood, J; Nixon, G; Beebeejaun, K; Shute, J; Poh, J; Millar, A; Ijaz, S; Parry, J; Mandal, S; Ramsay, M; Amirthalingam, G

    2016-07-01

    Despite national guidance recommending testing and vaccination of household contacts of hepatitis B-infected pregnant women, provision and uptake of this is sub-optimal. The aim of this study was to evaluate the use of in-home dried blood spot (DBS) testing to increase testing and vaccination of household contacts of hepatitis B-infected pregnant women as an alternative approach to conventional primary-care follow-up. The study was conducted across two London maternity trusts (North Middlesex and Newham). All hepatitis B surface antigen-positive pregnant women identified through these trusts were eligible for inclusion. The intervention of in-home DBS testing for household contacts was introduced at North Middlesex Trust from November 2010 to December 2011. Data on testing and vaccination uptake from GP records across the two trusts were compared between baseline (2009) and intervention (2010-2011) periods. In-home DBS service increased testing uptake for all ages (P < 0·001) with the biggest impact seen in partners, where testing increased from 30·3% during the baseline period to 96·6% during the intervention period in North Middlesex Trust. Although impact on vaccine uptake was less marked, improvements were observed for adults. The provision of nurse-led home-based DBS may be useful in areas of high prevalence. PMID:26833270

  12. Strong Correlation Between Concentrations of Tenofovir (TFV) Emtricitabine (FTC) in Hair and TFV Diphosphate and FTC Triphosphate in Dried Blood Spots in the iPrEx Open Label Extension: Implications for Pre-exposure Prophylaxis Adherence Monitoring.

    PubMed

    Gandhi, Monica; Glidden, David V; Liu, Albert; Anderson, Peter L; Horng, Howard; Defechereux, Patricia; Guanira, Juan V; Grinsztejn, Beatriz; Chariyalertsak, Suwat; Bekker, Linda-Gail; Grant, Robert M

    2015-11-01

    Self-reported adherence to pre-exposure prophylaxis (PrEP) has limitations, raising interest in pharmacologic monitoring. Drug concentrations in hair and dried blood spots (DBS) are used to assess long-term-exposure; hair shipment/storage occurs at room temperature. The iPrEx Open Label Extension collected DBS routinely, with opt-in hair collection; concentrations were measured with liquid chromatography/tandem mass spectrometry. In 806 hair-DBS pairs, tenofovir (TFV) hair levels and TFV diphosphate (DP) in DBS were strongly correlated (Spearman coefficient r = 0.734; P < .001), as were hair TFV/DBS emtricitabine (FTC) triphosphate (TP) (r = 0.781; P < .001); hair FTC/DBS TFV-DP (r = 0.74; P < .001); hair FTC/DBS FTC-TP (r = 0.587; P < .001). Drug detectability was generally concordant by matrix. Hair TFV/FTC concentrations correlate strongly with DBS levels, which are predictive of PrEP outcomes. PMID:25895984

  13. Agreement for HPV genotyping detection between self-collected specimens on a FTA cartridge and clinician-collected specimens

    PubMed Central

    Guan, YaoYao; Gravitt, Patti E.; Howard, Roslyn; Eby, Yolanda J.; Wang, Shaoming; Li, Belinda; Feng, Changyan; Qiao, You-Lin; Castle, Philip E.

    2016-01-01

    The current method of transporting self-collected cervicovaginal specimen for HPV DNA testing relies on liquid based medium, which is challenging and expensive to transport. A novel, dry storage and transportation device, Whatman indicating FTA™ Elute Cartridge, avoids some of the pitfalls of liquid-based medium. This method has been shown to be comparable to liquid-based collection medium, but relative performance of self-collected (SC) and clinician-collected (CC) samples onto FTA cards has not been reported. The objective of this study is to compare the analytic performance of self- and clinician-collected samples onto FTA cartridges for the detection of carcinogenic HPV using Linear Array. There was a 91% agreement, 69% positive agreement, and kappa of 0.75 between the clinician-collected and self-collected specimens for detection of any carcinogenic HPV genotype. When the HPV results were categorized hierarchically according to cervical cancer risk, there was no difference in the distribution of the HPV results for the clinician- and self-collected specimens (p = 0.7). This study concludes that FTA elute cartridge is a promising method of specimen transport for cervical cancer screening programs considering using self-collected specimen and HPV testing. Larger studies with clinical endpoints are now needed to assess the clinical performance. PMID:23370404

  14. Multiphoton microspectroscopy of biological specimens

    NASA Astrophysics Data System (ADS)

    Lin, Bai-Ling; Kao, Fu-Jen; Cheng, Ping C.; Sun, Chi-Kuang; Chen, RangWu; Wang, YiMin; Chen, JianCheng; Wang, Yung-Shun; Liu, Tzu-Ming; Huang, Mao-Kuo

    2000-07-01

    The non-linear nature of multi-photon fluorescence excitation restricts the fluorescing volume to the vicinity of the focal point. As a result, the technology has the capacity for micro- spectroscopy of biological specimen at high spatial resolution. Chloroplasts in mesophyll protoplast of Arabidopsis thaliana and maize stem sections were used to demonstrate the feasibility of multi-photon fluorescence micro-spectroscopy at subcellular compartments. Time-lapse spectral recording provides a means for studying the response of cell organelles to high intensity illumination.

  15. Automated Tracking of Drosophila Specimens

    PubMed Central

    Chao, Rubén; Macía-Vázquez, Germán; Zalama, Eduardo; Gómez-García-Bermejo, Jaime; Perán, José-Ramón

    2015-01-01

    The fruit fly Drosophila Melanogaster has become a model organism in the study of neurobiology and behavior patterns. The analysis of the way the fly moves and its behavior is of great scientific interest for research on aspects such as drug tolerance, aggression or ageing in humans. In this article, a procedure for detecting, identifying and tracking numerous specimens of Drosophila by means of computer vision-based sensing systems is presented. This procedure allows dynamic information about each specimen to be collected at each moment, and then for its behavior to be quantitatively characterized. The proposed algorithm operates in three main steps: a pre-processing step, a detection and segmentation step, and tracking shape. The pre-processing and segmentation steps allow some limits of the image acquisition system and some visual artifacts (such as shadows and reflections) to be dealt with. The improvements introduced in the tracking step allow the problems corresponding to identity loss and swaps, caused by the interaction between individual flies, to be solved efficiently. Thus, a robust method that compares favorably to other existing methods is obtained. PMID:26258779

  16. Automated Tracking of Drosophila Specimens.

    PubMed

    Chao, Rubén; Macía-Vázquez, Germán; Zalama, Eduardo; Gómez-García-Bermejo, Jaime; Perán, José-Ramón

    2015-01-01

    The fruit fly Drosophila Melanogaster has become a model organism in the study of neurobiology and behavior patterns. The analysis of the way the fly moves and its behavior is of great scientific interest for research on aspects such as drug tolerance, aggression or ageing in humans. In this article, a procedure for detecting, identifying and tracking numerous specimens of Drosophila by means of computer vision-based sensing systems is presented. This procedure allows dynamic information about each specimen to be collected at each moment, and then for its behavior to be quantitatively characterized. The proposed algorithm operates in three main steps: a pre-processing step, a detection and segmentation step, and tracking shape. The pre-processing and segmentation steps allow some limits of the image acquisition system and some visual artifacts (such as shadows and reflections) to be dealt with. The improvements introduced in the tracking step allow the problems corresponding to identity loss and swaps, caused by the interaction between individual flies, to be solved efficiently. Thus, a robust method that compares favorably to other existing methods is obtained. PMID:26258779

  17. Hydraulically Driven Grips For Hot Tensile Specimens

    NASA Technical Reports Server (NTRS)

    Bird, R. Keith; Johnson, George W.

    1994-01-01

    Pair of grips for tensile and compressive test specimens operate at temperatures up to 1,500 degrees F. Grips include wedges holding specimen inside furnace, where heated to uniform temperature. Hydraulic pistons drive wedges, causing them to exert clamping force. Hydraulic pistons and hydraulic fluid remain outside furnace, at room temperature. Cooling water flows through parts of grips to reduce heat transferred to external components. Advantages over older devices for gripping specimens in high-temperature tests; no need to drill holes in specimens, maintains constant gripping force on specimens, and heated to same temperature as that of specimen without risk of heating hydraulic fluid and acuator components.

  18. Dynamic Tensile Strength of Coal under Dry and Saturated Conditions

    NASA Astrophysics Data System (ADS)

    Zhao, Yixin; Liu, Shimin; Jiang, Yaodong; Wang, Kai; Huang, Yaqiong

    2016-05-01

    The tensile failure characterization of dry and saturated coals under different impact loading conditions was experimentally investigated using a Split Hopkinson pressure bar. Indirect dynamic Brazilian disc tension tests for coals were carried out. The indirect tensile strengths for different bedding angles under different impact velocities, strain rates and loading rates are analyzed and discussed. A high-speed high-resolution digital camera was employed to capture and record the dynamic failure process of coal specimens. Based on the experimental results, it was found that the saturated specimens have stronger loading rate dependence than the dry specimens. The bedding angle has a smaller effect on the dynamic indirect tensile strength compared to the impact velocity. Both shear and tensile failures were observed in the tested coal specimens. Saturated coal specimens have higher indirect tensile strength than dry ones.

  19. Drying Thermoplastics

    NASA Technical Reports Server (NTRS)

    1976-01-01

    In searching for an improved method of removing water from polyester type resins without damaging the materials, Conair Inc. turned to the NASA Center at the University of Pittsburgh for assistance. Taking an organized, thorough look at existing technology before beginning research has helped many companies save significant time and money. They searched the NASA and other computerized files for microwave drying of thermoplastics. About 300 relevant citations were retrieved - eight of which were identified as directly applicable to the problem. Company estimates it saved a minimum of a full year in compiling research results assembled by the information center.

  20. Can wet roof insulation be dried out

    SciTech Connect

    Tobiasson, W.; Korhonen, C.; Coutermarsh, B.; Greatorex, A.

    1983-01-01

    Nondestructive techniques are being widely used to locate wet insulation in compact roofing systems. Now that wet insulation can be found, breather vents and so-called breathable membranes are being promoted to dry out wet insulation, thereby recovering its thermal effectiveness. Exposure tests in New Hampshire indicate that the above venting methods are all rather ineffective in drying sealed specimens of perlite and fibrous glass roof insulation. It would take many decades to dry our specimens at the rates measured over the past two years. Cross-ventilation within the insulation increased the rate of drying. For perlite insulation, the faster rate would still result in a drying time measured in decades. For fibrous glass insulation, the drying time was reduced to 13 years. Fibrous glass insulation in a roof was dried by removing the water with a vacuum cleaner. In a series of tests with a total duration of 134 h, about 0.4 2 m/sup 3/ (110 gal) of water was removed from a 17-m/sup 2/ (180-ft/sup 2/) area of 38-mm (1.5-in.)-thick insulation. Before the water was removed the insulation had only 21% of its dry insulating ability; afterward it had 83%.

  1. Can wet roof insulation be dried out

    SciTech Connect

    Tobiasson, W.; Coutermarsh, B.; Greatorex, A.; Korhonen, C.

