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

Science.gov (United States)

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. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Feasibility of self-sampled dried blood spot and saliva samples sent by mail in a population-based study.

Science.gov (United States)

Sakhi, Amrit Kaur; Bastani, Nasser Ezzatkhah; Ellingjord-Dale, Merete; Gundersen, Thomas Erik; Blomhoff, Rune; Ursin, Giske

2015-04-11

In large epidemiological studies it is often challenging to obtain biological samples. Self-sampling by study participants using dried blood spots (DBS) technique has been suggested to overcome this challenge. DBS is a type of biosampling where blood samples are obtained by a finger-prick lancet, blotted and dried on filter paper. However, the feasibility and efficacy of collecting DBS samples from study participants in large-scale epidemiological studies is not known. The aim of the present study was to test the feasibility and response rate of collecting self-sampled DBS and saliva samples in a population-based study of women above 50 years of age. We determined response proportions, number of phone calls to the study center with questions about sampling, and quality of the DBS. We recruited women through a study conducted within the Norwegian Breast Cancer Screening Program. Invitations, instructions and materials were sent to 4,597 women. The data collection took place over a 3 month period in the spring of 2009. Response proportions for the collection of DBS and saliva samples were 71.0% (3,263) and 70.9% (3,258), respectively. We received 312 phone calls (7% of the 4,597 women) with questions regarding sampling. Of the 3,263 individuals that returned DBS cards, 3,038 (93.1%) had been packaged and shipped according to instructions. A total of 3,032 DBS samples were sufficient for at least one biomarker analysis (i.e. 92.9% of DBS samples received by the laboratory). 2,418 (74.1%) of the DBS cards received by the laboratory were filled with blood according to the instructions (i.e. 10 completely filled spots with up to 7 punches per spot for up to 70 separate analyses). To assess the quality of the samples, we selected and measured two biomarkers (carotenoids and vitamin D). The biomarker levels were consistent with previous reports. Collecting self-sampled DBS and saliva samples through the postal services provides a low cost, effective and feasible

Evaluation of a novel dried blood spot collection device (HemaSpot™) to test blood samples collected from dogs for antibodies to Leishmania infantum.

Science.gov (United States)

Rosypal, Alexa C; Pick, Leanne D; Hernandez, Jaime O Esquivel; Lindsay, David S

2014-09-15

Collection of blood samples from veterinary and wildlife patients is often challenging because the samples have to be collected on farm or in the wild under various environmental conditions. This poses many technical problems associated with venipuncture materials, their safe use and disposal, transportation and processing of collected samples. Dried blood spot (DBS) sample collection techniques offer a simple and practical alternative to traditional blood collection methods to obtain blood samples from animals for parasite antibody evaluation. The DBS collection devices are compact, simple to use, and are particularly useful for large number of samples. Additionally, DBS samples take up less space and they are easier to transport than traditional venipuncture-collected blood samples. Visceral leishmaniasis (VL) is a potentially fatal parasitic disease of dogs and humans and it is frequently diagnosed by antibody tests. Immunochromatographic tests (ICT) for antibodies to Leishmania infantum are commercially available for dogs and they produce qualitative results in minutes. Measurement of canine antibodies to L. infantum with the ICT using traditional venipuncture has been validated previously, but the use of DBS samples has not been evaluated using this method. The purpose of the present study was to determine the ability of DBS samples to detect antibodies to L. infantum in dogs using a commercial ICT assay. One hundred plasma samples from dogs experimentally infected with the LIVT-1 strain of L. infantum were collected by venipuncture and frozen. Individual samples were thawed, and then 80 μl plasma (2 drops) was aliquotted onto the 8-spoked disk pad on individual DBS sample collection devices (HemaSpot™, Spot-On Sciences, Austin, TX), dried, and stored in the dark at room temperature. After one month and six months, respectively, 2 spokes of the 8 spokes of the disk pad of each DBS sample were removed and eluted in 200 μl PBS. The eluate was used to test

Analysis of multiple single nucleotide polymorphisms (SNP) on DNA traces from plasma and dried blood samples

NARCIS (Netherlands)

Catsburg, Arnold; van der Zwet, Wil C.; Morre, Servaas A.; Ouburg, Sander; Vandenbroucke-Grauls, Christina M. J. E.; Savelkoul, Paul H. M.

2007-01-01

Reliable analysis of single nucleotide polymorphisms (SNPs) in DNA derived from samples containing low numbers of cells or from suboptimal sources can be difficult. A new procedure to characterize multiple SNPs in traces of DNA from plasma and old dried blood samples was developed. Six SNPs in the

Feasibility of self-sampled dried blood spot and saliva samples sent by mail in a population-based study

International Nuclear Information System (INIS)

Sakhi, Amrit Kaur; Bastani, Nasser Ezzatkhah; Ellingjord-Dale, Merete; Gundersen, Thomas Erik; Blomhoff, Rune; Ursin, Giske

2015-01-01

In large epidemiological studies it is often challenging to obtain biological samples. Self-sampling by study participants using dried blood spots (DBS) technique has been suggested to overcome this challenge. DBS is a type of biosampling where blood samples are obtained by a finger-prick lancet, blotted and dried on filter paper. However, the feasibility and efficacy of collecting DBS samples from study participants in large-scale epidemiological studies is not known. The aim of the present study was to test the feasibility and response rate of collecting self-sampled DBS and saliva samples in a population–based study of women above 50 years of age. We determined response proportions, number of phone calls to the study center with questions about sampling, and quality of the DBS. We recruited women through a study conducted within the Norwegian Breast Cancer Screening Program. Invitations, instructions and materials were sent to 4,597 women. The data collection took place over a 3 month period in the spring of 2009. Response proportions for the collection of DBS and saliva samples were 71.0% (3,263) and 70.9% (3,258), respectively. We received 312 phone calls (7% of the 4,597 women) with questions regarding sampling. Of the 3,263 individuals that returned DBS cards, 3,038 (93.1%) had been packaged and shipped according to instructions. A total of 3,032 DBS samples were sufficient for at least one biomarker analysis (i.e. 92.9% of DBS samples received by the laboratory). 2,418 (74.1%) of the DBS cards received by the laboratory were filled with blood according to the instructions (i.e. 10 completely filled spots with up to 7 punches per spot for up to 70 separate analyses). To assess the quality of the samples, we selected and measured two biomarkers (carotenoids and vitamin D). The biomarker levels were consistent with previous reports. Collecting self-sampled DBS and saliva samples through the postal services provides a low cost, effective and feasible

A simple assay of paracetamol based on dried blood spot suitable ...

African Journals Online (AJOL)

Dried blood spots in Guthrie cards are a reliable means of blood sampling suitable for pharmacoki-netic analysis in children. The aim of this study was to develop a simple and reliable bioanalytical method to measure the concentration of paracetamol in dried blood spots. Paracetamol was ex-tracted from dry blood spots by ...

Blood and dried blood spot telomere length measurement by qPCR: assay considerations.

Directory of Open Access Journals (Sweden)

DeAnna L Zanet

Full Text Available Measurement of telomere length is crucial for the study of telomere maintenance and its role in molecular pathophysiology of diseases and in aging. Several methods are used to measure telomere length, the choice of which usually depends on the type and size of sample to be assayed, as well as cost and throughput considerations. The goal of this study was to investigate the factors that may influence the reliability of qPCR-based relative telomere length measurements in whole blood. Day to day intra-individual variability, types of blood anticoagulant, sample storage conditions, processing and site of blood draw were investigated. Two qPCR-based methods to measure telomere length (monoplex vs. multiplex were also investigated and showed a strong correlation between them. Freezing and thawing of the blood and storage of the blood at 4°C for up to 4 days did not affect telomere length values. Telomere lengths in dried blood spots were significantly higher than both whole blood and peripheral mononuclear blood cells, and were highly correlated with both. We found that telomere length measurements were significantly higher in dried blood spots collected directly from fingertip prick compared to dried blood spots prepared with anticoagulated whole blood collected from the finger, and non-blotted whole blood taken from both finger and arm venipuncture. This suggests that DNA from cells blotted on paper is not equivalent to that collected from venipuncture whole blood, and caution should be taken when comparing between blood sample types.

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

Science.gov (United States)

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.

Total reflection x-ray analysis of metals in blood samples

International Nuclear Information System (INIS)

Nakamura, Takuya; Matsui, Hiroshi; Kawamata, Masaya

2009-01-01

The sample preparation for TXRF (total reflection X-ray fluorescence) quantitative analysis of trace elements in human blood samples was investigated. In the TXRF analysis, a solution sample is dropped and dried on a flat substrate, and then the dried residue is measured. In this case, the dried residue should be flat not to disturb X-ray total reflection on the substrate. In addition, it is required to simply measure the whole blood sample by TXRF method, although a serum is analyzed in many cases. Thus, we studied the optimum conditions of the sample preparation of the whole blood by adding the pure water to apply Hemolysis phenomenon, where blood cells are destroyed due to different of the osmotic pressure, leading to flat residue. It was found that the best S/B ratio was obtained when the whole blood was diluted 8 times with pure water. Moreover, it was investigated the influence of the surface chemical condition of the glass substrate on the shape of the dried reside of the blood sample. When the surface of the glass substrate was hydrophilic, the shape of the dried residues was not uniform, as a result, the quantitative data of TXRF analysis gave a large deviation. On the other hand, when the surface of the glass was hydrophobic, the shape of the residue was almost uniform, as a result, a good reproducibility was obtained. Another problem was an outer ring of the dried residue of the blood. This uneven ring absorbs the primary X-rays, caused to low determined quantitative data. Thus, we tried the heating way of the dropped blood sample at a high temperature of 200 degrees. In this case, the blood sample was dried immediately, and a flat homogeneous dried residue was obtained without the outer ring. Using the optimized conditions for sample preparation, human blood sample was quantitatively measured by TXRF and ICP-AES. A good agreement was obtained in TXRF and ICP-AES determinations; however, the measurement of Cl and Br will be an advantage of TXRF, because

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

Science.gov (United States)

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.

Research results: preserving newborn blood samples.

Science.gov (United States)

Lewis, Michelle Huckaby; Scheurer, Michael E; Green, Robert C; McGuire, Amy L

2012-11-07

Retention and use, without explicit parental permission, of residual dried blood samples from newborn screening has generated public controversy over concerns about violations of family privacy rights and loss of parental autonomy. The public debate about this issue has included little discussion about the destruction of a potentially valuable public resource that can be used for research that may yield improvements in public health. The research community must advocate for policies and infrastructure that promote retention of residual dried blood samples and their use in biomedical research.

Adiponectin levels measured in dried blood spot samples from neonates born small and appropriate for gestational age

DEFF Research Database (Denmark)

Klamer, A; Skogstrand, Kristin; Hougaard, D M

2007-01-01

Adiponectin levels measured in neonatal dried blood spot samples (DBSS) might be affected by both prematurity and being born small for gestational age (SGA). The aim of the study was to measure adiponectin levels in routinely collected neonatal DBSS taken on day 5 (range 3-12) postnatal from...

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.

Science.gov (United States)

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.

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

Science.gov (United States)

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

Role of therapeutic drug monitoring in pulmonary infections : use and potential for expanded use of dried blood spot samples

NARCIS (Netherlands)

Hofman, Susan; Bolhuis, Mathieu S.; Koster, Remco A.; Akkerman, Onno W.; van Assen, Sander; Stove, Christophe; Alffenaar, Jan-Willem C.

Respiratory tract infections are among the most common infections in men. We reviewed literature to document their pharmacological treatments, and the extent to which therapeutic drug monitoring (TDM) is needed during treatment. We subsequently examined potential use of dried blood spots as sample

Direct analysis of site-specific N-glycopeptides of serological proteins in dried blood spot samples.

Science.gov (United States)

Choi, Na Young; Hwang, Heeyoun; Ji, Eun Sun; Park, Gun Wook; Lee, Ju Yeon; Lee, Hyun Kyoung; Kim, Jin Young; Yoo, Jong Shin

2017-08-01

Dried blood spot (DBS) samples have a number of advantages, especially with respect to ease of collection, transportation, and storage and to reduce biohazard risk. N-glycosylation is a major post-translational modification of proteins in human blood that is related to a variety of biological functions, including metastasis, cell-cell interactions, inflammation, and immunization. Here, we directly analyzed tryptic N-glycopeptides from glycoproteins in DBS samples using liquid chromatography-tandem mass spectrometry (LC-MS/MS) without centrifugation of blood samples, depletion of major proteins, desalting of tryptic peptides, and enrichment of N-glycopeptides. Using this simple method, we identified a total of 41 site-specific N-glycopeptides from 16 glycoproteins in the DBS samples, from immunoglobulin gamma 1 (IgG-1, 10 mg/mL) down to complement component C7 (50 μg/mL). Of these, 32 N-glycopeptides from 14 glycoproteins were consistently quantified over 180 days stored at room temperature. The major abundant glycoproteins in the DBS samples were IgG-1 and IgG-2, which contain nine asialo-fucosylated complex types of 16 different N-glycopeptide isoforms. Sialo-non-fucosylated complex types were primarily detected in the other glycoproteins such as alpha-1-acid glycoprotein 1, 2, alpha-1-antitypsin, alpha-2-macroglobulin, haptoglobin, hemopexin, Ig alpha 1, 2 chain C region, kininogen-1, prothrombin, and serotransferrin. We first report the characterization of site-specific N-glycoproteins in DBS samples by LC-MS/MS with minimal sample preparation.

A simple assay of paracetamol based on dried blood

African Journals Online (AJOL)

User

Dried blood spots in Guthrie cards are a reliable means of blood sampling suitable for pharmacoki- netic analysis in .... pers were then packed in plastic bags and stored at. 4ºC in the ..... of paracetamol and its metabolites in urine, plasma and ...

Suspected hypoglycaemia in out patient practice: accuracy of dried blood spot analysis.

Science.gov (United States)

Parker, D R; Bargiota, A; Cowan, F J; Corrall, R J

1997-12-01

The assay of dried blood spots on filter paper to determine blood glucose concentration has been used to detect hypoglycaemia in out patients. We assessed the accuracy of this approach in assaying blood glucose concentrations in the hypoglycaemic range. Volunteers were rendered hypoglycaemic by intravenous infusion of insulin. The glucose concentration in simultaneously taken blood samples was measured either fresh or after drying on filter paper. Twenty-four healthy young volunteers and 9 patients with insulin-dependent diabetes were studied. Plasma glucose concentrations were measured using a standard auto analyser glucose oxidase method. Whole blood taken simultaneously was placed on prepared filter paper and allowed to dry; glucose concentration was then measured using a well-established technique. A correction factor was applied to convert the glucose concentration of plasma to that of whole blood. The relationship between glucose concentrations measured by the two methods was determined by regression coefficient. In the unequivocally hypoglycaemic range (plasma dried blood spot glucose concentrations significantly correlated with standard plasma glucose concentrations (r = 0.81; P dried blood spot method had a sensitivity of 91%. In the range designated probable hypoglycaemia (plasma dried blood spot method was 100% in both ranges. Measurement of glucose concentrations in dried blood spots is specific and sensitive in the hypoglycaemic range. The present study indicates that hypoglycaemia may be excluded or confirmed respectively when levels in excess of 3.7 or below 2.8 mmol/l are found in uncorrected dried blood spot analysis.

Fibrinogen measurements in plasma and whole blood: a performance evaluation study of the dry-hematology system.

Science.gov (United States)

Ogawa, Satoru; Tanaka, Kenichi A; Nakajima, Yasufumi; Nakayama, Yoshinobu; Takeshita, Jun; Arai, Masatoshi; Mizobe, Toshiki

2015-01-01

An accurate and rapid determination of fibrinogen level is important during hemorrhage to establish a timely hemostatic intervention. The accuracy of fibrinogen measurements may be affected by the specific methodology for its determination, fluid therapies, and anticoagulant agents. The dry-hematology method (DRIHEMATO®) is a novel approach to determine fibrinogen levels in plasma and whole blood based on thrombin-activated coagulation time. We hypothesized that plasma or whole blood fibrinogen level using the dry-hematology method would be similar to those measured with conventional plasma fibrinogen assays. Acquired hypofibrinogenemia was modeled by serial dilutions of blood samples obtained from 12 healthy volunteers. Citrated whole blood samples were diluted with either normal saline, 5% human albumin, or 6% hydroxyethyl starch to achieve 25%, 50%, and 75% volume replacement. The dry-hematology method, the Clauss method, the prothrombin time (PT)-derived method, determination of antigen levels, and thromboelastometric fibrin formation were compared in plasma or whole blood samples. The effect of heparin on each assay was examined (0 to 6 IU/mL). Comparisons of dry-hematology and other methods were also conducted using ex vivo samples obtained from cardiac surgical patients (n = 60). In plasma samples, there were no significant differences between dry-hematology and the Clauss method, while dry-hematology showed lower fibrinogen levels compared with PT-derived and antigen level methods. The dry-hematology method yielded acceptable concordance correlation coefficients (Pc) with the Clauss method, the PT-derived method, and fibrinogen antigen levels (Pc = 0.91-0.99). The type of diluents and heparin affected the results of the PT-derived method and thromboelastometric assay, but not the dry-hematology method. In cardiac surgical patients, the overall correlation in fibrinogen levels between dry-hematology and the other methods was comparable to the results from

Evaluation of Glucose-6-Phosphate Dehydrogenase stability in stored blood samples.

Science.gov (United States)

Jalil, Norunaluwar; Azma, Raja Zahratul; Mohamed, Emida; Ithnin, Azlin; Alauddin, Hafiza; Baya, Siti Noor; Othman, Ainoon

2016-01-01

Glucose-6-Phosphate Dehydrogenase (G6PD) deficiency is the commonest cause of neonatal jaundice in Malaysia. Recently, OSMMR2000-D G6PD Assay Kit has been introduced to quantitate the level of G6PD activity in newborns delivered in Universiti Kebangsaan Malaysia Medical Centre (UKMMC). As duration of sample storage prior to analysis is one of the matters of concern, this study was conducted to identify the stability of G6PD enzyme during storage. A total of 188 cord blood samples from normal term newborns delivered at UKMMC were selected for this study. The cord bloods samples were collected in ethylene-diamine-tetra-acetic acid (EDTA) tubes and refrigerated at 2-8 °C. In addition, 32 out of 188 cord blood samples were spotted on chromatography paper, air-dried and stored at room temperature. G6PD enzyme activities were measured daily for 7 days using the OSMMR2000-D G6PD Assay Kit on both the EDTA blood and dried blood samples. The mean value for G6PD activity was compared between days of analysis using Student Paired T-Test. In this study, 172 out of 188 cord blood samples showed normal enzyme levels while 16 had levels corresponding to severe enzyme deficiency. The daily mean G6PD activity for EDTA blood samples of newborns with normal G6PD activity showed a significant drop on the fourth day of storage (p samples with severely deficient G6PD activity, significant drop was seen on third day of storage (p = 0.002). Analysis of dried cord blood showed a significant reduction in enzyme activity as early as the second day of storage (p = 0.001). It was also noted that mean G6PD activity for spotted blood samples were lower compared to those in EDTA tubes for all days (p = 0.001). Thus, EDTA blood samples stored at 2-8 °C appeared to have better stability in terms of their G6PD enzyme level as compared to dried blood samples on filter paper, giving a storage time of up to 3 days.

Rapid DNA extraction from dried blood spots on filter paper: potential applications in biobanking.

Science.gov (United States)

Choi, Eun-Hye; Lee, Sang Kwang; Ihm, Chunhwa; Sohn, Young-Hak

2014-12-01

Dried blood spot (DBS) technology is a microsampling alternative to traditional plasma or serum sampling for pharmaco- or toxicokinetic evaluation. DBS technology has been applied to diagnostic screening in drug discovery, nonclinical, and clinical settings. We have developed an improved elution protocol involving boiling of blood spots dried on Whatman filter paper. The purpose of this study was to compare the quality, purity, and quantity of DNA isolated from frozen blood samples and DBSs. We optimized a method for extraction and estimation of DNA from blood spots dried on filter paper (3-mm FTA card). A single DBS containing 40 μL blood was used. DNA was efficiently extracted in phosphate-buffered saline (PBS) or Tris-EDTA (TE) buffer by incubation at 37°C overnight. DNA was stable in DBSs that were stored at room temperature or frozen. The housekeeping genes GAPDH and beta-actin were used as positive standards for polymerase chain reaction (PCR) validation of general diagnostic screening. Our simple and convenient DBS storage and extraction methods are suitable for diagnostic screening by using very small volumes of blood collected on filter paper, and can be used in biobanks for blood sample storage.

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

Directory of Open Access Journals (Sweden)

Markus Kopp

Full Text Available 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.

Thyroxine and thyrotropin radioimmunoassays using dried blood samples on filter paper for screening of neonatal hypothyroidism

International Nuclear Information System (INIS)

Beckers, C.; Cornette, C.; Francois, B.; Bouckaert, A.; Lechat, M.

1977-01-01

A routine and automatized methodology for thyroxine (T4) and thyrotropin (TSH) radioimmunoassay (RIA) using dried blood samples on filter paper is described. Five mm diameter dots were prepared. One eluted dot, corresponding to 4 μl of plasma, was used for T4-RIA while two were necessary for TSH-RIA. Reference filter papers were introduced in each assay for quality control. In a preliminary study on 1903 newborns, samples were obtained, generally between the 5th-7th day. Mean dot T4 was 7.38 +- 2.5 μg/dl. Mean dot TSH was 11.83 +- 9.1 μU/ml, the equation of the regression line between dot TSH (y) and serum TSH (x) being Y = 10.29 + 0.623x. (orig.) [de

Archived neonatal dried blood spot samples can be used for accurate whole genome and exome-targeted next-generation sequencing

DEFF Research Database (Denmark)

Hollegaard, Mads Vilhelm; Grauholm, Jonas; Nielsen, Ronni

2013-01-01

Dried blood spot samples (DBSS) have been collected and stored for decades as part of newborn screening programmes worldwide. Representing almost an entire population under a certain age and collected with virtually no bias, the Newborn Screening Biobanks are of immense value in medical studies......, for example, to examine the genetics of various disorders. We have previously demonstrated that DNA extracted from a fraction (2×3.2mm discs) of an archived DBSS can be whole genome amplified (wgaDNA) and used for accurate array genotyping. However, until now, it has been uncertain whether wgaDNA from DBSS...... can be used for accurate whole genome sequencing (WGS) and exome sequencing (WES). This study examined two individuals represented by three different types of samples each: whole-blood (reference samples), 3-year-old DBSS spotted with reference material (refDBSS), and 27- to 29-year-old archived...

Evaluation of dry blood spot technique for quantification of an Anti-CD20 monoclonal antibody drug in human blood samples.

Science.gov (United States)

Lin, Yong-Qing; Zhang, Yilu; Li, Connie; Li, Louis; Zhang, Kelley; Li, Shawn

2012-01-01

To evaluate the dried blood spot (DBS) technique in ELISA quantification of larger biomolecular drugs, an anti-CD20 monoclonal antibody drug was used as an example. A method for the quantification of the anti-CD20 drug in human DBS was developed and validated. The drug standard and quality control samples prepared in fresh human blood were spotted on DBS cards and then extracted. A luminescent ELISA was used for quantification of the drug from DBS samples. The assay range of the anti-CD20 drug standards in DBS was 100-2500ng/mL. The intra-assay precision (%CV) ranged from 0.4% to 10.1%, and the accuracy (%Recovery) ranged from 77.9% to 113.9%. The inter assay precision (%CV) ranged from 5.9% to 17.4%, and the accuracy ranged from 81.5% to 110.5%. The DBS samples diluted 500 and 50-fold yielded recovery of 88.7% and 90.7%, respectively. The preparation of DBS in higher and lower hematocrit (53% and 35%) conditions did not affect the recovery of the drug. Furthermore, the storage stability of the anti-CD20 drug on DBS cards was tested at various conditions. It was found that the anti-CD20 drug was stable for one week in DBS stored at room temperature. However, it was determined that the stability was compro]mised in DBS stored at high humidity, high temperature (55°C), and exposed to direct daylight for a week, as well as for samples stored at room temperature and high humidity conditions for a month. Stability did not change significantly in samples that underwent 3 freeze/thaw cycles. Our results demonstrated a successful use of DBS technique in ELISA quantification of an anti-CD20 monoclonal antibody drug in human blood. The stability data provides information regarding sample storage and shipping for future clinical studies. It is, therefore, concluded that the DBS technique is applicable in the quantification of other large biomolecule drugs or biomarkers. Copyright © 2011 Elsevier Inc. All rights reserved.

Aflatoxin B1-lysine adduct in dried blood spot samples of animals and humans.

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Xue, Kathy S; Cai, Wenjie; Tang, Lili; Wang, Jia-Sheng

2016-12-01

Dried blood spots (DBS) were proposed as potentially viable method for exposure assessment of environmental toxicants in infant and young children. For this study, we validated an experimental protocol to quantify AFB 1 -lysine adduct in DBS samples of AFB 1 -treated F344 rats, as well as samples from human field study. Significant dose-response relationships in AFB 1 -lysine adduct formation were found in DBS samples of rats treated with single- and repeated-dose AFB 1 . AFB 1 -lysine levels in DBS samples were highly correlated with corresponding serum sample levels. The Person coefficients were 0.997 for the single-dose exposure, and 0.996 for the repeated-dose exposure. Levels of AFB 1 -lysine adduct had also good agreement between DBS and serum samples as shown by Bland-Altman plot analysis. For human field study samples (n = 36), a Pearson correlation coefficient of 0.784 was found between AFB 1 -lysine adduct levels of DBS and corresponding serum samples. Bland-Altman plots showed the distribution of the log differences between DBS and serum AFB 1 -lysine levels are within 95% confidence intervals. These results showed AFB 1 -lysine adduct levels in DBS cards and serum samples from animals and human samples are comparable, and the DBS technique and analytical protocol is a good means to assess AFB 1 exposure in infant and children populations. Copyright © 2016 Elsevier Ltd. All rights reserved.

Automated immunoradiometric assay of thyrotrophin (TSH) in dried blood filter paper spots

Energy Technology Data Exchange (ETDEWEB)

John, R.; Woodhead, J.S. (Welsh National School of Medicine, Cardiff (UK))

1982-11-10

An immunoradiometric two-site assay for thyrotrophin (TSH) in dried blood filter paper spots is described. The assay is automated by means of the Kemtek 3000 automated immunoassay system. The technique uses a 6.0 mm disc punched from the dried blood samples collected as part of the screening programme for phenylketonuria. The method is sensitive and precise, and results correlate well with those obtained in TSH assays of serum samples. The procedure is rapid, results being available within 24 h of receipt of samples. Of 25204 specimens so far screened by this assay, 99.9% have TSH levels less than 15 mU/l. One false positive result has been obtained and six confirmed cases of neonatal hypothyroidism detected, giving a prevalence of 1 in 4200.

Study of measurement of the alcohol biomarker phosphatidylethanol (PEth) in dried blood spot (DBS) samples and application of a volumetric DBS device.

Science.gov (United States)

Beck, Olof; Kenan Modén, Naama; Seferaj, Sabina; Lenk, Gabriel; Helander, Anders

2018-04-01

Phosphatidylethanol (PEth) is a group of phospholipids formed in cell membranes following alcohol consumption. PEth measurement in whole blood samples is established as a specific alcohol biomarker with clinical and medico-legal applications. This study further evaluated the usefulness of dried blood spot (DBS) samples collected on filter paper for PEth measurement. Specimens used were surplus volumes of venous whole blood sent for routine LC-MS/MS quantification of PEth 16:0/18:1, the major PEth homolog. DBS samples were prepared by pipetting blood on Whatman 903 Protein Saver Cards and onto a volumetric DBS device (Capitainer). The imprecision (CV) of the DBS sample amount based on area and weight measurements of spot punches were 23-28%. Investigation of the relationship between blood hematocrit and PEth concentration yielded a linear, positive correlation, and at around 1.0-1.5μmol/L PEth 16:0/18:1, the PEth concentration increased by ~0.1μmol/L for every 5% increase in hematocrit. There was a close agreement between the PEth concentrations obtained with whole blood samples and the corresponding results using Whatman 903 (PEth DBS =1.026 PEth WB +0.013) and volumetric device (PEth DBS =1.045 PEth WB +0.016) DBS samples. The CV of PEth quantification in DBS samples at concentrations≥0.05μmol/L were ≤15%. The present results further confirmed the usefulness of DBS samples for PEth measurement. Copyright © 2018 Elsevier B.V. All rights reserved.

Thyroxine (T4) radioimmunoassay using filter paper dried blood sample: an attempt for screening of neonates for hypothyroidism

International Nuclear Information System (INIS)

Afroz, S.; Hussain, R.; Ahmed, K.

1997-01-01

This paper describes a sensitive but simple and less expensive method suitable for estimation of thyroxine (T 4 ) level. Deficiency of iodine during fetal life results in neonatal hypothyroidism and critinism. Frequency of neonatal hypothyroidism is 1 in 5000 to 7000 in countries having iodine deficiency. It is therefore important to diagnose the neonatal hypothyroidism as soon as possible after birth. The estimation of thyroxine has been found to the a reliable index for diagnosis of hypothyroidism and has long been used for screening of neonatal hypothyroidism. In the present study, instead of serum sample, a 6 mm disc of filter paper containing dried blood sample was used. The test was carried out in the laboratory with 40 samples. As compared to the sensitivity of serum sample technique which is 15.19 n mol/L, the filter paper technique has the sensitivity of 17.23 n mol/L. The work revealed that the T 4 concentration do not depend upon the amount of blood on the filter paper. Effect of temperature on filter paper disc was evaluated at 4 o c, at 25 o c and at 37 o c. Results obtained showed significant variation and the best result was obtained for the sample kept at 4 o c. The method is simple, rapid, less expensive and needs a small amount of blood and is, therefore, a useful technique for mass screening of neonatal hypothyroidism. 6 refs., 4 tables (author)

Determination of branched chain amino acids, methionine, phenylalanine, tyrosine and alpha-keto acids in plasma and dried blood samples using HPLC with fluorescence detection.

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Kand'ár, Roman; Záková, Pavla; Jirosová, Jana; Sladká, Michaela

2009-01-01

The determination of branched chain amino acids [BCAA; valine (Val), leucine (Leu), isoleucine (Ile)], alpha-keto acids derived from BCAA [BCKA; alpha-ketoisovaleric acid (KIV), alpha-ketoisocaproic acid (KIC), alpha-ketomethylvaleric acid (KMV)], methionine (Met), phenylalanine (Phe) and tyrosine (Tyr) is currently the most reliable approach for the diagnosis of maple syrup urine disease (MSUD), hypermethioninemia, phenylketonuria (PKU) and tyrosinemia. The aim of this study was to develop rapid and simple HPLC methods for measurement of BCAA, Met, Phe, Tyr and BCKA in plasma and dried blood samples. Samples of peripheral venous blood with EDTA as anticoagulant were obtained from a group of healthy blood donors (n=70, 35 females, 27-41 years of age and 35 males, 28-43 years of age). Blood-spot samples from a group of newborns (n=80, 40 girls and 40 boys 3-5 days of age) were collected onto #903 Specimen Collection Paper and allowed to dry for at least 24 h before analysis. Prior to separation, the amino acids (AA) were derivatized with o-phthaldialdehyde (OPA) and BCKA with o-phenylenediamine (OPD). Reverse phase column chromatography (LiChroCart 125-4 Purospher RP-18e, 5 microm) was used for separation and fluorescence detection used to monitoring of effluent. For AA analysis, 25 mmol/L sodium hydrogenphosphate-methanol (90:10, v/v), pH 6.5+/-0.1 was used as mobile phase A and 100% methanol was used as mobile phase B. Measurement of BCKA used a mixture of methanol and deionized water (55:45, v/v) as mobile phase A and mobile phase B consisted of 100% methanol. Analytical performance of these methods was satisfactory for the determination of all AA and BCKA. The intra-assay and inter-assay coefficients of variation were below 10% and recovery ranged from 90%-110%. We have developed simple, rapid and selective HPLC methods with fluorescence detection for the determination of BCAA, Met, Phe, Tyr and BCKA in plasma and dried blood samples.

Studies of U in the blood of two population samples

International Nuclear Information System (INIS)

Segovia, N.; Olguin, M.E.; Romero, M.

1986-01-01

The present work, attempts to establish the statistical distribution of blood uranium in a population of the same community, similar in age and in living patterns. U traces were evaluated by a fission track technique both in whole blood and plasma samples. Dried samples were compressed into pellets and irradiated in a nuclear reactor using the external detector method. For U quantification, standard U samples were used. A comparative sampling of U content in blood samples from a group of radiation exposed workers and another of leukemia patients was also carried out. Results from the sampling groups are reported and discussed. (author)

"Center punch" and "whole spot" bioanalysis of apixaban in human dried blood spot samples by UHPLC-MS/MS.

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Zheng, Naiyu; Yuan, Long; Ji, Qin C; Mangus, Heidi; Song, Yan; Frost, Charles; Zeng, Jianing; Aubry, Anne-Françoise; Arnold, Mark E

2015-04-15

Apixaban (Eliquis™) was developed by Bristol-Myers Squibb (BMS) and Pfizer to use as an antithrombotic/anticoagulant agent and has been recently approved for the prevention of stroke and systemic embolism in patients with nonvalvular atrial fibrillation. A clinical study of apixaban, sponsored by BMS and Pfizer, included a pilot exploratory portion to evaluate the potential for future drug concentration monitoring using dried blood spot (DBS) sample collection. For DBS sample collection, a fixed blood volume was dispensed onto a DBS card by either regular volumetric pipette (venous blood collection) or capillary dispenser (finger prick blood collection). A 96-well semi-automated liquid-liquid extraction sample preparation procedure was developed to provide clean extracts for UHPLC-MS/MS quantitation. Assays using both partial-spot center punch and whole spot punch were developed and validated. The linear dynamic ranges for all the analyses were from 0.5 to 500 ng/mL. The coefficient of determination (r(2)) values was >0.9944 for all the validation runs. For the center punch approach, the intra-assay precision (%CV) was within 4.4% and inter-assay precision was within 2.6%. The assay accuracy, expressed as %Dev., was within ± 5.4% of the nominal concentrations. One accuracy and precision run was performed using the whole spot approach, the intra-assay precision (%CV) was within 7.1% and the accuracy was within ± 8.0% of the nominal concentrations. In contrast to the center punch approach, the whole spot approach eliminated the effect of hematocrit and high lipids on the analysis of apixaban in human DBS when an accurate sample blood volume was collected on DBS cards. Copyright © 2015 Elsevier B.V. All rights reserved.