    1981-12-01

    Nondestructive techniques are being widely used to locate wet insulation in compact roofing systems. Now that wet insulation can be found, breather vents and so called ''breathable'' membranes are being promoted to dry out wet insulation, thereby recovering its thermal effectiveness. Exposure tests in New Hampshire indicate that the above venting methods are all rather ineffective in drying sealed specimens of perlite and fibrous glass roof insulation. It would take many decades to dry specimens at the rates measured over the past two years. Cross-ventilation within the insulation increased the rate of drying. For perlite insulation, the faster rate would still result in a drying time measured in decades. For fibrous glass insulation, the drying time was reduced to 13 years. The authors have succeeded in drying fibrous glass insulation in a roof by removing the water with a vacuum cleaner. In a series of tests with a total duration of 134 h, about 0.42 m/sup 3/ (110 gal) of water was removed from a 17-m/sup 2/ (180-ft/sup 2/) area of 38-mm (1.5-in.)-thick insulation. Before the water was removed the insulation had only 21 percent of its dry insulating ability; afterward it had 83 percent.

  2. [Recovery of facultatives and anaerobes from frozen specimens with a polymicrobial nature].

    PubMed

    Kawamura, Chizuko; Nakamura, Toshihiko; Kaimori, Mitsuomi; Watanabe, Kunitomo

    2003-01-01

    Microbiological examination of frozen specimens is sometimes carried out in clinical microbiology and the result is used as an aid of diagnosis and/or treatment of polymicrobial infections. The study was carried out to reevaluate the merit of freezing specimens in clinical microbiology. A total of 10 specimens with a polymicrobial nature were included in this study. Before and after freezing specimens, we isolated facultative and anaerobic bacteria using a set of primary isolation media, consisting of three aerobic agar plates (MacConkey agar, blood agar and chocolate agar) and four pre-reduced anaerobic agar plates (HK Blood agar, HK blood agar with paromomycin (PM) and vancomycin (VM), phenyl ethyl-alcohol (PEA) agar and Bacteroides bile esculin (BBE) agar). All the procedures were done in a properly controlled anaerobic chamber. The number of isolates before and after freezing was 79 and 70, respectively. Among the strains isolated before freezing, 33 strains were recovered on the same kin of media artery freezing, without a remarkable decrease in the quantity. But 26 strains were not recovered and 2 strains were recovered with a remarkable decrease. Among 26 strains, 15 strains could be successfully backed up on the different kind of media. In conclusion, an anaerobic technique with an anaerobic chamber and a set of isolatin plates including blood agar, chocolate agar, HK blood agar, PEA blood agar, HK blood agar with PM and VM enable us to estimate the bacteriology before freezing from frozen specimens. PMID:14984303

  3. Evaluation of the intercept oral specimen collection device with HIV assays versus paired serum/plasma specimens.

    PubMed

    Beelaert, G; Van Heddegem, L; Van Frankenhuijsen, M; Vandewalle, G; Compernolle, V; Florence, E; Fransen, K

    2016-08-01

    Oral fluid has many advantages over blood-based techniques: it is less invasive, eliminates the occupational risk associated with needle stick accidents and collection can be self-administrated. Each individual test is packaged with a corresponding collection device. This study tested the suitability of the Intercept Oral Specimen Collection Device for different HIV diagnostic tests: three different rapid HIV tests and two adapted ELISAs, which were evaluated and compared with a gold standard on blood. In addition a total IgG quantification was performed to demonstrate the quality of the specimen. HIV antibodies were detected with a sensitivity of 100%, 99.3%, 98.6%, 100% and 95.7% for, DPP, OraQuick, Aware, Genscreen and Vironostika respectively using the Intercept Collection Device. Respective specificities were 100%, 100%, 99.3%, 97.3% and 100%. PMID:27142112

  4. Morphine-3-D glucuronide stability in postmortem specimens exposed to bacterial enzymatic hydrolysis.

    PubMed

    Carroll, F T; Marraccini, J V; Lewis, S; Wright, W

    2000-12-01

    Medical examiners frequently rely on the finding of free morphine present in postmortem specimens to assist in certifying deaths associated with narcotics. In vitro hydrolysis of morphine-3-D glucuronide (M3DG) to free morphine was studied using variable specimen pH, initial degree of specimen putrefaction, storage temperature and time, and the effectiveness of sodium fluoride (NaF) preservation. Reagent M3DG was added to opiate-free fresh blood and urine and to autopsy-derived blood specimens. Reagent bovine glucuronidase was also added to certain specimens. Freshly collected and refrigerated NaF-preserved blood produced minimal free morphine, whereas four of five autopsy blood specimens produced free morphine from M3DG. Increased storage time, temperature, and initial degree of putrefaction resulted in greater free morphine generation despite the absence of viable bacteria. Hydrolysis occurring during specimen storage can generate free morphine from M3DG and may result in erroneous conclusions in certifying narcotic deaths. PMID:11111790

  5. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... RULES OF PRACTICE IN TRADEMARK CASES Drawing § 2.56 Specimens. (a) An application under section 1(a) of..., is acceptable. However, a photocopy of the drawing required by § 2.51 is not a proper specimen....

  6. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... RULES OF PRACTICE IN TRADEMARK CASES Drawing § 2.56 Specimens. (a) An application under section 1(a) of..., is acceptable. However, a photocopy of the drawing required by § 2.51 is not a proper specimen....

  7. Testing Biopsy and Cytology Specimens for Cancer

    MedlinePlus

    ... articles window. My Saved Articles » My ACS » Testing Biopsy and Cytology Specimens for Cancer Download Printable Version [ ... on the topics below to get started. Testing Biopsy and Cytology Specimens for Cancer How is cancer ...

  8. Implementation and Operational Research: Programmatic Feasibility of Dried Blood Spots for the Virological Follow-up of Patients on Antiretroviral Treatment in Nord Kivu, Democratic Republic of the Congo

    PubMed Central

    Serrano, Laetitia; Muwonga, Jeremie; Kabuayi, Jean Pierre; Kambale, Alain; Mutaka, Fidèle; Fujiwara, Paula I.; Decosas, Josef; Peeters, Martine; Delaporte, Eric

    2016-01-01

    Background: As part of its policy to shift monitoring of antiretroviral therapy (ART) to primary health care (PHC) workers, the Ministry of Health of the Democratic Republic of Congo (DRC) tested the feasibility of using dried blood spots (DBS) for viral load (VL) quantification and genotypic drug resistance testing in off-site high-throughput laboratories. Methods: DBS samples from adults on ART were collected in 13 decentralized PHC facilities in the Nord-Kivu province and shipped during program quarterly supervision to a reference laboratory 2000 km away, where VL was quantified with a commercial assay (m2000rt, Abbott). A second DBS was sent to a World Health Organization (WHO)-accredited laboratory for repeat VL quantification on a subset of samples with a generic assay (Biocentric) and genotypic drug resistance testing when VL >1000 copies per milliliter. Findings: Constraints arose because of an interruption in national laboratory funding rather than to technical or logistic problems. All samples were assessed by both VL assays to allow ART adjustment. Median DBS turnaround time was 37 days (interquartile range: 9–59). Assays performed unequally with DBS, impacting clinical decisions, quality assurance, and overall cost-effectiveness. Based on m2000rt or generic assay, 31.3% of patients were on virological failure (VF) and 14.8% presented resistance mutations versus 50.3% and 15.4%, respectively. Conclusion: This study confirms that current technologies involving DBS make virological monitoring of ART possible at PHC level, including in challenging environments, provided organizational issues are addressed. Adequate core funding of HIV laboratories and adapted choice of VL assays require urgent attention to control resistance to ART as coverage expands. PMID:26413848

  9. A Simple and Rapid Method Based on Liquid Chromatography-Tandem Mass Spectrometry for the Measurement of α-L-Iduronidase Activity in Dried Blood Spots: An Application to Mucopolysaccharidosis I (Hurler) Screening

    PubMed Central

    Yang, Jeong Soo; Min, Hye Kyeong; Oh, Hyeon Ju; Woo, Hye In; Lee, Soo-Youn; Kim, Jong-Won; Song, Junghan

    2015-01-01

    Background We developed an analytical method to measure α-L-iduronidase (IDUA) activity in dried blood spots. This was achieved by using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) with electrospray ionization in the positive ion mode. Methods Chromatographic separation was completed using mobile phase involving water-formic acid and acetonitrile-formic acid over 2.8 min of run time on a column with a Kinetex XB-C18 (Phenomenex, USA). The detection of column effluent was performed using a Xevo TQ-S triple quadrupole mass spectrometer (Waters, USA) in the multiple-reaction monitoring mode. This method was verified with blank and control samples at four activity levels: base, low, medium, and high. Control materials were provided from Centers for Disease Control and Prevention (CDC). Results Intra- and inter-day precisions were between 2.6% and 16.5% and between 7.9% and 17.0%, respectively. A correlative regression study on the IDUA activity in CDC-control samples performed to assess the validity of the developed method showed a highly significant linear association (r2=0.9976) between the calculated and CDC-reported values and an obvious difference in activity among the four levels. This reliable analytical method was applied to mucopolysaccharidosis I (Hurler) screening of patients under treatment (n=4) and in normal controls (n=129). IDUA activity ranged from 8.98 to 77.12 µmol/hr/L) in normal controls, and patients undergoing medical treatment showed low IDUA activity. Conclusions This method had advantages of simplicity, rapid sample preparation, and liquid chromatographic separation, which efficiently inhibited ionization suppression induced by matrix effects in mass spectrometric detection. PMID:25553279

  10. Blood Disorders

    MedlinePlus

    ... and protein. Over half of your blood is plasma. The solid part of your blood contains red blood cells, white blood cells and platelets. Blood disorders affect one or more parts of the blood and prevent ...

  11. Apparatus for automated testing of biological specimens

    DOEpatents

    Layne, Scott P.; Beugelsdijk, Tony J.

    1999-01-01

    An apparatus for performing automated testing of infections biological specimens is disclosed. The apparatus comprise a process controller for translating user commands into test instrument suite commands, and a test instrument suite comprising a means to treat the specimen to manifest an observable result, and a detector for measuring the observable result to generate specimen test results.

  12. Specimen mass measurement. [during space environment simulation

    NASA Technical Reports Server (NTRS)

    Thornton, W. E.; Ord, J.

    1973-01-01

    The Skylab specimen mass measurement device was operated throughout the altitude test in close simulation of the 56-day Skylab mission. It performed operational specimen measurements well until it was passed out of the chamber for replacement of the specimen hold-down and was autoclaved prior to return. Fecal measurements were typically made with less than one percent error.

  13. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 1 2013-01-01 2013-01-01 false Split specimens. 26.135 Section 26.135 Energy NUCLEAR REGULATORY COMMISSION FITNESS FOR DUTY PROGRAMS Licensee Testing Facilities § 26.135 Split specimens. (a) If the FFD program follows split-specimen procedures, as described in § 26.113, the licensee...

  14. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 1 2012-01-01 2012-01-01 false Split specimens. 26.135 Section 26.135 Energy NUCLEAR REGULATORY COMMISSION FITNESS FOR DUTY PROGRAMS Licensee Testing Facilities § 26.135 Split specimens. (a) If the FFD program follows split-specimen procedures, as described in § 26.113, the licensee...

  15. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 1 2014-01-01 2014-01-01 false Split specimens. 26.135 Section 26.135 Energy NUCLEAR REGULATORY COMMISSION FITNESS FOR DUTY PROGRAMS Licensee Testing Facilities § 26.135 Split specimens. (a) If the FFD program follows split-specimen procedures, as described in § 26.113, the licensee...

  16. Dry Mouth or Xerostomia

    MedlinePlus

    ... or Xerostomia Request Permissions Print to PDF Dry Mouth or Xerostomia Approved by the Cancer.Net Editorial ... a dry mouth. Signs and symptoms of dry mouth The signs and symptoms of dry mouth include ...