Evaluation of dried blood spot samples for hepatitis C virus detection and quantification.

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Marques, Brunna Lemos Crespo; do Espírito-Santo, Márcia Paschoal; Marques, Vanessa Alves; Miguel, Juliana Custódio; da Silva, Elisangela Ferreira; Villela-Nogueira, Cristiane Alves; Lewis-Ximenez, Lia Laura; Lampe, Elisabeth; Villar, Livia Melo

2016-09-01

Dried blood spots (DBS) could be an excellent alternative for HCV diagnosis, since it is less invasive and can be stored and transported without refrigeration. The aim of this study was to optimize quantitative and qualitative methods for HCV detection in DBS. DBS and serum samples were collected from 99 subjects (59 anti-HCV/HCV RNA positive and 40 seronegative samples). Seven extraction methods and different PCR parameters were evaluated in DBS samples in the quantitative RT-PCR (qRT-PCR) developed to amplify the 5' noncoding region of HCV. A qualitative PCR for amplification of NS5B region of HCV was also valued and the nested-PCR sequenced. The qRT-PCR showed good correlation to commercial assay for HCV viral measurement in serum. To quantify HCV RNA in DBS, it was necessary to increase reverse transcriptase and cDNA concentration. HCV RNA quantification in DBS demonstrated sensitivity of 65.9%, 100% of specificity and kappa statistic of 0.65. The median viral load of DBS samples was 5.38 log10 copies/ml (minimum value=1.76 and maximum value=10.48 log10 copies/ml). HCV RNA was detected in NS5B regions and nucleotide sequences obtained in 43 serum and 11 DBS samples. The presence of the same subtype was observed in paired serum and DBS samples. In this study, it was possible to demonstrate that, despite the low sensitivity, the optimized protocol was able to determine the viral load, as well as, the infecting HCV genotype, validating the usefulness of DBS for viral load determination and molecular epidemiology studies of HCV. Copyright © 2016 Elsevier B.V. All rights reserved.

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

Science.gov (United States)

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. Copyright © 2016 Elsevier B.V. All rights reserved.

Determination of Efavirenz in Human Dried Blood Spots by Reversed-Phase High Performance Liquid Chromatography with UV Detection

Science.gov (United States)

Hoffman, Justin T; Rossi, Steven S; Espina-Quinto, Rowena; Letendre, Scott; Capparelli, Edmund V

2013-01-01

Background Previously published methods for determination of efavirenz (EFV) in human dried blood spots (DBS) employ costly and complex liquid chromatography/mass spectrometry. We describe the validation and evaluation of a simple and inexpensive high-performance liquid chromatography (HPLC) method for EFV quantification in human DBS and dried plasma spots (DPS), using ultraviolet (UV) detection appropriate for resource-limited settings. Methods 100μl of heparinized whole blood or plasma were spotted onto blood collection cards, dried, punched, and eluted. Eluates are injected onto a C-18 reversed phase HPLC column. EFV is separated isocratically using a potassium phosphate and ACN mobile phase. UV detection is at 245nm. Quantitation is by use of external calibration standards. Following validation, the method was evaluated using whole blood and plasma from HIV-positive patients undergoing EFV therapy. Results Mean recovery of drug from dried blood spots is 91.5%. The method is linear over the validated concentration range of 0.3125 – 20.0μg/mL. A good correlation (Spearman r=0.96) between paired plasma and DBS EFV concentrations from the clinical samples was observed, and hematocrit level was not found to be a significant determinant of the EFV DBS level. The mean observed CDBS/Cplasma ratio was 0.68. A good correlation (Spearman r=0.96) between paired plasma and DPS EFV concentrations from the clinical samples was observed. The mean percent deviation of DPS samples from plasma samples is 1.68%. Conclusions Dried whole blood spot or dried plasma spot sampling is well suited for monitoring EFV therapy in resource limited settings, particularly when high sensitivity is not essential. PMID:23503446

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

Science.gov (United States)

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. Copyright © 2015 Elsevier B.V. All rights reserved.

Relocation of cryopreserved umbilical cord blood samples using a high-capacity dry shipper to a new laboratory: a cord blood banking experience.

Science.gov (United States)

Thiagarajah, Kalaivani; Wong, Chee-Yin; Vijayan, Vickneswary Veera; Ooi, Ghee-Chien; Ng, Mei-Theng; Cheong, Soon-Keng; Then, Kong-Yong

2015-05-01

Processed umbilical cord blood (UCB) must be stored at cryogenic temperature at all times to maintain the quality and viability of the cells. However, a challenge is presented in the form of moving a large number of cryopreserved UCB samples to a new location. In this report, we share our experience on relocating more than 100,000 units of cryopreserved UCB samples stored in 12 liquid nitrogen freezers (LNFs) to our new laboratory. For quality control purposes, 2 weeks before relocation, donor UCB samples were processed, cryopreserved, and stored in each LNF. On relocation day, half of the samples were retrieved to determine total nucleated cell count, percentage of CD34+ cells, and cell viability as controls for later comparison. UCB samples were transferred into dry shippers before being relocated to the new laboratory. Upon arrival, LNFs were serviced before transferring UCB samples back into its original location within the LNF. The remaining donor UCB samples were retrieved and analyzed for the same tests mentioned. We found no significant differences in pre- and postrelocation values of the tests performed. All UCB samples were successfully relocated into the new laboratory without affecting the quality. © 2014 AABB.

Spreading of blood drops over dry porous substrate: complete wetting case.

Science.gov (United States)

Chao, Tzu Chieh; Arjmandi-Tash, Omid; Das, Diganta B; Starov, Victor M

2015-05-15

The process of dried blood spot sampling involves simultaneous spreading and penetration of blood into a porous filter paper with subsequent evaporation and drying. Spreading of small drops of blood, which is a non-Newtonian liquid, over a dry porous layer is investigated from both theoretical and experimental points of view. A system of two differential equations is derived, which describes the time evolution of radii of both the drop base and the wetted region inside the porous medium. The system of equations does not include any fitting parameters. The predicted time evolutions of both radii are compared with experimental data published earlier. For a given power law dependency of viscosity of blood with different hematocrit level, radii of both drop base and wetted region, and contact angle fell on three universal curves if appropriate scales are used with a plot of the dimensionless radii of the drop base and the wetted region inside the porous layer and dynamic contact angle on dimensionless time. The predicted theoretical relationships are three universal curves accounting satisfactorily for the experimental data. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

[Prospects of the integration of dry blood spot technology with human health and environmental population studies].

Science.gov (United States)

Pomelova, V G; Osin, N S

2007-01-01

This literature review is dedicated to prospects of the use of whole blood dried on special filter paper as a source of biological material for human health and environmental population studies. Evident advantages of this low-invasive approach include the following: it is easy to take a blood sample from a patient's finger ofa neonate's heel; the cost of sampling as well as transportation and storage of samples is low; paper blanks are safe to manipulate with and convenient to mail in sealed plastic packages. Many analytes, such as DNA, become more stable after drying, which allows for the detection of phenotypic and genotypic markers, as well as multiple gene mutations by multiplex DNA amplification. Modern diagnostic techniques make it possible to detect a wide spectrum of biomarkers characterizing the condition of the endocrine, cardiovascular, reproductive, and immune systems of the organism in a single drop of blood. This allows considering paper blanks with dry blood the key component of multilevel interdisciplinary population studies on neonatal screening, disease spread surveillance, seroepidemiological monitoring, and ecological and genetic research.

An automated immunoradiometric assay of thyrotrophin (TSH) in dried blood filter paper spots

International Nuclear Information System (INIS)

John, R.; Woodhead, J.S.

1982-01-01

An immunoradiometric two-site assay for thyrotrophin (TSH) in dried blood filter paper spots is described. The assay is automated by means of the Kemtek 3000 automated immunoassay system. The technique uses a 6.0 mm disc punched from the dried blood samples collected as part of the screening programme for phenylketonuria. The method is sensitive and precise, and results correlate well with those obtained in TSH assays of serum samples. The procedure is rapid, results being available within 24 h of receipt of samples. Of 25204 specimens so far screened by this assay, 99.9% have TSH levels less than 15 mU/l. One false positive result has been obtained and six confirmed cases of neonatal hypothyroidism detected, giving a prevalence of 1 in 4200. (Auth.)

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

Directory of Open Access Journals (Sweden)

Laura Russo Zillmer

2013-09-01

Full Text Available 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.

Performance of the biomerieux DBS puncher and dried blood spots ...

African Journals Online (AJOL)

Introduction: The latest World Health Organization recommendations request viral load (VL) testing, if possible, for monitoring HIV-1 infections. However, the use of plasma is an obstacle to realize this test in sub-Saharan Africa. In this context, the dried blood spot (DBS) is an interesting tool for sample collections.

A simple method to quantitate IP-10 in dried blood and plasma spots.

Directory of Open Access Journals (Sweden)

Martine G Aabye

Full Text Available BACKGROUND: Antigen specific release of IP-10 is an established marker for infection with M.tuberculosis. Compared to IFN-γ, IP-10 is released in 100-fold higher concentrations enabling the development of novel assays for detection. Dried blood spots are a convenient sample for high throughput newborn screening. AIM: To develop a robust and sensitive ELISA-based assay for IP-10 detection in plasma, dried blood spots (DBS and dried plasma spots (DPS; to validate the ELISA in clinically relevant samples; and to assess the performance of the assay for detection of Cytomegalovirus (CMV and M.tuberculosis specific immune responses. METHOD: We raised mice and rat monoclonal antibodies against human IP-10 and developed an ELISA. The assay was validated and applied to the detection of CMV and M.tuberculosis specific responses in 18 patients with immune reactivity towards M.tuberculosis and 32 healthy controls of which 22 had immune reactivity towards CMV and none towards M.tuberculosis. We compared the performance of this new assay to IFN-γ. RESULTS: The ELISA was reliable for IP-10 detection in both plasma and filter paper samples. The linear range of the ELISA was 2.5-600 pg/ml. IFN-γ was not readily detectable in DPS samples. IP-10 was stabile in filter paper samples for at least 4 weeks at 37 °C. The correlation between IP-10 detected in plasma, DPS and DBS samples was excellent (r(2>0.97. CONCLUSIONS: This newly developed assay is reliable for IP-10 quantification in plasma, DBS and DPS samples from antigen stimulated and non-stimulated whole blood. The filter paper assays enable easy sample acquisition and transport at ambient temperature e.g. via the postal system. The system can potentially simplify diagnostic assays for M.tuberculosis and CMV infection.

The influence of the dried blood spot drying time on the recoveries of six immunosuppressants

Directory of Open Access Journals (Sweden)

Remco A. Koster

2015-10-01

Full Text Available Investigation of the drying time of dried blood spots (DBS is currently not included in DBS validations. The influence of the DBS drying time on the recovery of tacrolimus, ascomycin, sirolimus, everolimus, cyclosporin A and temsirolimus was evaluated by measuring DBS with a fixed blood volume at a hematocrit range between 0.1 and 0.6 L/L at 3, 24 and 48 hours of drying time. Results showed that the recovery of sirolimus, everolimus, temsirolimus and cyclosporin A was influenced by the DBS drying time, while the recovery of tacrolimus and ascomycin was not. A drying time of at least 24 hours is advised in order to stabilize hematocrit and concentration related recovery effects of sirolimus, everolimus, temsirolimus and cyclosporin A.

Analysis of the stability of urea in dried blood spots collected and stored on filter paper.

Science.gov (United States)

Quraishi, Rizwana; Lakshmy, Ramakrishnan; Mukhopadhyay, Ashok Kumar; Jailkhani, Bansi Lal

2013-05-01

The ability to use dry blood spots (DBSs) on filter paper for the analysis of urea levels could be an important diagnostic tool for areas that have limited access to laboratory facilities. We developed a method for the extraction and quantification of urea from DBSs that were stored on 3M Whatman filter paper and investigated the effect of long-term storage on the level of urea in DBSs. DBSs of 4.5 mm in diameter were used for our assay, and we determined the urea levels in blood using a commercially available enzymatic kit (UV GLDH-method; Randox laboratories Ltd., UK). The DBSs on filter discs were stored at 4℃ or at 37℃ for 120 days. The mean intra- and inter-assay coefficient of variance for our method of urea extraction from dried blood was 4.2% and 6.3%, respectively. We collected 75 fresh blood samples and compared the urea content of each fresh sample with the urea content of DBSs taken from corresponding fresh blood samples. Regression analysis reported a regression coefficient (r) value of 0.97 and a recovery of urea from dried spots was 102.2%. Urea concentrations in DBSs were stable for up to 120 and 90 days when stored at 4℃ and 37℃, respectively. Our results show that urea can be stored and quantitatively recovered from small volumes of blood that was collected on filter paper.

A Novel, Nondestructive, Dried Blood Spot-Based Hematocrit Prediction Method Using Noncontact Diffuse Reflectance Spectroscopy

NARCIS (Netherlands)

Capiau, S.; Wilk, L.S.; Aalders, M.C.G.; Stove, C.P.

2016-01-01

Dried blood spot (DBS) sampling is recognized as a valuable alternative sampling strategy both in research and in clinical routine. Although many advantages are associated with DBS sampling, its more widespread use is hampered by several issues, of which the hematocrit effect on DBS-based

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

Science.gov (United States)

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

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

Science.gov (United States)

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

Water-Soluble Dried Blood Spot in Protein Analysis: A Proof-of-Concept Study.

Science.gov (United States)

Rosting, Cecilie; Gjelstad, Astrid; Halvorsen, Trine Grønhaug

2015-08-04

In the present work human chorionic gonadotropin (hCG) was used as a model protein in a proof-of-concept study combining water-soluble dried blood spot (DBS) material in liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based protein analysis. A water-soluble material consisting of commercially available carboxymethyl cellulose (CMC) was evaluated as sampling material for this purpose. The material dissolved readily at physiological pH. Different sample preparation methods were evaluated, and in the final method, 15 μL of whole blood was deposited and dried on CMC before the whole spot was dissolved prior to cleanup by immunoaffinity extraction, tryptic digest, and preconcentration by solid-phase extraction (SPE). The results indicated complete dissolution of hCG from the spots, acceptable limit of detection (LOD) (0.1 IU/mL), linearity (R(2) = 0.959), accuracy (16%), and precision (≤22%). Long-term stability (45 days) of hCG in dried spots at reduced temperatures (≤8 °C) was also demonstrated. The analyte recovery was comparable to the commercially available nonsolvable cellulose material (FTA DMPK-C card).

Radioimmunoassay of alpha-foetoprotein in the eluate of dried blood. A method for antenatal screening of neural tube defects

International Nuclear Information System (INIS)

Travert, G.; Herlicoviez, M.; Laroche, D.

1979-01-01

A radioimmunoassay for alpha-foetoprotein (AFP) in dried blood spots is reported. The main technical characteristics (reproducibility, sensitivity, recovery of exogenous AFP added and AFP stability in dried blood) are evaluated. They indicate that this method is feasible and well adapted to AFP measurement during pregnancy. AFP determination in maternal serum allows early detection of at least 80% of neural tube defects. The use of dried blood spots as samples for AFP assay makes our method a possible mass screening test for these malformations, which occur with an incidence of 12 for 10,000 [fr

Studies of an alpha-fetoprotein assay using dry blood-spot samples to be used for the detection of fetal neural tube defects.

Science.gov (United States)

Wong, P Y; Mee, A V; Doran, T A

1982-06-01

We modified the Pharmacia serum alpha-fetoprotein (AFP) kit to enable its use with dry blood-spots on filter paper. Reference values were established for blood from 253 women in the 16th to 18th weeks of gestation. The result by the present technique in a woman with a confirmed anencephalic fetus was elevated, and in agreement with the results of AFP assays in serum and amniotic fluid. Blood AFP was stable on dried filter paper sent by mail.

Contamination of dried blood spots - an underestimated risk in newborn screening.

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Winter, Theresa; Lange, Anja; Hannemann, Anke; Nauck, Matthias; Müller, Cornelia

2018-01-26

Newborn screening (NBS) is an established screening procedure in many countries worldwide, aiming at the early detection of inborn errors of metabolism. For decades, dried blood spots have been the standard specimen for NBS. The procedure of blood collection is well described and standardized and includes many critical pre-analytical steps. We examined the impact of contamination of some anticipated common substances on NBS results obtained from dry spot samples. This possible pre-analytical source of uncertainty has been poorly examined in the past. Capillary blood was obtained from 15 adult volunteers and applied to 10 screening filter papers per volunteer. Nine filter papers were contaminated without visible trace. The contaminants were baby diaper rash cream, baby wet wipes, disinfectant, liquid infant formula, liquid infant formula hypoallergenic (HA), ultrasonic gel, breast milk, feces, and urine. The differences between control and contaminated samples were evaluated for 45 NBS quantities. We estimated if the contaminations might lead to false-positive NBS results. Eight of nine investigated contaminants significantly altered NBS analyte concentrations and potentially caused false-positive screening outcomes. A contamination with feces was most influential, affecting 24 of 45 tested analytes followed by liquid infant formula (HA) and urine, affecting 19 and 13 of 45 analytes, respectively. A contamination of filter paper samples can have a substantial effect on the NBS results. Our results underline the importance of good pre-analytical training to make the staff aware of the threat and ensure reliable screening results.

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

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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. Copyright © 2015 John Wiley & Sons, Ltd.

Validity and Reliability of Perinatal Biomarkers after Storage as Dry Blood Spots on Paper

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Mihalopoulos, Nicole L.; Phillips, Terry M.; Slater, Hillarie; Thomson, J. Anne; Varner, Michael W.; Moyer-Mileur, Laurie J.

2013-01-01

Ojective To validate use of chip-based immunoaffinity capillary electrophoresis on dry blood spot samples (DBSS) to measure obesity-related cytokines. Methods Chip-based immunoaffinity capillary electrophoresis was used to measure adiponectin, leptin and insulin in serum and DBSS in pregnant women, cord blood, and infant heelstick at birth and 6 weeks. Concordance of measurements was determined with Pearson's correlation. Results We report high concordance between results obtained from serum and DBSS with the exception of cord blood specimens. Conclusions Ease of sample collection and storage makes DBSS an optimal method for use in studies involving neonates and young children. PMID:21735507

A novel approach for quantitation of glucosylceramide in human dried blood spot using LC-MS/MS.

Science.gov (United States)

Ji, Allena Ji; Wang, Haixing; Ziso-Qejvanaj, Enida; Zheng, Kefei; Chung, Lee Lee; Foley, Timothy; Chuang, Wei-Lien; Richards, Susan; Sung, Crystal

2015-01-01

Glucosylceramide, an efficacy biomarker for Gaucher Type 1 disease, exhibits poor solubility in polar solvents and whole blood which makes it difficult to prepare a homogenous blood standard. We developed a novel method using standard addition approach by spiking a small volume of analyte solution on the surface of prespotted dried blood spot. The whole spots were punched out for subsequent extraction and LC-MS/MS analysis. The assay performance met all validation acceptance criteria. Glucosylceramide concentrations in 50 paired plasma and dry blood spot samples obtained from Gaucher Type 1 patients were tested and the results demonstrated the feasibility of using the DBS method for clinical biomarker monitoring. The new approach greatly improves assay precision and accuracy.

Serum Dried Samples to Detect Dengue Antibodies: A Field Study

Directory of Open Access Journals (Sweden)

Angelica Maldonado-Rodríguez

2017-01-01

Full Text Available Background. Dried blood and serum samples are useful resources for detecting antiviral antibodies. The conditions for elution of the sample need to be optimized for each disease. Dengue is a widespread disease in Mexico which requires continuous surveillance. In this study, we standardized and validated a protocol for the specific detection of dengue antibodies from dried serum spots (DSSs. Methods. Paired serum and DSS samples from 66 suspected cases of dengue were collected in a clinic in Veracruz, Mexico. Samples were sent to our laboratory, where the conditions for optimal elution of DSSs were established. The presence of anti-dengue antibodies was determined in the paired samples. Results. DSS elution conditions were standardized as follows: 1 h at 4°C in 200 µl of DNase-, RNase-, and protease-free PBS (1x. The optimal volume of DSS eluate to be used in the IgG assay was 40 µl. Sensitivity of 94%, specificity of 93.3%, and kappa concordance of 0.87 were obtained when comparing the antidengue reactivity between DSSs and serum samples. Conclusion. DSS samples are useful for detecting anti-dengue IgG antibodies in the field.

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

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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. Copyright © 2012 John Wiley & Sons, Ltd.

Advantages and Challenges of Dried Blood Spot Analysis by Mass Spectrometry Across the Total Testing Process

Science.gov (United States)

Zakaria, Rosita; Allen, Katrina J.; Koplin, Jennifer J.; Roche, Peter

2016-01-01

Introduction Through the introduction of advanced analytical techniques and improved throughput, the scope of dried blood spot testing utilising mass spectrometric methods, has broadly expanded. Clinicians and researchers have become very enthusiastic about the potential applications of dried blood spot based mass spectrometric applications. Analysts on the other hand face challenges of sensitivity, reproducibility and overall accuracy of dried blood spot quantification. In this review, we aim to bring together these two facets to discuss the advantages and current challenges of non-newborn screening applications of dried blood spot quantification by mass spectrometry. Methods To address these aims we performed a key word search of the PubMed and MEDLINE online databases in conjunction with individual manual searches to gather information. Keywords for the initial search included; “blood spot” and “mass spectrometry”; while excluding “newborn”; and “neonate”. In addition, databases were restricted to English language and human specific. There was no time period limit applied. Results As a result of these selection criteria, 194 references were identified for review. For presentation, this information is divided into: 1) clinical applications; and 2) analytical considerations across the total testing process; being pre-analytical, analytical and post-analytical considerations. Conclusions DBS analysis using MS applications is now broadly applied, with drug monitoring for both therapeutic and toxicological analysis being the most extensively reported. Several parameters can affect the accuracy of DBS measurement and further bridge experiments are required to develop adjustment rules for comparability between dried blood spot measures and the equivalent serum/plasma values. Likewise, the establishment of independent reference intervals for dried blood spot sample matrix is required. PMID:28149263

The use of mass spectrometry to analyze dried blood spots.

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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

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

Energy Technology Data Exchange (ETDEWEB)

Kyle, Jennifer E. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Casey, Cameron P. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Stratton, Kelly G. [National Security Directorate, Pacific Northwest National Laboratory, Richland WA USA; Zink, Erika M. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Kim, Young-Mo [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Zheng, Xueyun [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Monroe, Matthew E. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Weitz, Karl K. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Bloodsworth, Kent J. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Orton, Daniel J. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Ibrahim, Yehia M. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Moore, Ronald J. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Lee, Christine G. [Department of Medicine, Bone and Mineral Unit, Oregon Health and Science University, Portland OR USA; Research Service, Portland Veterans Affairs Medical Center, Portland OR USA; Pedersen, Catherine [Department of Medicine, Bone and Mineral Unit, Oregon Health and Science University, Portland OR USA; Orwoll, Eric [Department of Medicine, Bone and Mineral Unit, Oregon Health and Science University, Portland OR USA; Smith, Richard D. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Burnum-Johnson, Kristin E. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA; Baker, Erin S. [Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, Richland WA USA

2017-02-05

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

A simple method to quantitate IP-10 in dried blood and plasma spots

DEFF Research Database (Denmark)

Aabye, Martine G; Eugen-Olsen, Jesper; Werlinrud, Anne Marie

2012-01-01

Antigen specific release of IP-10 is an established marker for infection with M.tuberculosis. Compared to IFN-γ, IP-10 is released in 100-fold higher concentrations enabling the development of novel assays for detection. Dried blood spots are a convenient sample for high throughput newborn...

Dried blood spots, valid screening for viral hepatitis and human immunodeficiency virus in real-life

DEFF Research Database (Denmark)

Mössner, Belinda K; Staugaard, Benjamin; Jensen, Janne

2016-01-01

AIM: To detect chronic hepatitis B (CHB), chronic hepatitis C (CHC) and human immunodeficiency virus (HIV) infections in dried blood spot (DBS) and compare these samples to venous blood sampling in real-life. METHODS: We included prospective patients with known viral infections from drug treatment......, but correctly classified 95% of the anti-HCV-positive patients with chronic and past infections. Anti-HBc and anti-HBS showed low sensitivity in DBS (68% and 42%). CONCLUSION: DBS sampling, combined with an automated analysis system, is a feasible screening method to diagnose chronic viral hepatitis and HIV...

Effect of ambient humidity on the rate at which blood spots dry and the size of the spot produced.

Science.gov (United States)

Denniff, Philip; Woodford, Lynsey; Spooner, Neil

2013-08-01

For shipping and storage, dried blood spot (DBS) samples must be sufficiently dry to protect the integrity of the sample. When the blood is spotted the humidity has the potential to affect the size of the spot created and the speed at which it dries. The area of DBS produced on three types of substrates were not affected by the humidity under which they were generated. DBS samples reached a steady moisture content 150 min after spotting and 90 min for humidities less than 60% relative humidity. All packaging materials examined provided some degree of protection from external extreme conditions. However, none of the packaging examined provided a total moisture barrier to extreme environmental conditions. Humidity was shown not to affect the spot area and DBS samples were ready for shipping and storage 2 h after spotting. The packing solutions examined all provided good protection from external high humidity conditions.

Diagnostic accuracy of serological diagnosis of hepatitis C and B using dried blood spot samples (DBS): two systematic reviews and meta-analyses.

Science.gov (United States)

Lange, Berit; Cohn, Jennifer; Roberts, Teri; Camp, Johannes; Chauffour, Jeanne; Gummadi, Nina; Ishizaki, Azumi; Nagarathnam, Anupriya; Tuaillon, Edouard; van de Perre, Philippe; Pichler, Christine; Easterbrook, Philippa; Denkinger, Claudia M

2017-11-01

Dried blood spots (DBS) are a convenient tool to enable diagnostic testing for viral diseases due to transport, handling and logistical advantages over conventional venous blood sampling. A better understanding of the performance of serological testing for hepatitis C (HCV) and hepatitis B virus (HBV) from DBS is important to enable more widespread use of this sampling approach in resource limited settings, and to inform the 2017 World Health Organization (WHO) guidance on testing for HBV/HCV. We conducted two systematic reviews and meta-analyses on the diagnostic accuracy of HCV antibody (HCV-Ab) and HBV surface antigen (HBsAg) from DBS samples compared to venous blood samples. MEDLINE, EMBASE, Global Health and Cochrane library were searched for studies that assessed diagnostic accuracy with DBS and agreement between DBS and venous sampling. Heterogeneity of results was assessed and where possible a pooled analysis of sensitivity and specificity was performed using a bivariate analysis with maximum likelihood estimate and 95% confidence intervals (95%CI). We conducted a narrative review on the impact of varying storage conditions or limits of detection in subsets of samples. The QUADAS-2 tool was used to assess risk of bias. For the diagnostic accuracy of HBsAg from DBS compared to venous blood, 19 studies were included in a quantitative meta-analysis, and 23 in a narrative review. Pooled sensitivity and specificity were 98% (95%CI:95%-99%) and 100% (95%CI:99-100%), respectively. For the diagnostic accuracy of HCV-Ab from DBS, 19 studies were included in a pooled quantitative meta-analysis, and 23 studies were included in a narrative review. Pooled estimates of sensitivity and specificity were 98% (CI95%:95-99) and 99% (CI95%:98-100), respectively. Overall quality of studies and heterogeneity were rated as moderate in both systematic reviews. HCV-Ab and HBsAg testing using DBS compared to venous blood sampling was associated with excellent diagnostic accuracy

Surface fixation of dried blood by glutaraldehyde and peracetic acid.

Science.gov (United States)

Kampf, G; Bloss, R; Martiny, H

2004-06-01

The difficulties of successful prion inactivation by chemical agents has led to changes in recommendations regarding the reprocessing of instruments including flexible endoscopes. One of the changes is the preference for peracetic acid instead of glutaraldehyde in order to avoid fixation of organic material, but the surface fixation by various active agents has not been fully investigated. We used a standardized amount of dried blood soil on metal carriers (on average 22 mg). One part of the carriers was exposed to different disinfectants (four based on peracetic acid, three based on glutaraldehyde, two based on quaternary ammonium compounds (QAC), one based on QAC and amines, one based on phenols and one cleaning agent) and air dried. The difference compared with the non-exposed soiled carrier was taken as the measure of blood removal by exposure to the disinfectants. In addition the other part of the carriers was exposed to a cleaning agent and air dried. The cleaning agent itself was capable of removing more than 99% of the dried blood and served as a control for non-fixation. The rate of fixation of dried blood was calculated as the ratio of the weight of residual soil on 'soiled, disinfected and cleaned' carriers and on 'soiled and disinfected' carriers. All experiments were repeated eight times. Blood removal varied between 90.3% +/- 1.5% (phenol-based disinfectant) and peracetic acid. No other preparations showed a potential for blood fixation (peracetic acid, and support the evidence that effective cleaning should precede the chemical disinfection. Copyright 2004 The Hospital Infection Society

Dried blood spots voor het bepalen van de serumconcentratie van tamoxifen en zijn actieve metaboliet endoxifen

NARCIS (Netherlands)

Jager, Nynke G L; Rosing, Hilde; Schellens, Jan H M; Beijnen, Jos H.; Linn, Sabine C.

2016-01-01

OBJECTIVE To establish the relationship between dried blood spot (DBS) and serum concentrations of tamoxifen and endoxifen in order to allow the use of DBS sampling, a simple and patient-friendly alternative to venous sampling, in clinical practice. The antiestrogenic effect of tamoxifen is

A filter paper dry blood spot procedure for acute intermittent porphyria population screening by use of whole blood uroporphyrinogen-I-synthase assay.

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Johansson, L; Thunell, S; Wetterberg, L

1984-03-13

A filter paper dry blood spot procedure for the determination of whole blood uroporphyrinogen-I-synthase (UIS) activity is presented. The method is based on the concept of enzyme specific activity, the enzyme activity being related to the haemoglobin concentration of the assay sample. The diagnostic capacity with regard to the acute intermittent porphyria (AIP) gene carrier state is shown to be equivalent to that of a washed red cell reference method. On grounds of easy capillary blood sampling, uncomplicated and safe mail specimen transport and simple laboratory reception routines, the method is stated to be well adapted for use in AIP preadolescent population screening.

Stability of Phosphatidylethanol in Dry Blood Spot Cards.

Science.gov (United States)

Bakhireva, Ludmila N; Shrestha, Shikhar; Gutierrez, Hilda L; Berry, Mike; Schmitt, Cheryl; Sarangarm, Dusadee

2016-05-01

The analysis of phosphatidylethanol, a promising direct ethanol metabolite, in dry blood spots (PEth-DBS) is advantageous due to ease of storage, transportation and minimal invasiveness of capillary blood collection. One potential application of PEth-DBS is to confirm prenatal alcohol exposure in newborns suspected of FASD; however, stability of PEth-DBS is largely unknown. Phlebotomized samples from 31 adults with a history of alcoholism, admitted to the University of New Mexico Emergency Department, were analyzed for blood alcohol content and pipetted onto DBS cards (13 spots per patient). The first spot was analyzed within 2 weeks of collection for a baseline PEth; the remaining 12 spots were allocated into three temperature conditions (room temperature, 4°C, -80°C) for the repeated measures analysis. In addition, 5 newborn DBS samples with a baseline PEth>LOD were obtained from a prospective cohort at UNM and re-analyzed at 4 months after storage at -80°C. A mixed linear model was fitted to examine the effects of temperature, time and temperature-time interaction on PEth degradation over the first 9 months. The baseline PEth levels were 592.8 ± 86.7 ng/ml and 18.3 ± 4.8 ng/ml in adult and newborn samples, respectively. All DBS samples remained positive in successive samples in all temperature conditions. Results of mixed linear model demonstrated a significant effect of temperature (P < 0.001) on PEth degradation over 9 months. PEth-DBS appears to be relatively stable, especially when stored at lower temperatures. These initial results are encouraging and highlight the PEth-DBS potential in retrospective assessment of alcohol exposure. © The Author 2015. Medical Council on Alcohol and Oxford University Press. All rights reserved.

Apparatus for freeze drying of biologic and sediment samples

International Nuclear Information System (INIS)

Anon.

1978-01-01

Freeze drying to obtain water from individual samples, though not complicated, usually requires considerable effort to maintain the cold traps on a 24-hr basis. In addition, the transfer of a sample from sample containers to freeze-dry flasks is usually made with some risk of contamination to the sample. If samples are large, 300 g to 600 g, usually several days are required to dry the samples. The use of an unattended system greatly improves personnel and drying efficiency. Commercial freeze dryers are not readily applicable to the problems of collecting water from individual samples, and lab-designed collectors required sample transfer and continual replenishment of the dry ice. A freeze-dry apparatus for collecting water from individual sediment and/or biological samples was constructed to determine the tritium concentrations in fish for dose calcaluations and the tritium distribution in sediment cores for water movement studies. The freeze, dry apparatus, which can handle eight samples simultaneously and conveniently, is set up for unattended 24-hr operation and is designed to avoid sample transfer problems

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

Science.gov (United States)

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. Copyright © 2012 Elsevier B.V. All rights reserved.

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

DEFF Research Database (Denmark)

Poulsen, Jesper Buchhave; Lescai, Francesco; Grove, Jakob

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...

Identification of bovine material in porcine spray-dried blood derivatives using the Polymerase Chain Reaction technique

Directory of Open Access Journals (Sweden)

Sánchez A.