  17. Imaging Inhomogeneities From Dry-Coupled Ultrasonic Scans

    NASA Technical Reports Server (NTRS)

    Roth, Don J.

    1995-01-01

    Method of imaging spatial distribution of selected physical properties and microstructure of specimen of material based on dry-coupled contact ultrasonic pulse/echo scanning. Ultrasonic transducer scanned across top surface of specimen. At each of many positions on two-dimensional grid on top surface, ultrasonic pulse/echo measurements taken and processed. Offers rapid, nondestructive alternative to destructive metallographic sectioning to obtain picture of inhomogeneity of specimen.

  18. Further studies of specimen volume changes during processing for SEM: including some plant tissue.

    PubMed

    Boyde, A; Boyde, S

    1980-01-01

    The dimensions of specimens undergoing preparation for examination in the SEM were measured throughout the preparative sequence or at various important stages. The tissues studied included 15-day mouse embryo limbs (MEL), small blocks of adult mouse liver and brain, and potato tuber. The animal tissues were fixed in 3% glutaraldehyde (GA) in 0.15M cacodylate buffer, and the potatoe tissue in 2% GA in water. The effects of various secondary fixation and other treatments were investigated. The results show that lithium salts cause a reduction in the shrinkage of MEL in 100% ethanol but this effect was not significant in the other tissues investigated, and did not persist in specimens stored after critical point drying (CPD). All CPD specimens were shrunken. However postosmication and treatment with uranyl acetate (UAc) and cetyl pyridinium chloride (CPC) all reduced specimen shrinkage in 100% ethanol and after critical point drying. The volume gains with Os + UAc and Os + CPC are both very significant, but it was found that these larger specimens shrank more on storage. Thus rapid examination in the SEM is recommended. Ethanol and Freon 113 were compared as intermediate fluids and it was found that ethanol-CO2 critical point dried specimens shrank more before and after CPD than Freon 113-CO2 specimens. The latter technique is, therefore, to be recommended. Potato tissue shrinks in 30% ethanol, whereas animal tissues all swell in this concentration. The potato tissue also shrank very litte on critical point drying in contrast to the animal tissue specimens. PMID:6999595

  19. Blood sugar test - blood

    MedlinePlus

    ... drink a certain amount of glucose ( oral glucose tolerance test ) How the Test will Feel When the ... a fasting blood glucose, HbA1c test , or glucose tolerance test , depending on your random blood glucose test ...

  20. A novel approach to managing hemolyzed specimens.

    PubMed

    Pretlow, Lester; Johnson, Shamala; Russell, Barbara; Evans, Bridget

    2013-01-01

    Hemolyzed specimens continue to cost the laboratory time and money. However, the core laboratory at Georgia Regents Health System, Inc. has instituted a novel approach to managing this problem. The purpose of this study was to determine whether the laboratory's new approach had a significant impact on the turn-around time (TAT) and cost of processing hemolyzed and non-hemolyzed specimens in the laboratory. The investigators queried the laboratory information systems for hemolyzed and non-hemolyzed specimens categorized as routine or STAT from the core laboratory and calculated statistical differences between the groups with respect to TAT and cost.The investigators found a statistically significant difference in the time it takes to process STAT hemolyzed specimens versus non-hemolyzed specimens with the new approach. Because of the new approach, hemolyzed specimens were actually processed as fast as, or faster than non-hemolyzed specimens in the core laboratory. PMID:23967544

  1. Alfredo Dugès' type specimens of amphibians and reptiles revisited.

    PubMed

    Flores-Villela, Oscar; Ríos-Muñoz, César A; Magaña-Cota, Gloria E; Quezadas-Tapia, Néstor L

    2016-01-01

    The type specimens of amphibians and reptiles of the Museo de Historia Natural Alfredo Dugès, at the University of Guanajuato (MADUG) were reviewed following Smith & Necker's (1943) summary. Owing to this collection's eventful history and its historical importance as the oldest herpetological collection in Mexico, a review of its conservation status was needed. After many years, the collection has received proper recognition at the University of Guanajuato with a portion of the herpetological types considered "Precious Assets" of the university. We found 34 type specimens pertaining to 18 taxa; six are additional specimens to those previously reported; six herpetological types are missing, including the body of the type of Adelophis copei. All specimens are in good to reasonable condition except for the type of Rhinocheilus antonii, which has dried out completely. All specimens are illustrated to show their condition. PMID:27394365

  2. Survival of female Anopheles gambiae Giles through a 9-month dry season in Sudan*

    PubMed Central

    Omer, Salah M.; Cloudsley-Thompson, J. L.

    1970-01-01

    The dry-season biology of a member of the Anopheles gambiae complex (probably species B) was studied in 2 areas in the Khartoum region of Sudan. It was found that in the valley of the White Nile the species maintained itself by low-level breeding, as shown by the continuing presence of larvae, male mosquitos and parous females through the dry months (9 months in the year). In the scattered villages of arid areas situated more than 20 km from the Nile Valley, on the other hand, regular sampling through the cool dry and hot dry months of the year failed to detect any An. gambiae except nulliparous females. These were found in occupied huts, deserted huts, dry wells and animal burrows. The great majority of 213 females collected in the 11 dry months between November 1966 and December 1967 had fresh or older blood-meals but the abdomen was never found fully distended in the dry season. Examination of the ovaries showed that they did not develop beyond Christophers' stage II in the period from November to February, stage III in March and April, or beyond stage IV in May. But, in June and July stage IV and V ovaries predominated and few specimens remained in stage late-II. It is inferred from these observations that the local population of An. gambiae is highly adapted to survive in the adult stage through the severe drought and heat of the arid zone of Sudan. Some feeding activity continues but ovarian development is extremely retarded, and only one batch of eggs matures during the whole 9-month period. Evidence collected in the Nile Valley indicated that female An. gambiae in that area were not subjected to similar retardation of the ovarian cycle; in fact, clear evidence was obtained there of continuous year-round breeding by the mosquito. PMID:5310144

  3. [The German Environmental Specimen Bank].

    PubMed

    Schröter-Kermani, Christa; Gies, Andreas; Kolossa-Gehring, Marike

    2016-03-01

    The main objective of the German Environmental Specimen Bank (ESB) is the long-term storage of environmental and human samples under stable deep-freeze conditions for future research. The ESB is unique in providing a continuous historical record of environmental and human exposure to chemicals in Germany. ESB was started parallel to the development of the first German Chemicals Legislation in the late 1970s. In 1979, the ESB test operation began. After the Chemicals Law came into force in 1982, the ESB was established as a permanent facility in 1985. With the new European Chemicals Legislation, REACH, in 2007 responsibility for the safety of commercial chemicals and risk assessment was assigned to the industry. Since then, the ESB has become even more important in verifying the self-assessment of the industry, in evaluating the effectiveness of regulations, thus ensuring the protection of humans and the environment against adverse effects caused by exposure to chemicals. These objectives are pursued by the regular monitoring of contaminations and the assessment of temporal trends. Demonstrating the necessity of deriving exposure reduction measures, ESB results serve as key information for policy-makers. Information on preventing exposure to chemicals is available to the general public and to the public health services. The ESB is thus an important monitoring instrument of the Federal Ministry for the Environment, Nature Conservation, Building and Nuclear Safety. The Federal Environment Agency operates the ESB based on its own concepts, heads the scientific data evaluation and transfers results into the environmental policy arena and to the general public. PMID:26753867

  4. Vomiting blood

    MedlinePlus

    ... first part of the small intestine, or esophagus Blood clotting disorders Defects in the blood vessels of the ... as a complete blood count (CBC), blood chemistries, blood clotting tests, and liver function tests Esophagogastroduodenoscopy (EGD) (placing ...

  5. Blood pressure

    MedlinePlus Videos and Cool Tools

    Normal blood pressure is important for proper blood flow to the body’s organs and tissues. The force of the blood on the walls of the arteries is called blood pressure. Blood pressure is measured both as the heart ...

  6. Blood transfusions

    MedlinePlus

    ... homologous blood donation. Many communities have a blood bank at which any healthy person can donate blood. ... to arrange with your hospital or local blood bank before your surgery to have directed donor blood. ...

  7. Blood pressure

    MedlinePlus Videos and Cool Tools

    Normal blood pressure is important for proper blood flow to the body’s organs and tissues. The force of the blood on the walls of the arteries is called blood pressure. Blood pressure is measured both ...

  8. Standardizing the Handling of Surgical Specimens.

    PubMed

    Shirey, Cheryl; Perrego, Kristen

    2015-11-01

    To standardize the handling of surgical specimens, the OR clinical educators in a community hospital setting devised a series of departmental changes as a quality improvement project. A newly created skill validation was reviewed in an hour-long educational meeting with all OR staff members. In addition to creating a new annual skill validation, discussions about specimens were included in the hand over, the time out, and a newly instituted debriefing tool to be used toward the end of a procedure. This interdisciplinary group devised interventions to improve the process of handling specimens such as standardizing the labeling process and changing the transparency of the specimen container. The goal was to assure standardization of specimen handling, specifically to assist novice staff members, and to harmonize inconsistencies between specialties within the practice of existing staff members. These combined methods helped to ensure accurate communication and procurement of specimens for all procedures. PMID:26514715

  9. Flat tensile specimen design for advanced composites

    NASA Technical Reports Server (NTRS)

    Worthem, Dennis W.

    1990-01-01

    Finite element analyses of flat, reduced gage section tensile specimens with various transition region contours were performed. Within dimensional constraints, such as maximum length, tab region width, gage width, gage length, and minimum tab length, a transition contour radius of 41.9 cm produced the lowest stress values in the specimen transition region. The stresses in the transition region were not sensitive to specimen material properties. The stresses in the tab region were sensitive to specimen composite and/or tab material properties. An evaluation of stresses with different specimen composite and tab material combinations must account for material nonlinearity of both the tab and the specimen composite. Material nonlinearity can either relieve stresses in the composite under the tab or elevate them to cause failure under the tab.

  10. Characterizing DNA preservation in degraded specimens of Amara alpina (Carabidae: Coleoptera).

    PubMed

    Heintzman, Peter D; Elias, Scott A; Moore, Karen; Paszkiewicz, Konrad; Barnes, Ian

    2014-05-01

    DNA preserved in degraded beetle (Coleoptera) specimens, including those derived from dry-stored museum and ancient permafrost-preserved environments, could provide a valuable resource for researchers interested in species and population histories over timescales from decades to millenia. However, the potential of these samples as genetic resources is currently unassessed. Here, using Sanger and Illumina shotgun sequence data, we explored DNA preservation in specimens of the ground beetle Amara alpina, from both museum and ancient environments. Nearly all museum specimens had amplifiable DNA, with the maximum amplifiable fragment length decreasing with age. Amplification of DNA was only possible in 45% of ancient specimens. Preserved mitochondrial DNA fragments were significantly longer than those of nuclear DNA in both museum and ancient specimens. Metagenomic characterization of extracted DNA demonstrated that parasite-derived sequences, including Wolbachia and Spiroplasma, are recoverable from museum beetle specimens. Ancient DNA extracts contained beetle DNA in amounts comparable to museum specimens. Overall, our data demonstrate that there is great potential for both museum and ancient specimens of beetles in future genetic studies, and we see no reason why this would not be the case for other orders of insect. PMID:24266987

  11. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 36 Parks, Forests, and Public Property 3 2014-07-01 2014-07-01 false Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... representative of a reputable scientific or educational institution or a State or Federal agency for the...