2004-01-01

Full Text Available Due to the widely supported theory of bovine spongiform encephalopathy (BSE spread in cattle by contaminated animal feeds, screening of feed products has become essential. For many years, manufacturers have used blood and plasma proteins as high quality ingredients of foods for both pets and farm animals. However, in Europe, the Commission Regulation 1234/2003/EC temporally bans the use of processed animal proteins, including blood-derivative products, in feedstuffs for all farm animals which are fattened or bred for the production of food. This regulation has some exceptions, such as the use of non ruminant blood products into the feed of farm fish. Authorization of the re-introduction of these proteins into animal feed formulations, especially non ruminant proteins into the feed for non ruminant farm animals, is expected when adequate control methods to discriminate ruminant proteins exist. Currently, the number of validated methods to differentiate the species of origin for most of the animal by-products is limited. Here we report the development of a rapid and sensitive polymerase chain reaction (PCR-based assay, which allows detection of bovine or porcine specific mitochondrial DNAfrom spray-dried blood derivate products (plasma, whole blood and red cells, as a marker for bovine contamination in porcine products. Sample extracts, suitable for PCR, were easily and quickly obtained with the commercial PrepManTM Ultra reagent (Applied Biosystems. To confirm the porcine origin of the samples, primers targeting a specific region of 134 bp of the porcine cytochrome b coding sequence were designed (cytbporc1-F and cytbporc2-R. Previously published PCR primers (L8129 and H8357, specific for a 271 bp fragment of the bovine mitochondrial ATPase 8-ATPase 6 genes, were chosen to accomplish amplification of bovine DNA. The limit of detection (LOD of the bovine PCR assay was at least of 0.05% (v/v of bovine inclusion in spray-dried porcine plasma or red

Manual versus automated blood sampling

DEFF Research Database (Denmark)

Teilmann, A C; Kalliokoski, Otto; Sørensen, Dorte B

2014-01-01

Facial vein (cheek blood) and caudal vein (tail blood) phlebotomy are two commonly used techniques for obtaining blood samples from laboratory mice, while automated blood sampling through a permanent catheter is a relatively new technique in mice. The present study compared physiological parameters......, glucocorticoid dynamics as well as the behavior of mice sampled repeatedly for 24 h by cheek blood, tail blood or automated blood sampling from the carotid artery. Mice subjected to cheek blood sampling lost significantly more body weight, had elevated levels of plasma corticosterone, excreted more fecal...... corticosterone metabolites, and expressed more anxious behavior than did the mice of the other groups. Plasma corticosterone levels of mice subjected to tail blood sampling were also elevated, although less significantly. Mice subjected to automated blood sampling were less affected with regard to the parameters...

Confirmation of congenital adrenal hyperplasia by adrenal steroid profiling of filter paper dried blood samples using ultra-performance liquid chromatography-tandem mass spectrometry.

Science.gov (United States)

Rossi, Claudia; Calton, Lisa; Brown, Heather A; Gillingwater, Scott; Wallace, A Michael; Petrucci, Francesca; Ciavardelli, Domenico; Urbani, Andrea; Sacchetta, Paolo; Morris, Michael

2011-04-01

The specificity of screening for congenital adrenal hyperplasia by direct measurement of 17-hydroxyprogesterone in filter paper dried blood spot samples by immunoassay is low and has a high false-positive rate. In order to reduce the false-positive rate of this test, we developed a rapid, robust, specific confirmatory procedure in which cortisol, 4-androstene-3,17-dione and 17-hydroxyprogesterone were measured simultaneously by ultra-performance liquid chromatography-tandem mass spectrometry. After extraction, samples were analysed by ultra-performance liquid chromatography-tandem mass spectrometry and 17-hydroxyprogesterone was quantified accurately. Other steroids were determined using stable deuterated internal standards. In total, 25 patient blood spot samples and 92 control samples were analysed. The assay was linear for 17-hydroxyprogesterone, with a coefficient of determination >0.997 and imprecision ≤ 6.5%. An upper limit of normal for 17-hydroxyprogester-one of 4.45 nmol/L was established by analysing a cohort of samples from unaffected newborns. In addition, a cut-off of 3.5 for the peak areas ratio (17-hydroxyprogesterone+4-androstene-3,17-dione)/cortisol, allows confirmation of the affected steroidogenic enzyme. A high throughput method for the detection of steroids related to congenital adrenal hyperplasia has been developed, allowing the false-positive rate associated with screening for 17-hydroxyprogesterone by immunoassay to be determined.

A rapid liquid chromatography tandem mass spectrometry-based method for measuring propranolol on dried blood spots.

Science.gov (United States)

Della Bona, Maria Luisa; Malvagia, Sabrina; Villanelli, Fabio; Giocaliere, Elisa; Ombrone, Daniela; Funghini, Silvia; Filippi, Luca; Cavallaro, Giacomo; Bagnoli, Paola; Guerrini, Renzo; la Marca, Giancarlo

2013-05-05

Propranolol, a non-selective beta blocker drug, is used in young infants and newborns for treating several heart diseases; its pharmacokinetics has been extensively evaluated in adult patients using extrapolation to treat pediatric population. The purpose of the present study was to develop and validate a method to measure propranolol levels in dried blood spots. The analysis was performed by using liquid chromatography/tandem mass spectrometry operating in multiple reaction monitoring mode. The calibration curve in matrix was linear in the concentration range of 2.5-200 μg/L with correlation coefficient r=0.9996. Intra-day and inter-day precisions and biases were less than 8.0% (n=10) and 11.5% (n=10) respectively. The recoveries ranged from 94 to 100% and the matrix effect did not result in a severe signal suppression. Propranolol on dried blood spot showed a good stability at three different temperatures for one month. This paper describes a micromethod for measuring propranolol levels on dried blood spot, which determines a great advantage in neonates or young infants during pharmacokinetic studies because of less invasive sampling and small blood volume required. Copyright © 2013 Elsevier B.V. All rights reserved.

Dried Blood Spot Analysis Suitable for Therapeutic Drug Monitoring of Voriconazole, Fluconazole, and Posaconazole

Science.gov (United States)

van der Elst, Kim C. M.; Span, Lambert F. R.; van Hateren, Kai; Vermeulen, Karin M.; van der Werf, Tjip S.; Greijdanus, Ben; Kosterink, Jos G. W.; Uges, Donald R. A.

2013-01-01

Invasive aspergillosis and candidemia are important causes of morbidity and mortality in immunocompromised and critically ill patients. The triazoles voriconazole, fluconazole, and posaconazole are widely used for the treatment and prophylaxis of these fungal infections. Due to the variability of the pharmacokinetics of the triazoles among and within individual patients, therapeutic drug monitoring is important for optimizing the efficacy and safety of antifungal treatment. A dried blood spot (DBS) analysis was developed and was clinically validated for voriconazole, fluconazole, and posaconazole in 28 patients. Furthermore, a questionnaire was administered to evaluate the patients' opinions of the sampling method. The DBS analytical method showed linearity over the concentration range measured for all triazoles. Results for accuracy and precision were within accepted ranges; samples were stable at room temperature for at least 12 days; and different hematocrit values and blood spot volumes had no significant influence. The ratio of the drug concentration in DBS samples to that in plasma was 1.0 for voriconazole and fluconazole and 0.9 for posaconazole. Sixty percent of the patients preferred DBS analysis as a sampling method; 15% preferred venous blood sampling; and 25% had no preferred method. There was significantly less perception of pain with the DBS sampling method (P = 0.021). In conclusion, DBS analysis is a reliable alternative to venous blood sampling and can be used for therapeutic drug monitoring of voriconazole, fluconazole, and posaconazole. Patients were satisfied with DBS sampling and had less pain than with venous sampling. Most patients preferred DBS sampling to venous blood sampling. PMID:23896473

Hyperspectral imaging and multivariate analysis in the dried blood spots investigations

Science.gov (United States)

Majda, Alicja; Wietecha-Posłuszny, Renata; Mendys, Agata; Wójtowicz, Anna; Łydżba-Kopczyńska, Barbara

2018-04-01

The aim of this study was to apply a new methodology using the combination of the hyperspectral imaging and the dry blood spot (DBS) collecting. Application of the hyperspectral imaging is fast and non-destructive. DBS method offers the advantage also on the micro-invasive blood collecting and low volume of required sample. During experimental step, the reflected light was recorded by two hyperspectral systems. The collection of 776 spectral bands in the VIS-NIR range (400-1000 nm) and 256 spectral bands in the SWIR range (970-2500 nm) was applied. Pixel has the size of 8 × 8 and 30 × 30 µm for VIS-NIR and SWIR camera, respectively. The obtained data in the form of hyperspectral cubes were treated with chemometric methods, i.e., minimum noise fraction and principal component analysis. It has been shown that the application of these methods on this type of data, by analyzing the scatter plots, allows a rapid analysis of the homogeneity of DBS, and the selection of representative areas for further analysis. It also gives the possibility of tracking the dynamics of changes occurring in biological traces applied on the surface. For the analyzed 28 blood samples, described method allowed to distinguish those blood stains because of time of apply.

The antisickling effects of dried fish (tilapia) And dried prawn ...

African Journals Online (AJOL)

The antisickling effect of dried fish (Tilapia) and dried prawn (Astacus red) were investigated to ascertain the ability of the extracts of these samples to inhibit polymerisation of sickle cell haemoglobin (HbS), improve the Fe 2+/Fe 3+ ratio and lower the activity of lactate dehydrogenase (LDH) in blood plasma. The samples ...

Freeze dried blood and development of an artificial diet for blood feeding arthropods

International Nuclear Information System (INIS)

DeLoach, J.R.; Spates, G.E.; Kapatsa, G.M.; Sheffield, C.L.; Kabayo, J.P.

1990-01-01

The goals of the research were to determine the biochemical differences between freeze dried bovine and porcine blood relative to their nutritional value to Glossina palpalis palpalis and Stomoxys calcitrans and to develop an artificial diet for mass rearing these flies. Freeze dried bovine and porcine blood were found to differ in their amino acid content; total dietary lipids did not significantly differ, but some notable exceptions were found in fatty acid content. Both sonication and addition of foetal bovine serum to freeze dried bovine blood improved its nutritional value for G. p. palpalis. A two component, semi-defined artificial diet was developed for G. p. palpalis and S. calcitrans. The College Station diet consisted of lipid contaminated bovine haemoglobin (BHb) and bovine serum albumin (BSA). To conduct dietary deletion tests, a process was developed for preparing large quantities of ultrapure lipid free bovine haemoglobin. S. calcitrans fed on lipid free BHb plus BSA had zero fecundity. Lipids were re-added to the protein diet in three forms: (1) lipid contaminated BHb, (2) pure erythrocyte ghosts, and (3) pure lipids. It was found that membrane lipid from the erythrocyte is required by S. calcitrans. A defined artificial diet consisting of lipid free BHb, BSA, sphingomyelin, phosphatidyl ethanolamine, phosphatidyl serine and cholesterol gave normal adult survival, as well as near normal fecundity and percentage egg hatch for S. calcitrans. Knowing the identity of the lipids, it is now possible to prepare dietary formulations to alleviate dependency on the blood proteins BHb and BSA. (author). 34 refs, 1 fig., 15 tabs

Use of a dry-plasma collection device to overcome problems with storage and transportation of blood samples for epidemiology studies in developing countries.

Science.gov (United States)

Nurgalieva, Z Z; Almuchambetova, R; Machmudova, A; Kapsultanova, D; Osato, M S; Peacock, J; Zoltek, R P; Marchildon, P A; Graham, D Y; Zhangabylov, A

2000-11-01

Studies are difficult in areas lacking modern facilities due to the inability to reliably collect, store, and ship samples. Thus, we sought to evaluate the use of a dry plasma collection device for seroepidemiology studies. Plasma was obtained by fingerstick using a commercial dry plasma collection device (Chemcard Plasma Collection Device) and serum (venipuncture) from individuals in Kazakhstan. Plasma samples were air dried for 15 min and then stored desiccated in foil zip-lock pouches at 4 to 6 degrees C and subsequently shipped to the United States by air at ambient temperature. Serum samples remained frozen at -20 degrees C until assayed. Helicobacter pylori status was determined by enzyme-linked immunosorbent assay (HM-CAP EIA) for the dry plasma and the serum samples. The results were concordant in 250 of the 289 cases (86.5%). In 25 cases (8.6%), the dry plasma samples gave indeterminate results and could not be retested because only one sample was collected. Five serum samples were positive, and the corresponding dry plasma samples were negative; one serum sample was negative, and the corresponding plasma sample was positive. The relative sensitivity and specificity of the Chemcard samples to serum were 97.6 and 97.9%, respectively, excluding those with indeterminate results. Repeated freeze-thawing had no adverse effect on the accuracy of the test. We found the dry plasma collection device to provide an accurate and practical alternative to serum when venipuncture may be difficult or inconvenient and sample storage and handling present difficulties, especially for seroepidemiologic studies in rural areas or developing countries and where freeze-thawing may be unavoidable.

TSH IRMA of dried blood spots

International Nuclear Information System (INIS)

Tojinda, N.; Pattanachak, C.; Chongchirasiri, S.; Pattanachak, S.; Putrasreni, N.; Pleehachinda, R.; Suwanik, R.

1990-01-01

TSH determination is most useful for screening of neonatal hypothyroid in the population in iodine deficient areas. The NETRIA IRMA method for serum TSH was applied for blood-spot TSH. Cord blood on SS No. 903 filter paper was left dry overnight. The spot of 6 mm diameter, one/tube, was mixed with an assay buffer, diluted labelled m-anti-TSH, and diluted anti-TSH-solid phase. The mixture was rotated for 22-24 hours. After washing twice with wash buffer, it was counted for 1 minute. The standard curve with 0, 5, 10, 25, 50, 100, and 150 mIU/L whole blood was obtained with the maximum binding of 25%. The precision profile was satisfactory with %CV of 0 C) or 4 0 C or -20 0 C. The correlation between serum and blood-spot TSH values (n=120) showed r of 0.9541 and y=1.6123 (BS-TSH) +1.382. The mean of normal cord blood spot TSH (n=142) was 5.27 mIU/L. The technique was found to be precise, sensitive and easy to perform. Mass screening with this developed method is underway

Energy dispersive X-ray fluorescence spectrometry for the direct multi-element analysis of dried blood spots

Science.gov (United States)

Marguí, E.; Queralt, I.; García-Ruiz, E.; García-González, E.; Rello, L.; Resano, M.

2018-01-01

Home-based collection protocols for clinical specimens are actively pursued as a means of improving life quality of patients. In this sense, dried blood spots (DBS) are proposed as a non-invasive and even self-administered alternative to sampling whole venous blood. This contribution explores the potential of energy dispersive X-ray fluorescence spectrometry for the simultaneous and direct determination of some major (S, Cl, K, Na), minor (P, Fe) and trace (Ca, Cu, Zn) elements in blood, after its deposition onto clinical filter papers, thus giving rise to DBS. For quantification purposes the best strategy was to use matrix-matched blood samples of known analyte concentrations. The accuracy and precision of the method were evaluated by analysis of a blood reference material (Seronorm™ trace elements whole blood L3). Quantitative results were obtained for the determination of P, S, Cl, K and Fe, and limits of detection for these elements were adequate, taking into account their typical concentrations in real blood samples. Determination of Na, Ca, Cu and Zn was hampered by the occurrence of high sample support (Na, Ca) and instrumental blanks (Cu, Zn). Therefore, the quantitative determination of these elements at the levels expected in blood samples was not feasible. The methodology developed was applied to the analysis of several blood samples and the results obtained were compared with those reported by standard techniques. Overall, the performance of the method developed is promising and it could be used to determine the aforementioned elements in blood samples in a simple, fast and economic way. Furthermore, its non-destructive nature enables further analyses by means of complementary techniques to be carried out.

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

Science.gov (United States)

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 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 concentrations. PMID:27015245

Dried-blood spots: a cost-effective field method for the detection of Chikungunya virus circulation in remote areas.

Directory of Open Access Journals (Sweden)

Soa Fy Andriamandimby

Full Text Available 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.

Fully automated drug screening of dried blood spots using online LC-MS/MS analysis

Directory of Open Access Journals (Sweden)

Stefan Gaugler

2018-01-01

Full Text Available A new and fully automated workflow for the cost effective drug screening of large populations based on the dried blood spot (DBS technology was introduced in this study. DBS were prepared by spotting 15 μL of whole blood, previously spiked with alprazolam, amphetamine, cocaine, codeine, diazepam, fentanyl, lysergic acid diethylamide (LSD, 3,4-methylenedioxymethamphet-amine (MDMA, methadone, methamphetamine, morphine and oxycodone onto filter paper cards. The dried spots were scanned, spiked with deuterated standards and directly extracted. The extract was transferred online to an analytical LC column and then to the electrospray ionization tandem mass spectrometry system. All drugs were quantified at their cut-off level and good precision and correlation within the calibration range was obtained. The method was finally applied to DBS samples from two patients with back pain and codeine and oxycodone could be identified and quantified accurately below the level of misuse of 89.6 ng/mL and 39.6 ng/mL respectively.

The relative nutritive value of irradiated spray-dried blood powder and heat-sterilized blood meal as measured in combination with whey protein

International Nuclear Information System (INIS)

Downes, T.E.H.; Nourse, L.D.; Siebrits, F.K.; Hastings, J.W.

1987-01-01

A method of processing blood meal in which nutritive value of the protein is preserved is described, since appreciable losses occur in the nutritive value of the protein when prepared by heat sterilization with drying at atmospheric pressure in steam jacketed vessels. Blood was spray dried and irradiated at an intensity of 10 kGy. Collectively the heat of spray drying and irradiation was effective in killing both the virus plaque-forming units and the bacteria, thus producing a commercially acceptable sterile product of higher nutritive value. The relative nutritive values (RNV) of 50:50 protein were 0,56 for whey protein concentrate plus heat-sterilized blood meal and 0.90 for whey protein concentrate plus irradiated spray-dried blood powder. Whey protein concentrate used as a control has a RNV of 1,0

Whole-genome amplified DNA from stored dried blood spots is reliable in high resolution melting curve and sequencing analysis

DEFF Research Database (Denmark)

Winkel, Bo G; Hollegaard, Mads V; Olesen, Morten S

2011-01-01

BACKGROUND: The use of dried blood spots (DBS) samples in genomic workup has been limited by the relative low amounts of genomic DNA (gDNA) they contain. It remains to be proven that whole genome amplified DNA (wgaDNA) from stored DBS samples, constitutes a reliable alternative to gDNA.We wanted...

Evaluation of HBsAg and anti-HBc assays in saliva and dried blood spot samples according HIV status.

Science.gov (United States)

Flores, Geane Lopes; Cruz, Helena Medina; Potsch, Denise Vigo; May, Silvia Beatriz; Brandão-Mello, Carlos Eduardo; Pires, Marcia Maria Amendola; Pilotto, Jose Henrique; Lewis-Ximenez, Lia Laura; Lampe, Elisabeth; Villar, Livia Melo

2017-09-01

Influence of HIV status in HBV markers detection in saliva and dried blood spots (DBS) was not well established. This study aims to evaluate the performance of optimized commercial immunoassay for identifying HBsAg and anti-HBc in saliva and DBS according HIV status. A sum of 535 individuals grouped as HIV + , HBV + , HIV/HBV + and HIV/HBV- were recruited where 347 and 188 were included for HBsAg and anti-HBc evaluation, respectively. Serum, DBS collected in Whatman 903 paper and saliva obtained using salivette device were analyzed using EIA. Increased sample volume and ROC curve analysis for cut off determination were used for DBS and saliva testing. HBsAg detection in saliva and DBS exhibited sensitivities of 80.9% and 85.6% and specificities of 86.8% and 96.3%. Sensitivity of anti-HBc in saliva and DBS were 82.4% and 76.9% and specificities in saliva and DBS were 96.9% and 91.7%. Low sensitivities were observed for HBsAg (62%) and anti-HBc (47%) detection in saliva of HIV/HBV+ individuals. OD values were also lower for HBsAg detection in DBS and saliva of HIV/HBV+ individuals compared to their serum samples. Statistical significance was found for sensitivities in HBsAg detection between saliva and DBS demonstrating high sensitivity for DBS specimens. In conclusion, HIV status or antiretroviral treatment appears to interfere in the performance of HBsAg and anti-HBc detection in DBS and saliva samples using the adapted commercial EIA. Copyright © 2017 Elsevier B.V. All rights reserved.

Methods for Determination of α-Glycosidase, β-Glycosidase, and α-Galactosidase Activities in Dried Blood Spot Samples.

Science.gov (United States)

Sozmen, Eser Yıldırım; Sezer, Ebru Demirel

2017-01-01

The lysosomal storage diseases (LDSs) are a heterogeneous group of inherited genetic disorders caused by defects of lysosomal proteins. The accumulation of undigested substrates from different catabolic pathways leads to cellular dysfunction. LSDs generally presents during early childhood and have a devastating impact on the families and on public health. Over the years, approaches for treatment of some LSDs have been developed with different strategies. Increasing availability of treatments of these diseases has accelerated the development of new methods and techniques for rapid diagnosis in patients with clinical indication.The use of dried blood spot (DBS) test has been proposed as a first tier test to identify patients with Gaucher, Pompe, and Fabry diseases. DBS usage is advantageous for the purpose of screening as it is non-invasive, sensitive, has low-cost and fast turnaround time compared to measurements in leucocyte and/or fibroblast culture. This chapter focuses on the activity measurement of three lysosomal enzymes (α-glucosidase, β-glucosidase, and α galactosidase) in DBS samples by using fluorescent substrates and by the LC-MS/MS (liquid chromatography-mass spectrometry) method. All steps of the methods, from preparation of the solutions to calculation of the enzyme activity, will be explained in detail.

Isolation of human genomic DNA for genetic analysis from premature neonates: a comparison between newborn dried blood spots, whole blood and umbilical cord tissue

Science.gov (United States)

2013-01-01

Background Genotyping requires biological sample collection that must be reliable, convenient and acceptable for patients and clinicians. Finding the most optimal procedure of sample collection for premature neonates who have a very limited blood volume is a particular challenge. The aim of the current study was to evaluate the use of umbilical cord (UC) tissue and newborn dried blood spot (DBS)-extracted genomic DNA (gDNA) as an alternative to venous blood-derived gDNA from premature neonates for molecular genetic analysis. All samples were obtained from premature newborn infants between 24-32 weeks of gestation. Paired blood and UC samples were collected from 31 study participants. gDNA was extracted from ethylenediaminetetraacetic acid (EDTA) anticoagulant-treated blood samples (~500 μl) and newborn DBSs (n = 723) using QIAamp DNA Micro kit (Qiagen Ltd., Crawley, UK); and from UC using Qiagen DNAeasy Blood and Tissue kit (Qiagen Ltd., Crawley, UK). gDNA was quantified and purity confirmed by measuring the A260:A280 ratio. PCR amplification and pyrosequencing was carried out to determine suitability of the gDNA for molecular genetic analysis. Minor allele frequency of two unrelated single nucleotide polymorphisms (SNPs) was calculated using the entire cohort. Results Both whole blood samples and UC tissue provided good quality and yield of gDNA, which was considerably less from newborn DBS. The gDNA purity was also reduced after 3 years of storage of the newborn DBS. PCR amplification of three unrelated genes resulted in clear products in all whole blood and UC samples and 86%-100% of newborn DBS. Genotyping using pyrosequencing showed 100% concordance in the paired UC and whole blood samples. Minor allele frequencies of the two SNPs indicated that no maternal gDNA contamination occurred in the genotyping of the UC samples. Conclusions gDNAs from all three sources are suitable for standard PCR and pyrosequencing assays. Given that UC provide good quality

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

Science.gov (United States)

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

Delay in blood sampling for routine newborn screening is associated with increased risk of schizophrenia

DEFF Research Database (Denmark)

Nordentoft, Merete; Tidselbak Larsen, Janne; Pedersen, Carsten Bøcker

2015-01-01

for this association. Therefore, we investigated whether the increased risk can be explained by other risk factors for schizophrenia. METHODS: A case-control design was applied. A total of 846 cases with schizophrenia were selected from the Danish Psychiatric Case Register. One control was selected for each case......BACKGROUND: The Danish Neonatal Screening Biobank, containing dried blood spot samples from all newborn in Denmark, is a unique source of data that can be utilized for analyses of genetic and environmental exposures related to schizophrenia and other mental disorders. In previous analyses, we have...... found that early and late blood sampling, compared to sampling at day 5, was associated with increased risk of schizophrenia. As delay in sampling of blood for neonatal screening cannot in itself influence the risk of schizophrenia, it must be seen as a proxy for unknown underlying causes responsible...

Retinol analysis in dried blood spots by HPLC.

Science.gov (United States)

Craft, N E; Haitema, T; Brindle, L K; Yamini, S; Humphrey, J H; West, K P

2000-04-01

There are many advantages to measuring vitamin A in dried blood spots (DBS) from a finger prick as compared to plasma collected by venipuncture. The advantages include easier collection, transport and storage; accessibility to younger and more remote populations; and decreased risk of disease transmission. We describe a method for the extraction of retinol from DBS for analysis by HPLC and initial comparison to plasma retinol. The effects of various buffers, detergents, antioxidants and chelators were evaluated to establish the most effective approach to elute the retinol: retinol binding protein (holo-RBP) complex from the blood collection cards. The process involves ultrasonic agitation to elute holo-RBP into a phosphate buffer containing an antioxidant and metal chelator. The holo-RBP complex was denatured by the addition of ethanol containing additional antioxidants permitting the extraction of free retinol into hexane. Following solvent evaporation, the extract was dissolved in methanol for HPLC analysis. The initial measured retinol levels in freshly collected DBS declined for 6-10 d whether stored at 25, 4 or -20 degrees C, but remained consistent thereafter (homeostatic). By incorporating a "recovery/volume adjustment" factor, measured retinol values in homeostatic DBS were adjusted to the equivalent of plasma retinol. For 17 normal adults, the correlation coefficient was 0.90 between plasma retinol and adjusted DBS retinol in samples that had been stored at -70 degrees C for < 9 mo. The use of this new sample matrix for vitamin A assessment will allow access to previously unavailable populations.

Silica Coated Paper Substrate for Paper-Spray Analysis of Therapeutic Drugs in Dried Blood Spots

Science.gov (United States)

Zhang, Zhiping; Xu, Wei; Manicke, Nicholas E.; Cooks, R. Graham; Ouyang, Zheng

2011-01-01

Paper spray is a newly developed ambient ionization method that has been applied for direct qualitative and quantitative analysis of biological samples. The properties of the paper substrate and spray solution have a significant impact on the release of chemical compounds from complex sample matrices, the diffusion of the analytes through the substrate, and the formation of ions for mass spectrometry analysis. In this study, a commercially available silica-coated paper was explored in an attempt to improve the analysis of therapeutic drugs in dried blood spots (DBS). The dichloromethane/isopropanol solvent has been identified as an optimal spray solvent for the analysis. The comparison was made with paper spray using chromatography paper as substrate with methanol/water as solvent for the analysis of verapamil, citalopram, amitriptyline, lidocaine and sunitinib in dried blood spots. It has been demonstrated the efficiency of recovery of the analytes was notably improved with the silica coated paper and the limit of quantitation (LOQ) for the drug analysis was 0.1 ng mL−1 using a commercial triple quadrupole mass spectrometer. The use of silica paper substrate also resulted in a sensitivity improvement of 5-50 fold in comparison with chromatography papers, including the Whatmann ET31 paper used for blood card. Analysis using a handheld miniature mass spectrometer Mini 11 gave LOQs of 10~20 ng mL−1 for the tested drugs, which is sufficient to cover the therapeutic ranges of these drugs. PMID:22145627

Quantification of sulfatides in dried blood and urine spots from metachromatic leukodystrophy patients by liquid chromatography/electrospray tandem mass spectrometry.

Science.gov (United States)

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

2014-06-10

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. 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. 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. Sulfatide accumulation studies in urine, but not in DBS, emerges as the method of choice if newborn screening is to be proposed for MLD. Copyright © 2013 Elsevier B.V. All rights reserved.

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

International Nuclear Information System (INIS)

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

1989-01-01

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

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

Directory of Open Access Journals (Sweden)

Luz-María Torres

Full Text Available 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.

Simplifying sampling for African swine fever surveillance: Assessment of antibody and pathogen detection from blood swabs.

Science.gov (United States)

Carlson, J; Zani, L; Schwaiger, T; Nurmoja, I; Viltrop, A; Vilem, A; Beer, M; Blome, S

2018-02-01

African swine fever (ASF) is a notifiable disease with serious socio-economic consequences that has been present in wild boar in the Baltic States and Poland since 2014. An introduction of ASF is usually accompanied by increased mortality, making fallen wild boar and hunted animals with signs of disease the main target for early warning and passive surveillance. It is difficult, however, to encourage hunters and foresters to report and take samples from these cases. A pragmatic and easy sampling approach with quick-drying swabs could facilitate this. In this study, we further evaluated the use of dry blood swabs for the detection of ASFV antibody and genome with samples from animal trials and diagnostic submissions (blood, bone and organs) from Estonia. Compared to serum samples, dried blood swabs yielded 93.1% (95% confidence interval: [83.3, 98.1]) sensitivity and 100% [95.9, 100.0] specificity in a commercial ASFV antibody ELISA. Similarly, the swabs gave a sensitivity of 98.9% [93.4, 100.0] and a specificity of 98.1% [90.1, 100.0] for genome detection by a standard ASFV p72 qPCR when compared to EDTA blood. The same swabs were tested in a VP72-antibody lateral flow device, with a sensitivity of 94.7% [85.4, 98.9] and specificity of 96.1% [89.0, 99.2] compared to the serum ELISA. When GenoTube samples tested in ELISA and LFD were compared, the sensitivity was 96.3% [87.3, 99.5] and the specificity was 93.8% [86.0, 97.9]. This study demonstrates reliable detection of ASFV antibody and genome from swabs. A field test of the swabs with decomposed wild boar carcasses in an endemic area in Estonia also gave promising results. Thus, this technique is a practical approach for surveillance of ASF in both free and endemic areas. © 2017 Blackwell Verlag GmbH.

Homocysteine measurement in dried blood spot for neonatal detection of homocystinurias.

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Alodaib, Ahmad N; Carpenter, Kevin; Wiley, Veronica; Wotton, Tiffany; Christodoulou, John; Wilcken, Bridget

2012-01-01

Expanded newborn screening (NBS) leads to an increased number of false positive results, causing parental anxiety, greater follow-up costs, and the need for further metabolic investigations. We developed and validated a second-tier approach for NBS of homocystinurias by measuring the total homocysteine (tHcy) on the initial dried blood spot (DBS) samples to reduce the need for further investigation, and investigated newborn DBS homocysteine values in patients with homocystinuria. Total DBS homocysteine was measured in normal newborns, and retrospectively in newborns with established disorders, using liquid chromatography tandem mass spectrometry (LC-MS/MS) with stable isotope-labelled internal standards for homocysteine. Analytes were separated using reverse phase chromatography with a total run time of 3 min. The method was linear over the range of 10-100 μmol/L of tHcy and showed excellent precision; intra-batch CV was 4% and inter-batch precision 6.5%. Comparison of 59 plasma values with DBS for tHcy taken at the same time showed excellent correlation, (r (2)>0.97). The reference range for current neonatal samples was 5.4-10.7 μmol/L (n=99), and for the stored neonatal samples (stored dry, sealed in plastic at room temperature for 10 years) was 1.7-5.5 μmol/L, (n=50), both being normally distributed. The clinical utility of this method was checked by retrospective analysis of stored NBS samples from patients with different forms of homocystinuria, including four different remethylating disorders. All had clear elevations of tHcy.

Patient identification in blood sampling.

Science.gov (United States)

Davidson, Anne; Bolton-Maggs, Paula

The majority of adverse reports relating to blood transfusions result from human error, including misidentification of patients and incorrect labelling of samples. This article outlines best practice in blood sampling for transfusion (but is recommended for all pathology samples) and the role of patient empowerment in improving safety.

Simultaneous measurement of 25 inflammatory markers and neurotrophins in neonatal dried blood spots by immunoassay with xMAP technology

DEFF Research Database (Denmark)

Skogstrand, Kristin; Thorsen, Poul; Nørgaard-Pedersen, Bent

2005-01-01

BACKGROUND: Inflammatory reactions and other events in early life may be part of the etiology of late-onset diseases, including cerebral palsy, autism, and type 1 diabetes. Most neonatal screening programs for congenital disorders are based on analysis of dried blood spot samples (DBSS), and stor...

A novel dried blood spot-LCMS method for the quantification of methotrexate polyglutamates as a potential marker for methotrexate use in children.

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Ahmed F Hawwa

Full Text Available Development and validation of a selective and sensitive LCMS method for the determination of methotrexate polyglutamates in dried blood spots (DBS.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.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.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.

A field trial of a PCR-based Mansonella ozzardi diagnosis assay detects high-levels of submicroscopic M. ozzardi infections in both venous blood samples and FTA card dried blood spots.

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Medeiros, Jansen Fernandes; Almeida, Tatiana Amaral Pires; Silva, Lucyane Bastos Tavares; Rubio, Jose Miguel; Crainey, James Lee; Pessoa, Felipe Arley Costa; Luz, Sergio Luiz Bessa

2015-05-20

Mansonella ozzardi is a poorly understood human filarial parasite with a broad distribution throughout Latin America. Most of what is known about its parasitism has come from epidemiological studies that have estimated parasite incidence using light microscopy. Light microscopy can, however, miss lighter, submicroscopic, infections. In this study we have compared M. ozzardi incidence estimates made using light microscopy, with estimates made using PCR. 214 DNA extracts made from Large Volume Venous Blood Samples (LVVBS) were taken from volunteers from two study sites in the Rio Solimões region: Codajás [n = 109] and Tefé [n = 105] and were subsequently assayed for M. ozzardi parasitism using a diagnostic PCR (Mo-dPCR). Peripheral finger-prick blood samples were taken from the same individuals and used for microscopic examination. Finger-prick blood, taken from individuals from Tefé, was also used for the creation of FTAcard dried blood spots (DBS) that were subsequently subjected to Mo-dPCR. Overall M. ozzardi incidence estimates made with LVVBS PCRs were 1.8 times higher than those made using microscopy (44.9% [96/214] compared with 24.3% [52/214]) and 1.5 times higher than the PCR estimates made from FTAcard DBS (48/105 versus 31/105). PCR-based detection of FTAcard DBS proved 1.3 times more sensitive at diagnosing infections from peripheral blood samples than light microscopy did: detecting 24/105 compared with 31/105. PCR of LVVBS reported the fewest number of false negatives, detecting: 44 of 52 (84.6%) individuals diagnosed by microscopy; 27 of 31 (87.1%) of those diagnosed positive from DBSs and 17 out of 18 (94.4%) of those diagnosed as positive by both alternative methodologies. In this study, Mo-dPCR of LVVBS was by far the most sensitive method of detecting M. ozzardi infections and detected submicroscopic infections. Mo-dPCR FTAcard DBS also provided a more sensitive test for M. ozzardi diagnosis than light microscopy based diagnosis did and

Use of serum and blood samples on filter paper to improve the surveillance of Dengue in Pacific Island Countries.