  12. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 3 2012-07-01 2012-07-01 false Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... representative of a reputable scientific or educational institution or a State or Federal agency for the...

  13. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 3 2013-07-01 2012-07-01 true Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... representative of a reputable scientific or educational institution or a State or Federal agency for the...

  14. 36 CFR 2.5 - Research specimens.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Research specimens. 2.5 Section 2.5 Parks, Forests, and Public Property NATIONAL PARK SERVICE, DEPARTMENT OF THE INTERIOR RESOURCE PROTECTION, PUBLIC USE AND RECREATION § 2.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in accordance with...

  15. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... shall store Bottles A and B of the specimen in a secure manner until the facility has finished testing. If the initial validity and drug test results are negative and the specimen in Bottle A will not be forwarded to the HHS-certified laboratory, the licensee testing facility may discard both Bottle A...

  16. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... shall store Bottles A and B of the specimen in a secure manner until the facility has finished testing. If the initial validity and drug test results are negative and the specimen in Bottle A will not be forwarded to the HHS-certified laboratory, the licensee testing facility may discard both Bottle A...

  17. Machining technique prevents undercutting in tensile specimens

    NASA Technical Reports Server (NTRS)

    Moscater, R. E.; Royster, D. M.

    1968-01-01

    Machining technique prevents undercutting at the test section in tensile specimens when machining the four corners of the reduced section. Made with a gradual taper in the test section, the width of the center of the tensile specimen is less than the width at the four corners of the reduced section.

  18. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... advertising of the services in commerce. (b)(1) A trademark specimen is a label, tag, or container for the... goods, or in the sale or advertising of the services, is acceptable. However, a photocopy of the drawing... specimen that meets the requirements of the rule (i.e., is flat and no larger than 81/2 inches (21.6...

  19. 37 CFR 1.166 - Specimens.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... COMMERCE GENERAL RULES OF PRACTICE IN PATENT CASES National Processing Provisions Plant Patents § 1.166 Specimens. The applicant may be required to furnish specimens of the plant, or its flower or fruit, in a quantity and at a time in its stage of growth as may be designated, for study and inspection....

  20. 37 CFR 1.166 - Specimens.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... COMMERCE GENERAL RULES OF PRACTICE IN PATENT CASES National Processing Provisions Plant Patents § 1.166 Specimens. The applicant may be required to furnish specimens of the plant, or its flower or fruit, in a quantity and at a time in its stage of growth as may be designated, for study and inspection....

  1. 37 CFR 1.166 - Specimens.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2014-07-01 2014-07-01 false Specimens. 1.166 Section 1.166 Patents, Trademarks, and Copyrights UNITED STATES PATENT AND TRADEMARK OFFICE, DEPARTMENT OF COMMERCE GENERAL RULES OF PRACTICE IN PATENT CASES National Processing Provisions Plant Patents § 1.166 Specimens. The applicant may be required to...

  2. Making Ceramic Reference Specimens Containing Seeded Voids

    NASA Technical Reports Server (NTRS)

    Baaklini, George Y.; Klima, Stanley J.; Roth, Don J.

    1994-01-01

    Internal and surface voids of known sizes incorporated into silicon carbide and silicon nitride ceramic reference specimens at prescribed locations. Specimens used to demonstrate sensitivity and resolution in nondestructive examination techniques like scanning laser acoustic microscopy and x-radiography, and to assist in establishing proper examination procedures.

  3. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 3 2010-07-01 2010-07-01 false Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... of research, baseline inventories, monitoring, impact analysis, group study, or museum display...

  4. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 3 2011-07-01 2011-07-01 false Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... of research, baseline inventories, monitoring, impact analysis, group study, or museum display...

  5. Layered Plating Specimens For Mechanical Tests

    NASA Technical Reports Server (NTRS)

    Thompson, Linda B.; Flowers, Cecil E.

    1991-01-01

    Layered specimens readily made in standard sizes for tensile and other tests of mechanical properties. Standard specimen of metal ordinarily difficult to plate to standard grip thickness or diameter made by augmentation with easier-to-plate material followed by machining to standard size and shape.

  6. Electron microprobe analysis program for biological specimens: BIOMAP

    NASA Technical Reports Server (NTRS)

    Edwards, B. F.

    1972-01-01

    BIOMAP is a Univac 1108 compatible program which facilitates the electron probe microanalysis of biological specimens. Input data are X-ray intensity data from biological samples, the X-ray intensity and composition data from a standard sample and the electron probe operating parameters. Outputs are estimates of the weight percentages of the analyzed elements, the distribution of these estimates for sets of red blood cells and the probabilities for correlation between elemental concentrations. An optional feature statistically estimates the X-ray intensity and residual background of a principal standard relative to a series of standards.

  7. Numbers and types of anaerobic bacteria isolated from clinical specimens since 1960.

    PubMed Central

    Holland, J W; Hill, E O; Altemeier, W A

    1977-01-01

    Between 1960 and 1974, 826 specimens, excluding stool, urine, sputum, and blood, yielded 689 (83%) positive cultures, of which 403 (58.5%) contained anaerobic bacteria. This represents 48.8% of the total specimens cultured. Isolates from 153 specimens obtained and stocked from 1965 to 1974 were reidentified by current criteria. Gram-negative bacilli, primarily bacteroides, were the most frequently isolated anaerobes, being found in 70% of 153 anaerobe-positive specimens and accounting for 42% of the total anaerobes isolated. Gram-positive cocci were second in occurrence, being found in 66% of 153 specimens and accounting for 40% of the total isolates. Bacteroides fragilis was by far the most frequently isolated species. Compairson of 14 years of cumulative data with data from current studies covering 1- to 2-year periods indicated that the anaerobes isolated from clinical material have not changed significantly in type or relative numbers. PMID:833266

  8. Donating Blood

    MedlinePlus

    ... can give blood every 56 days. Before Donating Blood donation starts before you walk in the door of ... regenerate the red blood cells lost during a blood donation. An iron-fortified diet plus daily iron tablets ...

  9. Blood culture

    MedlinePlus

    Culture - blood ... A blood sample is needed . The site where blood will be drawn is first cleaned with an antiseptic such ... organism from the skin getting into (contaminating) the blood sample and causing a false-positive result (see ...

  10. Blood Thinners

    MedlinePlus

    If you have some kinds of heart or blood vessel disease, or if you have poor blood flow to your brain, your doctor may recommend that you take a blood thinner. Blood thinners reduce the risk of heart ...

  11. High-intensity drying processes: Impulse drying

    SciTech Connect

    Orloff, D.I.

    1989-05-01

    Impulse drying is an innovative process for drying paper that holds great promise for reducing the energy consumed during manufacture of paper and similar web products. Impulse drying occurs when a wet paper web passes through a press nip where one of the rolls is heated to a very high temperature. Steam generated by contact with the hot roll expands and displaces water from the sheet in a very efficient manner. The energy required for water removal is much lower than that required for conventional evaporative drying. Tests have been completed that elucidate the unique displacement mechanism of water removal in the impulse drying process. A pilot roll press has been designed, installed and used to examine impulse drying under conditions that simulate commercial press conditions. The results of this earlier work have been reported in three previous reports. During this report period October, 1987 to September, 1988, the pilot press was equipped with a second impulse drying roll to facilitate studies of surface uniformity in impulse dried paper. Studies have also been completed which examine the origins of sheet delamination that has been been encountered during impulse drying of certain heavyweight paper grades, and which investigate approaches to prevent delamination in these grades. Finally, an experimental plan has been formalized to examine impulse drying of lightweight grades which are candidates for early commercialization. 7 refs., 30 figs., 3 tabs.

  12. Optimal design of biaxial tensile cruciform specimens

    NASA Astrophysics Data System (ADS)

    Demmerle, S.; Boehler, J. P.

    1993-01-01

    F OR EXPERIMENTAL investigations concerning the mechanical behaviour under biaxial stress states of rolled sheet metals, mostly cruciform flat specimens are used. By means of empirical methods, different specimen geometries have been proposed in the literature. In order to evaluate the suitability of a specimen design, a mathematically well defined criterion is developed, based on the standard deviations of the values of the stresses in the test section. Applied to the finite element method, the criterion is employed to realize the shape optimization of biaxial cruciform specimens for isotropic elastic materials. Furthermore, the performance of the obtained optimized specimen design is investigated in the case of off-axes tests on anisotropic materials. Therefore, for the first time, an original testing device, consisting of hinged fixtures with knife edges at each arm of the specimen, is applied to the biaxial test. The obtained results indicate the decisive superiority of the optimized specimens for the proper performance on isotropic materials, as well as the paramount importance of the proposed off-axes testing technique for biaxial tests on anisotropic materials.

  13. Blood sugar test - blood

    MedlinePlus

    ... in the way you normally talk or behave Fainting spells Seizures (for the first time) SCREENING FOR ... drawn are slight, but may include: Excessive bleeding Fainting or feeling lightheaded Hematoma (blood accumulating under the ...

  14. National Aeronautics and Space Administration Biological Specimen Repository

    NASA Technical Reports Server (NTRS)

    McMonigal, Kathleen A.; Pietrzyk, Robert a.; Johnson, Mary Anne

    2008-01-01

    The National Aeronautics and Space Administration Biological Specimen Repository (Repository) is a storage bank that is used to maintain biological specimens over extended periods of time and under well-controlled conditions. Samples from the International Space Station (ISS), including blood and urine, will be collected, processed and archived during the preflight, inflight and postflight phases of ISS missions. This investigation has been developed to archive biosamples for use as a resource for future space flight related research. The International Space Station (ISS) provides a platform to investigate the effects of microgravity on human physiology prior to lunar and exploration class missions. The storage of crewmember samples from many different ISS flights in a single repository will be a valuable resource with which researchers can study space flight related changes and investigate physiological markers. The development of the National Aeronautics and Space Administration Biological Specimen Repository will allow for the collection, processing, storage, maintenance, and ethical distribution of biosamples to meet goals of scientific and programmatic relevance to the space program. Archiving of the biosamples will provide future research opportunities including investigating patterns of physiological changes, analysis of components unknown at this time or analyses performed by new methodologies.

  15. Blood Vessel Tension Tester

    NASA Technical Reports Server (NTRS)

    1978-01-01

    In the photo, a medical researcher is using a specially designed laboratory apparatus for measuring blood vessel tension. It was designed by Langley Research Center as a service to researchers of Norfolk General Hospital and Eastern Virginia Medical School, Norfolk, Virginia. The investigators are studying how vascular smooth muscle-muscle in the walls of blood vessels-reacts to various stimulants, such as coffee, tea, alcohol or drugs. They sought help from Langley Research Center in devising a method of measuring the tension in blood vessel segments subjected to various stimuli. The task was complicated by the extremely small size of the specimens to be tested, blood vessel "loops" resembling small rubber bands, some only half a millimeter in diameter. Langley's Instrumentation Development Section responded with a miniaturized system whose key components are a "micropositioner" for stretching a length of blood vessel and a strain gage for measuring the smooth muscle tension developed. The micropositioner is a two-pronged holder. The loop of Mood vessel is hooked over the prongs and it is stretched by increasing the distance between the prongs in minute increments, fractions of a millimeter. At each increase, the tension developed is carefully measured. In some experiments, the holder and specimen are lowered into the test tubes shown, which contain a saline solution simulating body fluid; the effect of the compound on developed tension is then measured. The device has functioned well and the investigators say it has saved several months research time.