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Aubry, Maite; Roche, Claudine; Dupont-Rouzeyrol, Myrielle; Aaskov, John; Viallon, Jérôme; Marfel, Maria; Lalita, Paul; Elbourne-Duituturaga, Salanieta; Chanteau, Suzanne; Musso, Didier; Pavlin, Boris I; Harrison, Dustin; Kool, Jacob L; Cao-Lormeau, Van-Mai

2012-09-01

In Pacific Island Countries (PICs) the epidemiology of dengue is characterized by long-term transmission of a single dengue virus (DENV) serotype. The emergence of a new serotype in one island country often indicates major outbreaks with this serotype will follow in other PICs. Filter paper (FP) cards on which whole blood or serum from dengue suspected patients had been dried was evaluated as a method for transportation of this material by standard mail delivery throughout the Pacific. Twenty-two FP-dried whole blood samples collected from patients in New Caledonia and Wallis & Futuna Islands, during DENV-1 and DENV-4 transmission, and 76 FP-dried sera collected from patients in Yap State, Majuro (Republic of Marshall Islands), Tonga and Fiji, before and during outbreaks of DENV-2 in Yap State and DENV-4 in Majuro, were tested for the presence of DENV RNA, by serotype specific RT-PCR, at the Institut Louis Malardé in French Polynesia. The serotype of DENV could be determined, by a variety of RT-PCR procedures, in the FP-dried samples after more than three weeks of transport at ambient temperatures. In most cases, the sequencing of the envelope gene to genotype the viruses also was possible. The serotype and genotype of DENV can be determined from FP-dried serum or whole blood samples transported over thousands of kilometers at ambient, tropical, temperatures. This simple and low-cost approach to virus identification should be evaluated in isolated and resource poor settings for surveillance for a range of significant viral diseases. Copyright © 2012. Published by Elsevier B.V.

The effect of zeolite A supplementation in the dry period on blood mineral status around calving

DEFF Research Database (Denmark)

Thilsing-Hansen, T; Jørgensen, R J; Enemark, J M

2003-01-01

This article summarizes the results obtained in 6 separate studies concerned with the effect of zeolite A supplementation in the dry period on blood calcium, magnesium and phosphorus status around calving. The experiments were conducted on 5 different farms, and comprised a total of 117 cows. Two...... of the experiments (exp. 5 and 6) were conducted under extensive farming conditions whereas the rest (exp. 1-4) were conducted on intensively driven farms. All cows included in the experiments had completed at least 2 lactations. The cows were allocated as either untreated control cows or zeolite treated...... experimental cows according to expected date of calving and parity. The experimental cows were fed between 0.5 and 1.0 kg of zeolite A per day during the last 2 to 4 weeks of the dry period. Blood samples were drawn on the day of calving and day one and two after calving (all experiments), three weeks before...

A simplified method for determination of radioactive iron in whole-blood samples

DEFF Research Database (Denmark)

Bukhave, Klaus; Sørensen, Anne Dorthe; Hansen, M.

2001-01-01

in humans. The overall recovery of radioiron from blood is more than 90%, and the coefficient of variation, as judged by the variation in the ratio Fe-55/Fe-59 is in the order of 4%. Combined with whole-body counting of 59Fe and direct gamma -counting of Fe-59 on blood samples, this method represents......For studies on iron absorption in man radioisotopes represent an easy and simple tool. However, measurement of the orbital electron emitting radioiron, Fe-55, in blood is difficult and insufficiently described in the literature. The present study describes a relatively simple method...... for simultaneous determination of Fe-55 and Fe-59 in blood, using a dry-ashing procedure and recrystallization of the remaining iron. The detection Limit of the method permits measurements of 0.1 Bq/ml blood thus allowing detection of Less than 1% absorption from a 40 kBq dose, which is ethically acceptable...

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

Energy Technology Data Exchange (ETDEWEB)

Thomson, S.; Wallace, A.M.; Cook, B. (Stobhill Hospital, Glasgow (England))

1989-08-01

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

Alternative polymerase chain reaction method to identify Plasmodium species in human blood samples: the semi-nested multiplex malaria PCR (SnM-PCR)

NARCIS (Netherlands)

Rubio, J.M.; Post, R.J.; Docters van Leeuwen, W.M.; Henry, M.C.; Lindergard, G.; Hommel, M.

2002-01-01

A simplified protocol for the identification of Plasmodium species by semi-nested multiplex polymerase chain reaction (SnM-PCR) in human blood samples is compared with microscopical examination of thin and thick blood films in 2 field trials in Côte d'Ivoire and Cameroon. Also, dried blood spots or

HbA1c measurements from dried blood spots : validation and patient satisfaction

NARCIS (Netherlands)

Fokkema, Margaretha; Bakker, Andries J; de Boer, Fokje; Kooistra, Jeltsje; de Vries, Sifra; Wolthuis, Albert

2009-01-01

Background: This study evaluates HbA1c measurements from dried blood spots collected on filter paper and compares HbA1c from filter paper (capillary blood) with HbA1c measured in venous blood. Methods: Patient satisfaction was evaluated using a questionnaire. The performance with the filter paper

Evaluation of dried blood spot samples for screening of hepatitis C and human immunodeficiency virus in a real-world setting.

Science.gov (United States)

Vázquez-Morón, Sonia; Ryan, Pablo; Ardizone-Jiménez, Beatriz; Martín, Dolores; Troya, Jesus; Cuevas, Guillermo; Valencia, Jorge; Jimenez-Sousa, María A; Avellón, Ana; Resino, Salvador

2018-01-30

Both hepatitis C virus (HCV) infection and human immunodeficiency virus (HIV) infection are underdiagnosed, particularly in low-income countries and in difficult-to-access populations. Our aim was to develop and evaluate a methodology for the detection of HCV and HIV infection based on capillary dry blood spot (DBS) samples taken under real-world conditions. We carried out a cross-sectional study of 139 individuals (31 healthy controls, 68 HCV-monoinfected patients, and 40 HCV/HIV-coinfected patients). ELISA was used for anti-HCV and anti-HIV antibody detection; and SYBR Green RT-PCR was used for HCV-RNA detection. The HIV serological analysis revealed 100% sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). The HCV serological analysis revealed a sensitivity of 92.6%, specificity of 100%, PPV of 100%, and NPV of 79.5%. Finally, the HCV-RNA detection test revealed a detection limit of 5 copies/µl with an efficiency of 100% and sensitivity of 99.1%, specificity of 100%, PPV of 100%, and NPV of 96.9%. In conclusion, our methodology was able to detect both HCV infection and HIV infection from the same DBS sample with good diagnostic performance. Screening for HCV and HIV using DBS might be a key strategy in the implementation of national programs for the control of both infections.

A nested real-time PCR assay for the quantification of Plasmodium falciparum DNA extracted from dried blood spots.

Science.gov (United States)

Tran, Tuan M; Aghili, Amirali; Li, Shanping; Ongoiba, Aissata; Kayentao, Kassoum; Doumbo, Safiatou; Traore, Boubacar; Crompton, Peter D

2014-10-04

As public health efforts seek to eradicate malaria, there has been an emphasis on eliminating low-density parasite reservoirs in asymptomatic carriers. As such, diagnosing submicroscopic Plasmodium infections using PCR-based techniques has become important not only in clinical trials of malaria vaccines and therapeutics, but also in active malaria surveillance campaigns. However, PCR-based quantitative assays that rely on nucleic acid extracted from dried blood spots (DBS) have demonstrated lower sensitivity than assays that use cryopreserved whole blood as source material. The density of Plasmodium falciparum asexual parasites was quantified using genomic DNA extracted from dried blood spots (DBS) and the sensitivity of two approaches was compared: quantitative real-time PCR (qPCR) targeting the P. falciparum 18S ribosomal RNA gene, either with an initial conventional PCR amplification prior to qPCR (nested qPCR), or without an initial amplification (qPCR only). Parasite densities determined by nested qPCR, qPCR only, and light microscopy were compared. Nested qPCR results in 10-fold higher sensitivity (0.5 parasites/μl) when compared to qPCR only (five parasites/ul). Among microscopy-positive samples, parasite densities calculated by nested qPCR correlated strongly with microscopy for both asymptomatic (Pearson's r=0.58, PNested qPCR improves the sensitivity for the detection of P. falciparum blood-stage infection from clinical DBS samples. This approach may be useful for active malaria surveillance in areas where submicroscopic asymptomatic infections are prevalent.

Pooled HIV-1 viral load testing using dried blood spots to reduce the cost of monitoring antiretroviral treatment in a resource-limited setting.

Science.gov (United States)

Pannus, Pieter; Fajardo, Emmanuel; Metcalf, Carol; Coulborn, Rebecca M; Durán, Laura T; Bygrave, Helen; Ellman, Tom; Garone, Daniela; Murowa, Michael; Mwenda, Reuben; Reid, Tony; Preiser, Wolfgang

2013-10-01

Rollout of routine HIV-1 viral load monitoring is hampered by high costs and logistical difficulties associated with sample collection and transport. New strategies are needed to overcome these constraints. Dried blood spots from finger pricks have been shown to be more practical than the use of plasma specimens, and pooling strategies using plasma specimens have been demonstrated to be an efficient method to reduce costs. This study found that combination of finger-prick dried blood spots and a pooling strategy is a feasible and efficient option to reduce costs, while maintaining accuracy in the context of a district hospital in Malawi.

Detection of newly produced T and B lymphocytes by digital PCR in blood stored dry on nylon flocked swabs.

Science.gov (United States)

Tessitore, Marion Vaglio; Sottini, Alessandra; Roccaro, Aldo M; Ghidini, Claudia; Bernardi, Simona; Martellosio, Giovanni; Serana, Federico; Imberti, Luisa

2017-04-05

A normal number of T-cell receptor excision circles (TRECs) and K-deleting recombination excision circles (KRECs) is considered a biomarker for adequate new T- and B-cell production. In newborns, detection of TRECs and KRECs by real time PCR from dried blood spotted on filter paper is used for the screening of severe immunodeficiency. In adults, elderly and during diseases, where the number of TRECs is lower than in newborns and children, a large amount of DNA and a sensitive method of amplification are necessary to identify newly produced lymphocytes. DNA was prepared from blood of 203 healthy adults (range: 18-91 years old) absorbed for 10 s on flocked swabs and let to dry, or from peripheral blood mononuclear cells. DNA was subjected to digital PCR and to well established conventional real time PCR-based method using TREC- and KREC-specific primers and probes. The number of TRECs and KRECs was expressed per mL of blood. Statistical analysis was performed by nested ANOVA, Pearson coefficient of determination, and by linear regression tests. The novel method for the storage of dried blood on nylon flocked swabs and the use of digital PCR allow quantification of TRECs and KRECs with high degree of sensitivity, specificity, accuracy, and precision. TRECs and KRECs were amplified by digital PCR in all tested blood samples, including those obtained from elderly individuals (>70 years old) and that were negative by real time PCR. Furthermore, values of TRECs and KRECs obtained by digital PCR were in the range of those acquired by real time PCR. Our findings demonstrate that DNA isolation from dried blood on flocked swabs followed by digital PCR-based analysis represents a useful tool for studying new lymphocyte production in adults and elderly individuals. This suggests the potential use of the methodology when monitoring of clinical variables is limited by the number of molecules that can be amplified and detected, such as in patients with immunodeficiency or under

Radial line-scans as representative sampling strategy in dried-droplet laser ablation of liquid samples deposited on pre-cut filter paper disks

Energy Technology Data Exchange (ETDEWEB)

Nischkauer, Winfried [Institute of Chemical Technologies and Analytics, Vienna University of Technology, Vienna (Austria); Department of Analytical Chemistry, Ghent University, Ghent (Belgium); Vanhaecke, Frank [Department of Analytical Chemistry, Ghent University, Ghent (Belgium); Bernacchi, Sébastien; Herwig, Christoph [Institute of Chemical Engineering, Vienna University of Technology, Vienna (Austria); Limbeck, Andreas, E-mail: Andreas.Limbeck@tuwien.ac.at [Institute of Chemical Technologies and Analytics, Vienna University of Technology, Vienna (Austria)

2014-11-01

Nebulising liquid samples and using the aerosol thus obtained for further analysis is the standard method in many current analytical techniques, also with inductively coupled plasma (ICP)-based devices. With such a set-up, quantification via external calibration is usually straightforward for samples with aqueous or close-to-aqueous matrix composition. However, there is a variety of more complex samples. Such samples can be found in medical, biological, technological and industrial contexts and can range from body fluids, like blood or urine, to fuel additives or fermentation broths. Specialized nebulizer systems or careful digestion and dilution are required to tackle such demanding sample matrices. One alternative approach is to convert the liquid into a dried solid and to use laser ablation for sample introduction. Up to now, this approach required the application of internal standards or matrix-adjusted calibration due to matrix effects. In this contribution, we show a way to circumvent these matrix effects while using simple external calibration for quantification. The principle of representative sampling that we propose uses radial line-scans across the dried residue. This compensates for centro-symmetric inhomogeneities typically observed in dried spots. The effectiveness of the proposed sampling strategy is exemplified via the determination of phosphorus in biochemical fermentation media. However, the universal viability of the presented measurement protocol is postulated. Detection limits using laser ablation-ICP-optical emission spectrometry were in the order of 40 μg mL{sup −1} with a reproducibility of 10 % relative standard deviation (n = 4, concentration = 10 times the quantification limit). The reported sensitivity is fit-for-purpose in the biochemical context described here, but could be improved using ICP-mass spectrometry, if future analytical tasks would require it. Trueness of the proposed method was investigated by cross-validation with

Freeze-drying of mononuclear cells derived from umbilical cord blood followed by colony formation.

Directory of Open Access Journals (Sweden)

Dity Natan

Full Text Available 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 x 10(4+/-4.7, 3.49 x 10(4+/-6 and 6.31 x 10(4+/-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.

Blood drop patterns: Formation and applications.

Science.gov (United States)

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. Crown Copyright © 2016. Published by Elsevier B.V. All rights reserved.

Evaluation of PCR procedures for detecting and quantifying Leishmania donovani DNA in large numbers of dried human blood samples from a visceral leishmaniasis focus in Northern Ethiopia.

Science.gov (United States)

Abbasi, Ibrahim; Aramin, Samar; Hailu, Asrat; Shiferaw, Welelta; Kassahun, Aysheshm; Belay, Shewaye; Jaffe, Charles; Warburg, Alon

2013-03-27

Visceral Leishmaniasis (VL) is a disseminated protozoan infection caused by Leishmania donovani parasites which affects almost half a million persons annually. Most of these are from the Indian sub-continent, East Africa and Brazil. Our study was designed to elucidate the role of symptomatic and asymptomatic Leishmania donovani infected persons in the epidemiology of VL in Northern Ethiopia. The efficacy of quantitative real-time kinetoplast DNA/PCR (qRT-kDNA PCR) for detecting Leishmania donovani in dried-blood samples was assessed in volunteers living in an endemic focus. Of 4,757 samples, 680 (14.3%) were found positive for Leishmania k-DNA but most of those (69%) had less than 10 parasites/ml of blood. Samples were re-tested using identical protocols and only 59.3% of the samples with 10 parasite/ml or less were qRT-kDNA PCR positive the second time. Furthermore, 10.8% of the PCR negative samples were positive in the second test. Most samples with higher parasitemias remained positive upon re-examination (55/59 =93%). We also compared three different methods for DNA preparation. Phenol-chloroform was more efficient than sodium hydroxide or potassium acetate. DNA sequencing of ITS1 PCR products showed that 20/22 samples were Leishmania donovani while two had ITS1 sequences homologous to Leishmania major. Although qRT-kDNA PCR is a highly sensitive test, the dependability of low positives remains questionable. It is crucial to correlate between PCR parasitemia and infectivity to sand flies. While optimal sensitivity is achieved by targeting k-DNA, it is important to validate the causative species of VL by DNA sequencing.

A novel method for extracting nucleic acids from dried blood spots for ultrasensitive detection of low-density Plasmodium falciparum and Plasmodium vivax infections.