  16. Virtual blood bank

    PubMed Central

    Wong, Kit Fai

    2011-01-01

    Virtual blood bank is the computer-controlled, electronically linked information management system that allows online ordering and real-time, remote delivery of blood for transfusion. It connects the site of testing to the point of care at a remote site in a real-time fashion with networked computers thus maintaining the integrity of immunohematology test results. It has taken the advantages of information and communication technologies to ensure the accuracy of patient, specimen and blood component identification and to enhance personnel traceability and system security. The built-in logics and process constraints in the design of the virtual blood bank can guide the selection of appropriate blood and minimize transfusion risk. The quality of blood inventory is ascertained and monitored, and an audit trail for critical procedures in the transfusion process is provided by the paperless system. Thus, the virtual blood bank can help ensure that the right patient receives the right amount of the right blood component at the right time. PMID:21383930

  17. Virtual blood bank.

    PubMed

    Wong, Kit Fai

    2011-01-01

    Virtual blood bank is the computer-controlled, electronically linked information management system that allows online ordering and real-time, remote delivery of blood for transfusion. It connects the site of testing to the point of care at a remote site in a real-time fashion with networked computers thus maintaining the integrity of immunohematology test results. It has taken the advantages of information and communication technologies to ensure the accuracy of patient, specimen and blood component identification and to enhance personnel traceability and system security. The built-in logics and process constraints in the design of the virtual blood bank can guide the selection of appropriate blood and minimize transfusion risk. The quality of blood inventory is ascertained and monitored, and an audit trail for critical procedures in the transfusion process is provided by the paperless system. Thus, the virtual blood bank can help ensure that the right patient receives the right amount of the right blood component at the right time. PMID:21383930

  18. BIOMASS DRYING TECHNOLOGIES

    EPA Science Inventory

    The report examines the technologies used for drying of biomass and the energy requirements of biomass dryers. Biomass drying processes, drying methods, and the conventional types of dryers are surveyed generally. Drying methods and dryer studies using superheated steam as the d...

  19. Specimen for high-temperature tensile tests

    NASA Technical Reports Server (NTRS)

    Coulbert, C. D.

    1972-01-01

    Split nut with internal taper to hold specially formed specimen composed of filaments of refractory material provides means for holding at high temperature and under tension so that performance evaluations may be made.

  20. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... specimen”), the Office will create a digital facsimile of the specimen that meets the requirements of the...-bulky alternatives, the Office may accept an audio or video cassette tape recording, CD-ROM, or...

  1. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... exceeding these size requirements (a “bulky specimen”), the Office will create a digital facsimile of the... bulky specimen. (3) In the absence of non-bulky alternatives, the Office may accept an audio or...

  2. A Live Specimen Cell for the Microscope.

    ERIC Educational Resources Information Center

    McNeil, D. W.

    1991-01-01

    Provides background and instructions for the assembly of a microaquarium, or specimen cell, in which the dynamic world of living microorganisms can be viewed through a microscope overextended periods of time utilizing the simplest of materials in the process. (JJK)

  3. Impact of specimen adequacy on the assessment of renal allograft biopsy specimens

    PubMed Central

    Cimen, S.; Geldenhuys, L.; Guler, S.; Imamoglu, A.; Molinari, M.

    2016-01-01

    The Banff classification was introduced to achieve uniformity in the assessment of renal allograft biopsies. The primary aim of this study was to evaluate the impact of specimen adequacy on the Banff classification. All renal allograft biopsies obtained between July 2010 and June 2012 for suspicion of acute rejection were included. Pre-biopsy clinical data on suspected diagnosis and time from renal transplantation were provided to a nephropathologist who was blinded to the original pathological report. Second pathological readings were compared with the original to assess agreement stratified by specimen adequacy. Cohen's kappa test and Fisher's exact test were used for statistical analyses. Forty-nine specimens were reviewed. Among these specimens, 81.6% were classified as adequate, 6.12% as minimal, and 12.24% as unsatisfactory. The agreement analysis among the first and second readings revealed a kappa value of 0.97. Full agreement between readings was found in 75% of the adequate specimens, 66.7 and 50% for minimal and unsatisfactory specimens, respectively. There was no agreement between readings in 5% of the adequate specimens and 16.7% of the unsatisfactory specimens. For the entire sample full agreement was found in 71.4%, partial agreement in 20.4% and no agreement in 8.2% of the specimens. Statistical analysis using Fisher's exact test yielded a P value above 0.25 showing that - probably due to small sample size - the results were not statistically significant. Specimen adequacy may be a determinant of a diagnostic agreement in renal allograft specimen assessment. While additional studies including larger case numbers are required to further delineate the impact of specimen adequacy on the reliability of histopathological assessments, specimen quality must be considered during clinical decision making while dealing with biopsy reports based on minimal or unsatisfactory specimens. PMID:27119314

  4. Mode 2 fatigue crack growth specimen development

    NASA Technical Reports Server (NTRS)

    Buzzard, R. J.; Gross, B.; Srawley, J. E.

    1983-01-01

    A Mode II test specimen was developed which has potential application in understanding phemonena associated with mixed mode fatigue failures in high performance aircraft engine bearing races. The attributes of the specimen are: it contains one single ended notch, which simplifiers data gathering and reduction; the fatigue crack grous in-line with the direction of load application; a single axis test machine is sufficient to perform testing; and the Mode I component is vanishingly small.

  5. A curved beam test specimen for determining the interlaminar tensile strength of a laminated composite

    NASA Technical Reports Server (NTRS)

    Hiel, Clement C.; Sumich, Mark; Chappell, David P.

    1990-01-01

    A curved beam type of test specimen is evaluated for use in determining the through-the-thickness strength of laminated composites. Two variations of a curved beam specimen configuration (semi-circular and elliptical) were tested to failure using static and fatigue loads. The static failure load for the semi-circular specimens was found to be highly sensitive to flaw content, with the specimens falling into two distinct groups. This result supports the use of proof testing for structural validation. Static design allowables are derived based on the Weibull distribution. Fatigue data indicates no measured increase in specimen compliance prior to final fracture. All static and fatigue failures at room temperature dry conditions occurred catastrophically. The elliptical specimens demonstrated unusually high failure strengths indicating the presence of phenomena requiring further study. Results are also included for specimens exposed to a wet environment showing a matrix strength degradation due to moisture content. Further testing is under way to evaluate a fatigue methodology for matrix dominated failures based on residual static strength (wearout).

  6. A curved beam test specimen for determining the interlaminar tensile strength of a laminated composite

    NASA Technical Reports Server (NTRS)

    Hiel, Clement C.; Sumich, Mark; Chappell, David P.

    1991-01-01

    A curved beam type of test specimen is evaluated for use in determining the through-the-thickness strength of laminated composites. Two variations of a curved beam specimen configuration (semicircular and elliptical) were tested to failure using static and fatigue loads. The static failure load for the semicircular specimens was found to be highly sensitive to flaw content, with the specimens falling into two distinct groups. This result supports the use of proof testing for structural validation. Static design allowables are derived based on the Weibull distribution. Fatigue data indicates no measured increase in specimen compliance prior to final fracture. All static and fatigue failures at room temperature dry conditions occurred catastrophically. The elliptical specimens demonstrated unusually high failure strengths indicating the presence of phenomena requiring further study. Results are also included for specimens exposed to a wet environment showing a matrix strength degradation due to moisture content. Further testing is underway to evaluate a fatigue methodology for matrix dominated failures based on residual static strength (wearout).

  7. A curved beam test specimen for determining the interlaminar tensile strength of a laminated composite

    SciTech Connect

    Hiel, C.C.; Sumich, M.; Chappell, D.P. )

    1991-07-01

    A curved beam type of test specimen is evaluated for use in determining the through-the-thickness strength of laminated composites. Two variations of a curved beam specimen configuration (semicircular and elliptical) were tested to failure using static and fatigue loads. The static failure load for the semicircular specimens was found to be highly sensitive to flaw content, with the specimens falling into two distinct groups. This result supports the use of proof testing for structural validation. Static design allowables are derived based on the Weibull distribution. Fatigue data indicates no measured increase in specimen compliance prior to final fracture. All static and fatigue failures at room temperature dry conditions occurred catastrophically. The elliptical specimens demonstrated unusually high failure strengths indicating the presence of phenomena requiring further study. Results are also included for specimens exposed to a wet environment showing a matrix strength degradation due to moisture content. Further testing is underway to evaluate a fatigue methodology for matrix dominated failures based on residual static strength (wearout). 10 refs.

  8. Blood drop patterns: Formation and applications.

    PubMed

    Chen, Ruoyang; Zhang, Liyuan; Zang, Duyang; Shen, Wei

    2016-05-01

    The drying of a drop of blood or plasma on a solid substrate leads to the formation of interesting and complex patterns. Inter- and intra-cellular and macromolecular interactions in the drying plasma or blood drop are responsible for the final morphologies of the dried patterns. Changes in these cellular and macromolecular components in blood caused by diseases have been suspected to cause changes in the dried drop patterns of plasma and whole blood, which could be used as simple diagnostic tools to identify the health of humans and livestock. However, complex physicochemical driving forces involved in the pattern formation are not fully understood. This review focuses on the scientific development in microscopic observations and pattern interpretation of dried plasma and whole blood samples, as well as the diagnostic applications of pattern analysis. Dried drop patterns of plasma consist of intricate visible cracks in the outer region and fine structures in the central region, which are mainly influenced by the presence and concentration of inorganic salts and proteins during drying. The shrinkage of macromolecular gel and its adhesion to the substrate surface have been thought to be responsible for the formation of the cracks. Dried drop patterns of whole blood have three characteristic zones; their formation as functions of drying time has been reported in the literature. Some research works have applied engineering treatment to the evaporation process of whole blood samples. The sensitivities of the resultant patterns to the relative humidity of the environment, the wettability of the substrates, and the size of the drop have been reported. These research works shed light on the mechanisms of spreading, evaporation, gelation, and crack formation of the blood drops on solid substrates, as well as on the potential applications of dried drop patterns of plasma and whole blood in diagnosis. PMID:26988066

  9. Human Blood Typing: A Forensic Science Approach. Part I: Background.

    ERIC Educational Resources Information Center

    Kobilinsky, Lawrence; Sheehan, Francis X.

    1988-01-01

    In this article, part I of a series, the forensic methods used in "typing" human blood, which as physical evidence is often found in the dried state, are outlined. Background information about individualization, antibody typing, fresh blood, dried blood, and additional systems is provided. (CW)

  10. Solution Preserves Nucleic Acids in Body-Fluid Specimens

    NASA Technical Reports Server (NTRS)

    Pierson, Duane L.; Stowe, Raymond P.

    2004-01-01

    A solution has been formulated to preserve deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) in specimens of blood, saliva, and other bodily fluids. Specimens of this type are collected for diagnostic molecular pathology, which is becoming the method of choice for diagnosis of many diseases. The solution makes it possible to store such specimens at room temperature, without risk of decomposition, for subsequent analysis in a laboratory that could be remote from the sampling location. Thus, the solution could be a means to bring the benefits of diagnostic molecular pathology to geographic regions where refrigeration equipment and diagnostic laboratories are not available. The table lists the ingredients of the solution. The functions of the ingredients are the following: EDTA chelates divalent cations that are necessary cofactors for nuclease activity. In so doing, it functionally removes these cations and thereby retards the action of nucleases. EDTA also stabilizes the DNA helix. Tris serves as a buffering agent, which is needed because minor contaminants in an unbuffered solution can exert pronounced effects on pH and thereby cause spontaneous degradation of DNA. SDS is an ionic detergent that inhibits ribonuclease activity. SDS has been used in some lysis buffers and as a storage buffer for RNA after purification. The use of the solution is straightforward. For example, a sample of saliva is collected by placing a cotton roll around in the subject's mouth until it becomes saturated, then the cotton is placed in a collection tube. Next, 1.5 mL of the solution are injected directly into the cotton and the tube is capped for storage at room temperature. The effectiveness of the solution has been demonstrated in tests on specimens of saliva containing herpes simplex virus. In the tests, the viral DNA, as amplified by polymerase chain reaction, was detected even after storage for 120 days.

  11. STRESS CORROSION CRACKING IN TEAR DROP SPECIMENS

    SciTech Connect

    Lam, P; Philip Zapp, P; Jonathan Duffey, J; Kerry Dunn, K

    2009-05-01

    Laboratory tests were conducted to investigate the stress corrosion cracking (SCC) of 304L stainless steel used to construct the containment vessels for the storage of plutonium-bearing materials. The tear drop corrosion specimens each with an autogenous weld in the center were placed in contact with moist plutonium oxide and chloride salt mixtures. Cracking was found in two of the specimens in the heat affected zone (HAZ) at the apex area. Finite element analysis was performed to simulate the specimen fabrication for determining the internal stress which caused SCC to occur. It was found that the tensile stress at the crack initiation site was about 30% lower than the highest stress which had been shifted to the shoulders of the specimen due to the specimen fabrication process. This finding appears to indicate that the SCC initiation took place in favor of the possibly weaker weld/base metal interface at a sufficiently high level of background stress. The base material, even subject to a higher tensile stress, was not cracked. The relieving of tensile stress due to SCC initiation and growth in the HAZ and the weld might have foreclosed the potential for cracking at the specimen shoulders where higher stress was found.

  12. Dry mouth during cancer treatment

    MedlinePlus

    Chemotherapy - dry mouth; Radiation therapy - dry mouth; Transplant - dry mouth; Transplantation - dry mouth ... Some cancer treatments and medicines can cause dry mouth. Symptoms you may have include: Mouth sores Thick ...

  13. Dried-Plasma Transport Using a Novel Matrix and Collection System for Human Immunodeficiency Virus and Hepatitis C Virus Virologic Testing▿

    PubMed Central

    Lloyd, R. M.; Burns, D. A.; Huong, J. T.; Mathis, R. L.; Winters, M. A.; Tanner, M.; De La Rosa, A.; Yen-Lieberman, B.; Armstrong, W.; Taege, A.; McClernon, D. R.; Wetshtein, J. L.; Friedrich, Brian M.; Ferguson, Monique R.; O'Brien, William; Feorino, P. M.; Holodniy, M.

    2009-01-01

    A novel method for the collection and transportation of dried-blood-plasma samples, SampleTanker (ST), was developed and compared to standard shipping protocols for frozen-plasma specimens containing human immunodeficiency virus type 1 (HIV-1) and/or hepatitis C virus (HCV). Matched frozen and dried 1-ml EDTA-containing plasma samples were collected and analyzed by several molecular-based virologic assays. After addition of 1.175 ml of reconstitution buffer, 1.035 ml of dried plasma was recovered. Mean intra-assay variances were 0.05, 0.05, and 0.06 log10 copies/ml for the Versant, Amplicor, and NucliSens QT HIV-1 load assays, respectively (P, not significant). However, mean HIV-1 viral load was consistently reduced in dried samples by 0.32 to 0.51 log10 copies/ml, depending on assay type (P < 0.05). Infectious HIV-1 was not recovered from dried ST plasma. There was no significant difference in HIV-1 viral load results obtained using ST after 8 weeks of storage at ambient temperature. Compared to frozen plasma, HIV-1 genotypic results were >99% concordant at the nucleotide and amino acid levels, as well as for resistance-associated mutations. We further demonstrated successful detection of multiple analytes, including HIV-1 viral load, HIV-1 antiretroviral resistance genotype, and HCV genotype, from a single ST unit. Dried plasma collected with ST yielded comparable results to frozen samples for multiple-analyte clinical testing. As such, ST could be a useful alternative for virologic tests and clinical trials worldwide by significantly diminishing transportation cost and the sample volume restrictions associated with dried-blood-spot technology. PMID:19321732

  14. Blood typing

    MedlinePlus

    ... typing. The liquid part of your blood without cells (serum) is mixed with blood that is known to be type ... ABO typing: If your blood cells stick together when mixed with: Anti-A serum, you have type A blood Anti-B serum, you have type B blood Both anti-A and ...

  15. Closeout of JOYO-1 Specimen Fabrication Efforts

    SciTech Connect

    ME Petrichek; JL Bump; RF Luther

    2005-10-31

    Fabrication was well under way for the JOYO biaxial creep and tensile specimens when the NR Space program was canceled. Tubes of FS-85, ASTAR-811C, and T-111 for biaxial creep specimens had been drawn at True Tube (Paso Robles, CA), while tubes of Mo-47.5 Re were being drawn at Rhenium Alloys (Cleveland, OH). The Mo-47.5 Re tubes are now approximately 95% complete. Their fabrication and the quantities produced will be documented at a later date. End cap material for FS-85, ASTAR-811C, and T-111 had been swaged at Pittsburgh Materials Technology, Inc. (PMTI) (Large, PA) and machined at Vangura (Clairton, PA). Cutting of tubes, pickling, annealing, and laser engraving were in process at PMTI. Several biaxial creep specimen sets of FS-85, ASTAR-811C, and T-111 had already been sent to Pacific Northwest National Laboratory (PNNL) for weld development. In addition, tensile specimens of FS-85, ASTAR-811C, T-111, and Mo-47.5 Re had been machined at Kin-Tech (North Huntington, PA). Actual machining of the other specimen types had not been initiated. Flowcharts 1-3 detail the major processing steps each piece of material has experienced. A more detailed description of processing will be provided in a separate document [B-MT(SRME)-51]. Table 1 lists the in-process materials and finished specimens. Also included are current metallurgical condition of these materials and specimens. The available chemical analyses for these alloys at various points in the process are provided in Table 2.

  16. Application of PCR to multiple specimen types for diagnosis of cytomegalovirus infection: comparison with cell culture and shell vial assay.

    PubMed

    Miller, M J; Bovey, S; Pado, K; Bruckner, D A; Wagar, E A

    1994-01-01

    Human cytomegalovirus (CMV) is a herpesvirus that is responsible for significant morbidity and mortality in congenitally infected infants and immunocompromised patients. Antiviral therapies are available, thus making timely diagnosis of significant importance to at-risk patients. A PCR system was devised. The newly devised system, unlike previously described systems, can be applied to a wide variety of specimen types in a clinical microbiology laboratory setting. Specimens from all sites routinely accepted for CMV culture were shown to be acceptable for CMV PCR. Sensitivity and specificity were established in comparison with those of both monolayer culture and shell vial assay (SVA). The sensitivity and specificity of PCR for detection of CMV in specimens exclusive of urine and blood were 97.5 (77 of 79 specimens) and 87.2% (41 of 47 specimens), respectively. The sensitivity and specificity of PCR for urine and blood specimens were 100 (10 of 10) and 95.7% (45 of 47) and 66.7 (4 of 6) and 78.8% (41 of 52), respectively. Discrepancies of positive PCR results with negative culture or SVA results occurred for specimens flanked chronologically by other culture- or SVA-positive specimens and were likely culture failures, increasing the specificity (100%) of PCR. Discrepancies of negative PCR results with positive culture or SVA results occurred in specimens with few cells or infectious foci by SVA or culture and may represent sampling variability associated with low virus titers. PMID:8126204

  17. Investigations in space-related molecular biology. [cryo-electron microscopic and diffraction studies on terrestrial and extraterrestrial specimens

    NASA Technical Reports Server (NTRS)

    Fernandez-Moran, H.; Pritzker, A. N.

    1974-01-01

    Improved instrumentation and preparation techniques for high resolution, high voltage cryo-electron microscopic and diffraction studies on terrestrial and extraterrestrial specimens are reported. Computer correlated ultrastructural and biochemical work on hydrated and dried cell membranes and related biological systems provided information on membrane organization, ice crystal formation and ordered water, RNA virus linked to cancer, lunar rock samples, and organometallic superconducting compounds. Apollo 11, 12, 14, and 15 specimens were analyzed

  18. Characterization of some biological specimens using TEM and SEM

    NASA Astrophysics Data System (ADS)

    Ghosh, Nabarun; Smith, Don W.

    2009-05-01

    The advent of novel techniques using the Transmission and Scanning Electron Microscopes improved observation on various biological specimens to characterize them. We studied some biological specimens using Transmission and Scanning Electron Microscopes. We followed negative staining technique with Phosphotungstic acid using bacterial culture of Bacillus subtilis. Negative staining is very convenient technique to view the structural morphology of different samples including bacteria, phage viruses and filaments in a cell. We could observe the bacterial cell wall and flagellum very well when trapped the negative stained biofilm from bacterial culture on a TEM grid. We cut ultra thin sections from the fixed root tips of Pisum sativum (Garden pea). Root tips were pre fixed with osmium tetroxide and post fixed with uranium acetate and placed in the BEEM capsule for block making. The ultrathin sections on the grid under TEM showed the granular chromatin in the nucleus. The protein bodies and large vacuoles with the storage materials were conspicuous. We followed fixation, critical point drying and sputter coating with gold to view the tissues with SEM after placing on stubs. SEM view of the leaf surface of a dangerous weed Tragia hispida showed the surface trichomes. These trichomes when break on touching releases poisonous content causing skin irritation. The cultured tissue from in vitro culture of Albizia lebbeck, a tree revealed the regenerative structures including leaf buds and stomata on the tissue surface. SEM and TEM allow investigating the minute details characteristic morphological features that can be used for classroom teaching.

  19. 46 CFR 4.06-50 - Specimen analysis and follow-up procedures.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... sent to the Medical Review Officer meeting the requirements of 49 CFR 40.121, as designated by the... required by 49 CFR part 40, subpart G, and submit his or her findings to the marine employer. Blood test... 46 Shipping 1 2013-10-01 2013-10-01 false Specimen analysis and follow-up procedures....

  20. 46 CFR 4.06-50 - Specimen analysis and follow-up procedures.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... sent to the Medical Review Officer meeting the requirements of 49 CFR 40.121, as designated by the... required by 49 CFR part 40, subpart G, and submit his or her findings to the marine employer. Blood test... 46 Shipping 1 2014-10-01 2014-10-01 false Specimen analysis and follow-up procedures....

  1. 46 CFR 4.06-50 - Specimen analysis and follow-up procedures.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... sent to the Medical Review Officer meeting the requirements of 49 CFR 40.121, as designated by the... required by 49 CFR part 40, subpart G, and submit his or her findings to the marine employer. Blood test... 46 Shipping 1 2011-10-01 2011-10-01 false Specimen analysis and follow-up procedures....

  2. 46 CFR 4.06-50 - Specimen analysis and follow-up procedures.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... sent to the Medical Review Officer meeting the requirements of 49 CFR 40.121, as designated by the... required by 49 CFR part 40, subpart G, and submit his or her findings to the marine employer. Blood test... 46 Shipping 1 2010-10-01 2010-10-01 false Specimen analysis and follow-up procedures....

  3. 46 CFR 4.06-50 - Specimen analysis and follow-up procedures.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... sent to the Medical Review Officer meeting the requirements of 49 CFR 40.121, as designated by the... required by 49 CFR part 40, subpart G, and submit his or her findings to the marine employer. Blood test... 46 Shipping 1 2012-10-01 2012-10-01 false Specimen analysis and follow-up procedures....

  4. Novel Selective Medium for Isolation of Staphylococcus lugdunensis from Wound Specimens

    PubMed Central

    Leung, Sammy Man-Him; Tse, Herman; Chow, Kin-Hung; Cheng, Vincent Chi-Chung; Que, Tak-Lun

    2014-01-01

    We compared a novel selective Staphylococcus lugdunensis (SSL) medium with routine media (blood and chocolate agars) for the detection of S. lugdunensis in 990 clinical specimens (from tissue, pus, or wound swabs). Significantly more S. lugdunensis isolates were detected on SSL medium (34/990) than on routine medium (7/990) (P = 0.001, McNemar's test). PMID:24759715

  5. Unusual Histopathological Findings in Childhood Appendectomy Specimens.

    PubMed

    Buyukbese Sarsu, Sevgi; Ucak, Ramazan; Buyukbese, Mehmet Akif; Karakus, Suleyman Cuneyt; Deniz, Hale

    2015-12-01

    The purpose of this study was to find the unusual findings in the childhood appendectomy specimens and their incidence. The clinicopathological data of 1,306 patients whose ages ranged from 3 to 16 were retrospectively collected. Histopathological findings in appendectomy specimens taken from patients who had a prediagnosis of appendicitis were obtained. Incidental appendectomies were not included in the research. Unusual findings were reevaluated in the histopathological assessment of appendectomy specimens. The number of patients whose pathological findings are considered unusual is 25 (1.91 %). Nine of the patients were girls and 16 of them were boys. Their ages ranged from 6 to 15. Pathological results revealed that there were 16 (1.22 %) cases of parasitosis, 3 (0.23 %) cases of granulomatosis, 3 (0.23 %) cases of eosinophilic appendicitis, 2 (0.15 %) cases of carcinoid tumors, and 1 (0.08 %) case of appendiceal non-Hodgkin's lymphoma. All patients underwent a standard appendectomy. Uncommon histopathological findings in childhood appendectomy specimens are more common than those in adulthood. This kind of certain unexpected lesions of the appendix may require advanced diagnostics, careful clinical care, follow-up for years, and a multidisciplinary approach. Therefore, histopathological examinations of appendectomy specimens must be performed routinely. PMID:26730070

  6. Histopathologic findings in breast reduction specimens.

    PubMed

    Kececi, Yavuz; Tasli, Funda Alkan; Yagcı, Ayse; Sır, Emin; Canpolat, Selin; Vardar, Enver

    2014-04-01

    Reduction mammaplasty is a commonly performed operation for treatment of breast hypertrophy. It allows examination of specimens from a seemingly healthy population. Although there are many publications reporting the incidence of occult breast carcinomas, only a few studies have specifically examined the incidence of other breast lesions in reduction mammaplasty specimens. The authors conducted a single-centre retrospective chart review examining the incidence of benign and precancerous lesions in breast reduction specimens. Both age and the number of tissue sections were evaluated for the association with important pathologic findings. Of the 95 patients who underwent reduction mammaplasty, eight patients (8.4%) had atypical lesions. Fourteen patients (15%) had proliferative and 54 patients (57%) had non-proliferative breast lesions. No occult invasive breast cancer was identified in the breast reduction specimens. The existence of significant pathologic findings was not associated with age (p = 0.231, student t-test). On the other hand, it was found to be associated with the number of tissue sections (p = 0.046, Mann-Whitney U-test). This study reveals that breast reduction specimens should be analyzed histologically since a considerable amount of patients have breast lesions with increased cancer risk. Therefore, this analysis would guide the management of these patients in the follow-up period. PMID:23879776

  7. Blood Sugar

    MedlinePlus

    Blood sugar, or glucose, is the main sugar found in your blood. It comes from the food you eat, and is your body's main source of energy. Your blood carries glucose to all of your body's cells to use ...

  8. Blood transfusions

    MedlinePlus

    ... are many reasons you may need a blood transfusion: After knee or hip replacement surgery, or other ... your body cannot make enough blood A blood transfusion is a safe and common procedure during which ...

  9. Blood typing

    MedlinePlus

    ... whether or not there are certain proteins, called antigens, on your red blood cells. Blood is often ... There are many antigens besides the major ones (A, B, and Rh). Many minor ones are not routinely detected during blood typing. If ...

  10. Genomic treasure troves: complete genome sequencing of herbarium and insect museum specimens.

    PubMed

    Staats, Martijn; Erkens, Roy H J; van de Vossenberg, Bart; Wieringa, Jan J; Kraaijeveld, Ken; Stielow, Benjamin; Geml, József; Richardson, James E; Bakker, Freek T

    2013-01-01

    Unlocking the vast genomic diversity stored in natural history collections would create unprecedented opportunities for genome-scale evolutionary, phylogenetic, domestication and population genomic studies. Many researchers have been discouraged from using historical specimens in molecular studies because of both generally limited success of DNA extraction and the challenges associated with PCR-amplifying highly degraded DNA. In today's next-generation sequencing (NGS) world, opportunities and prospects for historical DNA have changed dramatically, as most NGS methods are actually designed for taking short fragmented DNA molecules as templates. Here we show that using a standard multiplex and paired-end Illumina sequencing approach, genome-scale sequence data can be generated reliably from dry-preserved plant, fungal and insect specimens collected up to 115 years ago, and with minimal destructive sampling. Using a reference-based assembly approach, we were able to produce the entire nuclear genome of a 43-year-old Arabidopsis thaliana (Brassicaceae) herbarium specimen with high and uniform sequence coverage. Nuclear genome sequences of three fungal specimens of 22-82 years of age (Agaricus bisporus, Laccaria bicolor, Pleurotus ostreatus) were generated with 81.4-97.9% exome coverage. Complete organellar genome sequences were assembled for all specimens. Using de novo assembly we retrieved between 16.2-71.0% of coding sequence regions, and hence remain somewhat cautious about prospects for de novo genome assembly from historical specimens. Non-target sequence contaminations were observed in 2 of our insect museum specimens. We anticipate that future museum genomics projects will perhaps not generate entire genome sequences in all cases (our specimens contained relatively small and low-complexity genomes), but at least generating vital comparative genomic data for testing (phylo)genetic, demographic and genetic hypotheses, that become increasingly more horizontal

  11. Dry Mouth (Xerostomia)

    MedlinePlus

    ... Gum Disease TMJ Disorders Oral Cancer Dry Mouth Burning Mouth Tooth Decay See All Oral Complications of Systemic ... mouth trouble chewing, swallowing, tasting, or speaking a burning feeling in the mouth a dry feeling in the throat cracked lips ...

  12. Dry eye syndrome

    MedlinePlus

    ... of dry eyes include: Dry environment or workplace (wind, air conditioning) Sun exposure Smoking or second-hand ... NOT smoke and avoid second-hand smoke, direct wind, and air conditioning. Use a humidifier, especially in ...

  13. Dry Skin (Xerosis)

    MedlinePlus

    ... skin, which may bleed if severe. Chapped or cracked lips. When dry skin cracks, germs can get ... cause the skin to become dry, raw, and cracked. Swimming : Some pools have high levels of chlorine, ...

  14. Surface analysis of space telescope material specimens

    NASA Technical Reports Server (NTRS)

    Fromhold, A. T.; Daneshvar, K.

    1985-01-01

    Qualitative and quantitative data on Space Telescope materials which were exposed to low Earth orbital atomic oxygen in a controlled experiment during the 41-G (STS-17) mission were obtained utilizing the experimental techniques of Rutherford backscattering (RBS), particle induced X-ray emission (PIXE), and ellipsometry (ELL). The techniques employed were chosen with a view towards appropriateness for the sample in question, after consultation with NASA scientific personnel who provided the material specimens. A group of eight samples and their controls selected by NASA scientists were measured before and after flight. Information reported herein include specimen surface characterization by ellipsometry techniques, a determination of the thickness of the evaporated metal specimens by RBS, and a determination of trace impurity species present on and within the surface by PIXE.

  15. Stress intensity factor in a tapered specimen

    NASA Technical Reports Server (NTRS)

    Xue-Hui, L.; Erdogan, F.

    1985-01-01

    The general problem of a tapered specimen containing an edge crack is formulated in terms of a system of singular integral equations. The equations are solved and the stress intensity factor is calculated for a compact and for a slender tapered specimen, the latter simulating the double cantilever beam. The results are obtained primarily for a pair of concentrated forces and for crack surface wedge forces. The stress intensity factors are also obtained for a long strip under uniform tension which contains inclined edge cracks.

  16. Attitudes of pregnant women towards collection of biological specimens during pregnancy and at birth.

    PubMed

    Nechuta, Sarah; Mudd, Lanay M; Elliott, Michael R; Lepkowski, James M; Paneth, Nigel

    2012-05-01

    Epidemiological investigations of maternal and child health may involve the collection of biological specimens, including cord blood and the placenta; however, the attitudes of pregnant women towards participation in the collection of biological specimens have been studied rarely. We evaluated attitudes towards collection and storage of biological specimens, and determined whether attitudes differed by maternal characteristics, in a cross-sectional study of pregnant women residing in Kent County, Michigan. Women were interviewed at their first visit for prenatal care between April and October 2006 (n = 311). Willingness to participate was highest for maternal blood collection (72%), followed by storage of biological specimens (68%), placenta collection (64%), and cord blood collection (63%). About one-quarter of women (25-28% by procedure) would not participate even if compensated. Hispanic ethnicity was associated with unwillingness to participate in maternal blood collection (OR = 2.16 [95% CI 1.15, 4.04]). Primiparity was associated with unwillingness to participate in cord blood collection (OR = 1.72 [95% CI 1.23, 2.42]). Among women willing to participate, Hispanic women were less likely to require compensation; while higher educated, married and primiparous women were more likely to require compensation. In conclusion, while many pregnant women were willing to participate in biological specimen collection, some women were more resistant, in particular Hispanic and primiparous women. Targeting these groups of women for enhanced recruitment efforts may improve overall participation rates and the representativeness of participants in future studies of maternal and child health. PMID:22471686

  17. Marking planes of surgical excision on breast biopsy specimens: use of artists' pigments suspended in acetone.

    PubMed Central

    Paterson, D A; Davies, J D

    1988-01-01

    The performance of carbon and metallic inks, silver nitrate solution, and artists' pigments mounted in acetone was compared for marking the surface of surgical biopsy specimens. Using India ink is an unsatisfactory procedure because of slow drying, messiness, and spreading of the ink. It is concluded that use of artists' pigments has many advantages over other reagents, because of their rapid drying, resistance to tissue processing, and the ability to mark simultaneously many different planes of excision. Furthermore, the pigments are readily visible, are distinguishable from each other on microscopical examination, and the method entails little extra cost. Images Fig 1 Fig 2 PMID:3056982

  18. Observation of the freeze-drying process of biological materials with a scanning electron microscope.

    PubMed

    Nei, T; Fujikawa, S

    1976-10-01

    Over the past few decades, numerous studies have been done on the freeze-drying of biological materials from a physical, chemical and biological point of view. Morphological observation of the freeze-drying process of specimens, however, has been tried by only a few investigators. In those studies, thin-layered aqueous specimens, which were sandwiched between two cover slips, were mostly observed with an optical microscope. For ultrastructural and stereoscopic observation, the scanning electron microscope has a great advantage, unlike that of the optical microscope. A specially designed cryo-scanning electron microscope, employed in the present study, made it possible to observe the freezing patterns of the specimens and also the sublimation process of ice in frozen specimens under vacuum. With this specially designed microscope, shrinkage of some specimens due to dehydration during the freeze-drying process was revealed and the extent of such shrinkage was quantitatively determined. PMID:1036327

  19. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 16 Commercial Practices 2 2010-01-01 2010-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  20. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 16 Commercial Practices 2 2013-01-01 2013-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  1. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 16 Commercial Practices 2 2014-01-01 2014-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  2. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 16 Commercial Practices 2 2011-01-01 2011-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  3. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 16 Commercial Practices 2 2013-01-01 2013-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt. 1610, Fig. 3 Figure 3 to Part...

  4. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 16 Commercial Practices 2 2012-01-01 2012-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt.1610, Fig. 3 Figure 3 to Part...

  5. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 16 Commercial Practices 2 2014-01-01 2014-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt. 1610, Fig. 3 Figure 3 to Part...

  6. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 16 Commercial Practices 2 2011-01-01 2011-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt.1610, Fig. 3 Figure 3 to Part...

  7. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 16 Commercial Practices 2 2012-01-01 2012-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt. A, Fig. 6 Figure 6 to Subpart A of Part...

  8. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 16 Commercial Practices 2 2010-01-01 2010-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt.1610, Fig. 3 Figure 3 to Part...

  9. High-intensity drying processes: Impulse drying

    SciTech Connect

    Orloff, D.I.

    1990-09-01

    Impulse drying is an innovative process for drying paper that holds great promise for reducing the energy consumed during the manufacture of paper and similar web products. Impulse drying occurs when a wet paper web passes through a press nip in which one of the rolls is heated to a high temperature. A steam layer adjacent to the heated surface grows and displaces water from the sheet in a very efficient manner. The energy required for water removal is very much less than that required for conventional evaporative drying. Hence, it has been projected that wide commercialization of impulse drying would result in at least a 10% industry-wide energy saving. This report covers work completed between October, 1988 and September, 1989. During this period, pilot press trails demonstrated that newsprint as well as linerboard experience delamination. Hence, the major focus of the research was the resolution of the delamination problem. In order to document potential process improvements, measurement methods were developed to quantify sheet delamination. Using these methods, low thermal diffusivity ceramic roll surfaces were shown to extend the range of impulse drying operating conditions while avoiding sheet delamination. As compared to steel surfaces, ceramics were found to provide significantly higher water volume without inducing sheet delamination. 46 figs., 4 tabs.

  10. Dry deposition velocities

    SciTech Connect

    Sehmel, G.A.

    1984-03-01

    Dry deposition velocities are very difficult to predict accurately. In this article, reported values of dry deposition velocities are summarized. This summary includes values from the literature on field measurements of gas and particle dry deposition velocities, and the uncertainties inherent in extrapolating field results to predict dry deposition velocities are discussed. A new method is described for predicting dry deposition velocity using a least-squares correlation of surface mass transfer resistances evaluated in wind tunnel experiments. 14 references, 4 figures, 1 table.

  11. The production of calibration specimens for impact testing of subsize Charpy specimens

    SciTech Connect

    Alexander, D.J.; Corwin, W.R.; Owings, T.D.

    1994-09-01

    Calibration specimens have been manufactured for checking the performance of a pendulum impact testing machine that has been configured for testing subsize specimens, both half-size (5.0 {times} 5.0 {times} 25.4 mm) and third-size (3.33 {times} 3.33 {times} 25.4 mm). Specimens were fabricated from quenched-and-tempered 4340 steel heat treated to produce different microstructures that would result in either high or low absorbed energy levels on testing. A large group of both half- and third-size specimens were tested at {minus}40{degrees}C. The results of the tests were analyzed for average value and standard deviation, and these values were used to establish calibration limits for the Charpy impact machine when testing subsize specimens. These average values plus or minus two standard deviations were set as the acceptable limits for the average of five tests for calibration of the impact testing machine.

  12. Polyphenol- and fibre-rich dried fruits with green tea attenuate starch-derived postprandial blood glucose and insulin: a randomised, controlled, single-blind, cross-over intervention.

    PubMed

    Nyambe-Silavwe, H; Williamson, G

    2016-08-01

    Polyphenol- and fibre-rich foods (PFRF) have the potential to affect postprandial glycaemic responses by reducing glucose absorption, and thus decreasing the glycaemic response of foods when consumed together. A randomised, single-blind, cross-over study was conducted on sixteen healthy volunteers to test whether PFRF could attenuate postprandial blood glucose in healthy volunteers when added to a source of carbohydrate (starch in bread). This is the first study to examine the effects of a meal comprised of components to inhibit each stage of the biochemical pathway, leading up to the appearance of glucose in the blood. The volunteers were fasted and attended four visits: two control visits (bread, water, balancing sugars) and two test visits (single and double dose of PFRF) where they consumed bread, water and PFRF. Blood samples were collected at 0 (fasted), 15, 30, 45, 60, 90, 120, 150 and 180 min after consumption. The PFRF components were tested for α-amylase and α-glucosidase inhibitory potential in vitro. Plasma glucose was lower after consumption of both doses compared with controls: lower dose, change in mean incremental areas under the glucose curves (IAUC)=-27·4 (sd 7·5) %, P<0·001; higher dose, IAUC=-49·0 (sd 15·3) %, P<0·001; insulin IAUC was also attenuated by-46·9 (sd 13·4) %, P<0·01. Consistent with this, the polyphenol components of the PFRF inhibited α-amylase (green tea, strawberry, blackberry and blackcurrant) and α-glucosidase (green tea) activities in vitro. The PFRF have a pronounced and significant lowering effect on postprandial blood glucose and insulin response in humans, due in part to inhibition of α-amylase and α-glucosidase, as well as glucose transport. PMID:27278405

  13. Some recent innovations in small specimen testing

    NASA Astrophysics Data System (ADS)

    Odette, G. R.; He, M.; Gragg, D.; Klingensmith, D.; Lucas, G. E.

    2002-12-01

    New innovative small specimen test techniques are described. Finite element simulations show that combinations of cone indentation pile-up geometry and load-penetration depth relations can be used to determine both the yield stress and strain-hardening behavior of a material. Techniques for pre-cracking and testing sub-miniaturized fracture toughness bend bars, with dimensions of 1.65×1.65×9 mm 3, or less, are described. The corresponding toughness-temperature curves have a very steep transition slope, primarily due to rapid loss of constraint, which has advantages in some experiments to characterize the effects of specified irradiation variables. As one example of using composite specimens, an approach to evaluating helium effects is proposed, involving diffusion bonding small wires of a 54Fe-based ferritic-martensitic alloy to a surrounding fracture specimen composed of an elemental Fe-based alloy. Finally, we briefly outline some potential approaches to multipurpose specimens and test automation.

  14. Recent progress in small specimen test technology

    NASA Astrophysics Data System (ADS)

    Lucas, G. E.; Odette, G. R.; Sokolov, M.; Spätig, P.; Yamamoto, T.; Jung, P.

    2002-12-01

    Small specimen test technology (SSTT) has enabled the development of fusion materials by efficiently using available irradiation volumes. The technology has also evolved in anticipation of the construction and operation of a high-energy neutron source for development and verification of an engineering database for materials for fusion power reactors. Work to date has brought SSTT to a robust state of maturity. SSTT specimens and techniques now routinely serve as the foundation for a number of ongoing and planned experimental programs. Moreover, the need to use small specimens has given rise to the development of new approaches to fracture assessment, such as the master curves-shifts method. Nonetheless a wealth of opportunities exists to further develop new and very innovative SSTT methods not only for characterizing standard mechanical properties but also to enable both large matrix single variable experiments and highly controlled basic mechanism studies. This paper reviews briefly the recent progress on developing a more science-based SSTT, including some future opportunities. The importance and utility of applying a variety of quasi-non-destructive evaluations to a single specimen and closely integrating finite element simulations and fundamental models of deformation and fracture are emphasized.

  15. Vee-notch tool cuts specimens

    NASA Technical Reports Server (NTRS)

    Spier, R. A.

    1970-01-01

    Triangular cutting tool uses carbide tips for notching heat-treated or abrasive materials, and alloys subjected to high structural stresses. The tool is rigidly mounted in a slot of mating contour to prevent deflection during cutting of tensile specimens. No other expensive machine equipment is required.

  16. Myocardial Sleeve Tissues in Surgical Lung Specimens.

    PubMed

    Yoshida, Akihiko; Kamata, Tsugumasa; Iwasa, Takeshi; Watanabe, Shun-ichi; Tsuta, Koji

    2015-10-01

    Left atrial myocardial extensions over the pulmonary veins (PVs), known as myocardial sleeves, are present in the physiological anatomy of most individuals. Although this structure has recently received clinical attention as a major origin of paroxysmal atrial fibrillation (AF), it has not been documented in surgical specimens. Here, we examine incidentally identified myocardial sleeve tissue in routinely processed lung resection specimens to determine its incidence and diagnostic implications. Among 694 lung resection specimens with evaluable PV margins, myocardial sleeve tissue was identified in 26 cases (3.7%). The tissue was located within the adventitia of the PVs, mostly in margin preparations, and existed outside the pericardium in the majority of cases. Carcinoma infiltration of the sleeves was evident in 6 cases. No heart injuries were observed, and no tumors invaded the heart. Preoperative electrocardiography showed sinus rhythm in all cases, whereas postoperative monitoring revealed sinus rhythm in all patients except one who showed AF and flutter. Myocardial sleeve tissue is an underrecognized incidental finding in lung resection specimens, and it is not indicative of heart injury. Cancer infiltration into this tissue indicates neither heart invasion nor, by itself, invasion into the pericardium. Although surgical transection of the myocardial sleeve did not evoke immediate arrhythmia in most cases, the overall influence of this procedure on the postsurgical risk of AF remains to be determined in further studies involving extensive rhythm assessment. PMID:26099012

  17. 36 CFR 2.5 - Research specimens.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 43 CFR part 24. Regulations concerning archeological resources are found in 43 CFR part 3. ... Federal agency for the purpose of research, baseline inventories, monitoring, impact analysis, group study... if removal of the specimen would result in damage to other natural or cultural resources,...

  18. 36 CFR 2.5 - Research specimens.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 43 CFR part 24. Regulations concerning archeological resources are found in 43 CFR part 3. ... Federal agency for the purpose of research, baseline inventories, monitoring, impact analysis, group study... if removal of the specimen would result in damage to other natural or cultural resources,...

  19. 3-D Volume Rendering of Sand Specimen

    NASA Technical Reports Server (NTRS)

    2004-01-01

    Computed tomography (CT) images of resin-impregnated Mechanics of Granular Materials (MGM) specimens are assembled to provide 3-D volume renderings of density patterns formed by dislocation under the external loading stress profile applied during the experiments. Experiments flown on STS-79 and STS-89. Principal Investigator: Dr. Stein Sture

  20. 7 CFR 97.8 - Specimen requirements.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... PLANT VARIETY AND PROTECTION The Application § 97.8 Specimen requirements. (a) The applicant may be..., in a quantity and at a specified stage of growth, as may be necessary to verify the statements in the... reimburse the Office for all costs, including travel, per diem or subsistence, and salary. (b)...