Science.gov (United States)

Zainabadi, Kayvan; Adams, Matthew; Han, Zay Yar; Lwin, Hnin Wai; Han, Kay Thwe; Ouattara, Amed; Thura, Si; Plowe, Christopher V; Nyunt, Myaing M

2017-09-18

Greater Mekong Subregion countries are committed to eliminating Plasmodium falciparum malaria by 2025. Current elimination interventions target infections at parasite densities that can be detected by standard microscopy or rapid diagnostic tests (RDTs). More sensitive detection methods have been developed to detect lower density "asymptomatic" infections that may represent an important transmission reservoir. These ultrasensitive polymerase chain reaction (usPCR) tests have been used to identify target populations for mass drug administration (MDA). To date, malaria usPCR tests have used either venous or capillary blood sampling, which entails complex sample collection, processing and shipping requirements. An ultrasensitive method performed on standard dried blood spots (DBS) would greatly facilitate the molecular surveillance studies needed for targeting elimination interventions. A highly sensitive method for detecting Plasmodium falciparum and P. vivax 18S ribosomal RNA from DBS was developed by empirically optimizing nucleic acid extraction conditions. The limit of detection (LoD) was determined using spiked DBS samples that were dried and stored under simulated field conditions. Further, to assess its utility for routine molecular surveillance, two cross-sectional surveys were performed in Myanmar during the wet and dry seasons. The lower LoD of the DBS-based ultrasensitive assay was 20 parasites/mL for DBS collected on Whatman 3MM filter paper and 23 parasites/mL for Whatman 903 Protein Saver cards-equivalent to 1 parasite per 50 µL DBS. This is about 5000-fold more sensitive than standard RDTs and similar to the LoD of ≤16-22 parasites/mL reported for other ultrasensitive methods based on whole blood. In two cross-sectional surveys in Myanmar, nearly identical prevalence estimates were obtained from contemporaneous DBS samples and capillary blood samples collected during the wet and dry season. The DBS-based ultrasensitive method described in this

Fetal scalp blood sampling during labor

DEFF Research Database (Denmark)

Chandraharan, Edwin; Wiberg, Nana

2014-01-01

Fetal cardiotocography is characterized by low specificity; therefore, in an attempt to ensure fetal well-being, fetal scalp blood sampling has been recommended by most obstetric societies in the case of a non-reassuring cardiotocography. The scientific agreement on the evidence for using fetal...... scalp blood sampling to decrease the rate of operative delivery for fetal distress is ambiguous. Based on the same studies, a Cochrane review states that fetal scalp blood sampling increases the rate of instrumental delivery while decreasing neonatal acidosis, whereas the National Institute of Health...... and Clinical Excellence guideline considers that fetal scalp blood sampling decreases instrumental delivery without differences in other outcome variables. The fetal scalp is supplied by vessels outside the skull below the level of the cranial vault, which is likely to be compressed during contractions...

Sampling for Beryllium Surface Contamination using Wet, Dry and Alcohol Wipe Sampling

Energy Technology Data Exchange (ETDEWEB)

Kerr, Kent [Central Missouri State Univ., Warrensburg, MO (United States)

2004-12-01

This research project was conducted at the National Nuclear Security Administration's Kansas City Plant, operated by Honeywell Federal Manufacturing and Technologies, in conjunction with the Safety Sciences Department of Central Missouri State University, to compare relative removal efficiencies of three wipe sampling techniques currently used at Department of Energy facilities. Efficiencies of removal of beryllium contamination from typical painted surfaces were tested by wipe sampling with dry Whatman 42 filter paper, with water-moistened (Ghost Wipe) materials, and by methanol-moistened wipes. Test plates were prepared using 100 mm X 15 mm Pyrex Petri dishes with interior surfaces spray painted with a bond coat primer. To achieve uniform deposition over the test plate surface, 10 ml aliquots of solution containing 1 beryllium and 0.1 ml of metal working fluid were transferred to the test plates and subsequently evaporated. Metal working fluid was added to simulate the slight oiliness common on surfaces in metal working shops where fugitive oil mist accumulates over time. Sixteen test plates for each wipe method (dry, water, and methanol) were processed and sampled using a modification of wiping patterns recommended by OSHA Method 125G. Laboratory and statistical analysis showed that methanol-moistened wipe sampling removed significantly more (about twice as much) beryllium/oil-film surface contamination as water-moistened wipes (p< 0.001), which removed significantly more (about twice as much) residue as dry wipes (p <0.001). Evidence for beryllium sensitization via skin exposure argues in favor of wipe sampling with wetting agents that provide enhanced residue removal efficiency.

Impact of blood sampling in very preterm infants

DEFF Research Database (Denmark)

Madsen, L P; Rasmussen, M K; Bjerregaard, L L

2000-01-01

; the groups were then subdivided into critically ill or not. Diagnostic blood sampling and blood transfusion events were recorded. In total, 1905 blood samples (5,253 analysis) were performed, corresponding to 0.7 samples (1.9 analysis) per day per infant. The highest frequencies were found during the first....../kg. For the extremely preterm infants a significant correlation between sampled and transfused blood volume was found (mean 37.1 and 33.3 ml/kg, respectively, r = + 0.71, p = 0.0003). The most frequently requested analyses were glucose, sodium and potassium. Few blood gas analyses were requested (1.9/ infant). No blood...... losses attributable to excessive generous sampling were detected. The results show an acceptable low frequency of sampling and transfusion events for infants of GA 28-32 weeks. The study emphasizes the necessity of thorough reflection and monitoring of blood losses when ordering blood sampling...

Heavy metals analysis in blood by the X-ray fluorescence method

International Nuclear Information System (INIS)

Perez Novara, Ana Ma.

1988-05-01

The analytical procedure for determination of heavy metals in blood is described. Blood was taken from active smelter workers, samples were stored in special flasks at 2 grades C. After freeze drying, dried samples was analyzed using X-ray fluorescence, sources excited: Pu-238 and Cd-109 was used. Br in Pb interferences were corrected. The agreement of the results with values in similar workers is satisfactory Median Pb level was 42.6 micro grams/100 ml (total blood), Cu was less 2.5 micro grams/g (dry blood) and Zn was 11 micro grams/g (dry blood). (author)

21 CFR 868.1100 - Arterial blood sampling kit.

Science.gov (United States)

2010-04-01

...) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Diagnostic Devices § 868.1100 Arterial blood sampling kit. (a) Identification. An arterial blood sampling kit is a device, in kit form, used to obtain arterial blood samples... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Arterial blood sampling kit. 868.1100 Section 868...

Blood sampling from adrenal gland vein

International Nuclear Information System (INIS)

Sun Yong; Ni Caifang

2009-01-01

Adrenal gland vein sampling is an interventional method to get the blood samples from the adrenal gland vein. The blood is obtained via a catheter which is selectively inserted in the adrenal gland vein. This technique is mainly used to be diagnostic for primary hyperaldosteronism. A full knowledge of the anatomy and variations of the adrenal gland vein, serious preoperative preparation and skilled catheterization manipulation are necessary for obtaining sufficient blood sample and for reducing the occurrence of complications. Providing the physicians with definite diagnostic evidence and being technically feasible, adrenal gland vein sampling should become one of the routine examinations for clarifying the cause of primary hyperaldosteronism. (authors)

The quantitative regional cerebral blood flow measurement with autoradiography method using 123I-IMP SPECT. Evaluation of arterialized venous blood sampling as a substitute for arterial blood sampling

International Nuclear Information System (INIS)

Ohnishi, Takashi; Yano, Takao; Nakano, Shinichi; Jinnouchi, Seishi; Nagamachi, Shigeki; Flores, L. II; Nakahara, Hiroshi; Watanabe, Katsushi.

1996-01-01

The purpose of this study is validation of calibrating a standard input function in autoradiography (ARG) method by one point venous blood sampling as a substitute for that by one point arterial blood sampling. Ten and 20 minutes after intravenous constant infusion of 123 I-IMP, arterialized venous blood sampling from a dorsal vein were performed on 15 patients having ischemic cerebrovascular disease. And arterial blood sampling from radial artery was performed 10 min after 123 I-IMP infusion. The mean difference rates of integrated input function between calibrated standard input function by arterial blood sampling at 10 min and that by venous blood sampling were 4.1±3% and 9.3±5.4% at 10 and 20 min after 123 I-IMP infusion, respectively. The ratio of venous blood radioactivity to arterial blood radioactivity at 10 min after 123 I-IMP infusion was 0.96±0.02. There was an excellent correlation between ARG method CBF values obtained by arterial blood sampling at 10 min and those obtained by arterialized venous blood sampling at 10 min. In conclusion, a substitution by arterialized venous blood sampling from dorsal hand vein for artery can be possible. The optimized time for arterialized venous blood sampling was 10 min after 123 I-IMP infusion. (author)

Dried blood spots as a source of anti-malarial antibodies for epidemiological studies

Science.gov (United States)

Corran, Patrick H; Cook, Jackie; Lynch, Caroline; Leendertse, Heleen; Manjurano, Alphaxard; Griffin, Jamie; Cox, Jonathan; Abeku, Tarekegn; Bousema, Teun; Ghani, Azra C; Drakeley, Chris; Riley, Eleanor

2008-01-01

Background Blood spots collected onto filter paper are an established and convenient source of antibodies for serological diagnosis and epidemiological surveys. Although recommendations for the storage and analysis of small molecule analytes in blood spots exist, there are no published systematic studies of the stability of antibodies under different storage conditions. Methods Blood spots, on filter paper or glass fibre mats and containing malaria-endemic plasma, were desiccated and stored at various temperatures for different times. Eluates of these spots were assayed for antibodies against two Plasmodium falciparum antigens, MSP-119 and MSP2, and calculated titres used to fit an exponential (first order kinetic) decay model. The first order rate constants (k) for each spot storage temperature were used to fit an Arrhenius equation, in order to estimate the thermal and temporal stability of antibodies in dried blood spots. The utility of blood spots for serological assays was confirmed by comparing antibodies eluted from blood spots with the equivalent plasma values in a series of samples from North Eastern Tanzania and by using blood spot-derived antibodies to estimate malaria transmission intensity in this site and for two localities in Uganda. Results Antibodies in spots on filter paper and glass fibre paper had similar stabilities but blood was more easily absorbed onto filter papers than glass fibre, spots were more regular and spot size was more closely correlated with blood volume for filter paper spots. Desiccated spots could be stored at or below 4°C for extended periods, but were stable for only very limited periods at ambient temperature. When desiccated, recoveries of antibodies that are predominantly of IgG1 or IgG3 subclasses were similar. Recoveries of antibodies from paired samples of serum and of blood spots from Tanzania which had been suitably stored showed similar recoveries of antibodies, but spots which had been stored for extended periods

Preanalytical Blood Sampling Errors in Clinical Settings

International Nuclear Information System (INIS)

Zehra, N.; Malik, A. H.; Arshad, Q.; Sarwar, S.; Aslam, S.

2016-01-01

Background: Blood sampling is one of the common procedures done in every ward for disease diagnosis and prognosis. Daily hundreds of samples are collected from different wards but lack of appropriate knowledge of blood sampling by paramedical staff and accidental errors make the samples inappropriate for testing. Thus the need to avoid these errors for better results still remains. We carried out this research with an aim to determine the common errors during blood sampling; find factors responsible and propose ways to reduce these errors. Methods: A cross sectional descriptive study was carried out at the Military and Combined Military Hospital Rawalpindi during February and March 2014. A Venous Blood Sampling questionnaire (VBSQ) was filled by the staff on voluntary basis in front of the researchers. The staff was briefed on the purpose of the survey before filling the questionnaire. Sample size was 228. Results were analysed using SPSS-21. Results: When asked in the questionnaire, around 61.6 percent of the paramedical staff stated that they cleaned the vein by moving the alcohol swab from inward to outwards while 20.8 percent of the staff reported that they felt the vein after disinfection. On contrary to WHO guidelines, 89.6 percent identified that they had a habit of placing blood in the test tube by holding it in the other hand, which should actually be done after inserting it into the stand. Although 86 percent thought that they had ample knowledge regarding the blood sampling process but they did not practice it properly. Conclusion: Pre analytical blood sampling errors are common in our setup. Eighty six percent participants though thought that they had adequate knowledge regarding blood sampling, but most of them were not adhering to standard protocols. There is a need of continued education and refresher courses. (author)

The direct measurement of free T4 in the eluate of dried blood. Usefulness as a confirmatory screening test for neonatal hypothyroidism

International Nuclear Information System (INIS)

Travert, G.; Lemonnier, F.; Fernandez, Y.

1984-01-01

In France neonatal screening for hypothyroidism is performed by mean of radioimmunoassay for TSH in dried blood spots, collected on the 5th day of life. In order to be able to get more data from the day 5 blood sample and thus confirm congenital hypothyroidism in all cases when elevated TSH levels have been found, we developed a simple radioimmunoassay for free T4 in eluates of dried blood spots on filter paper. We used reagents from the Amerlex FT4 RIA kit. The ready for use supplied anti-T4 antibody suspension and 125 I-T4 derivative solution are respectively 2-fold and 3-fold diluted in 0,154 M phosphate saline buffer-pH 7,4. Whole blood free T4 standards are prepared by adding serum standards in the kit to an equal volume of washed and packed red blood cells. Free T4 concentrations in these dried standards are expressed as picomoles per liter of whole blood. One blood spot 4.25 mm in diameter, equivalent to about 5-6 μl test blood or standard is incubated for 1 hour at room temperature in 500 μl of antibody suspension. The assay sensitivity (smallest detectable FT4 level) is 1.4 pmol.l -1 . Within assay and between assays coefficients of variation are respectively 9,5 and 13%. Free T4 levels measured in eluates of dried blood closely correlate (r=0,946) with plasma levels in the same subjects. Free T4 in 690 five day old neonates shows a non gaussian distribution, with a mean value: 13,6 pmol.l -1 and standard deviation: 4,3 (range 3,6-30 pmol.l -1 ). In 13 newborns with congenital hypothyroidism, day 5 free T4 levels were found within the range: 0 to 3,5 pmol.l -1 , and thus did not overlap with free T4 measured in age-matched euthyroid neonates [fr

DEVELOPMENT OF REAL-TIME MULTIPLEX PCR FOR THE QUANTITATIVE DETERMINATION OF TREC'S AND KREC'S IN WHOLE BLOOD AND IN DRIED BLOOD SPOTS

Directory of Open Access Journals (Sweden)

M. A. Gordukova

2015-01-01

Full Text Available Primary immunodeficiencies (PID such as severe combined immunodeficiency (SCID and X-linked agammaglobulinemia are characterized by the lack of functional Tand B-cells, respectively. Without early diagnosis and prompt treatment children with PID suffer from severe infectious diseases, leading to their death or disability. Our purpose was developing of simple, inexpensive, high throughput technique based on the quantitative determination of TREC and KREC molecules by real-time PCR, and its validation in a group of children with a verified diagnosis of SCID and X-linked agammaglobulinemia.In this study, we developed and validated multiplex real-time PCR for the TREC’s and KREC’s quantitative analysis. We have shown that linear range of Ct changes depending on the concentrations of targets with a correlation coefficient R2 not worse than 0.98 was observed at concentrations from 109 to 5 × 104 copies per ml. The lowest amount of targets reliably detected in a reaction volume was 10 TREC’s copies, 5 KREC ‘s copies and 5 copies of internal control (IL17RA. We determined the age-depended reference values of TRECs and KRECs in whole blood in 29 boys and 27 girls with normal immunological parameters. The normal cut-offs for TRECs and KRECs were defined in dry blood spots depending on the method of extraction.The proposed method showed 100% diagnostic sensitivity and specificity in the studied group. The method can be proposed as a screening tool for the diagnosis of SCID and X-linked agammaglobulinemia both in whole blood and in the dry blood spots. The further investigation is required with larger number of samples. 

Dried blood spot assay for the quantification of phenytoin using Liquid Chromatography-Mass Spectrometry.

Science.gov (United States)

Villanelli, Fabio; Giocaliere, Elisa; Malvagia, Sabrina; Rosati, Anna; Forni, Giulia; Funghini, Silvia; Shokry, Engy; Ombrone, Daniela; Della Bona, Maria Luisa; Guerrini, Renzo; la Marca, Giancarlo

2015-02-02

Phenytoin (PHT) is one of the most commonly used anticonvulsant drugs for the treatment of epilepsy and bipolar disorders. The large amount of plasma required by conventional methods for drug quantification makes mass spectrometry combined with dried blood spot (DBS) sampling crucial for pediatric patients where therapeutic drug monitoring or pharmacokinetic studies may be difficult to realize. DBS represents a new convenient sampling support requiring minimally invasive blood drawing and providing long-term stability of samples and less expensive shipment and storage. The aim of this study was to develop a LC-MS/MS method for the quantification of PHT on DBS. This analytical method was validated and gave good linearity (r(2)=0.999) in the range of 0-100mg/l. LOQ and LOD were 1.0mg/l and 0.3mg/l, respectively. The drug extraction from paper was performed in a few minutes using a mixture composed of organic solvent for 80%. The recovery ranged from 85 to 90%; PHT in DBS showed to be stable at different storage temperatures for one month. A good correlation was also obtained between PHT plasma and DBS concentrations. This method is both precise and accurate and appears to be particularly suitable to monitor treatment with a simple and convenient sample collection procedure. Copyright © 2014 Elsevier B.V. All rights reserved.

Fragile X protein in newborn dried blood spots.

Science.gov (United States)

Adayev, Tatyana; LaFauci, Giuseppe; Dobkin, Carl; Caggana, Michele; Wiley, Veronica; Field, Michael; Wotton, Tiffany; Kascsak, Richard; Nolin, Sarah L; Glicksman, Anne; Hosmer, Nicole; Brown, W Ted

2014-10-28

The fragile X syndrome (FXS) results from mutation of the FMR1 gene that prevents expression of its gene product, FMRP. We previously characterized 215 dried blood spots (DBS) representing different FMR1 genotypes and ages with a Luminex-based immunoassay (qFMRP). We found variable FMRP levels in the normal samples and identified affected males by the drastic reduction of FMRP. Here, to establish the variability of expression of FMRP in a larger random population we quantified FMRP in 2,000 anonymous fresh newborn DBS. We also evaluated the effect of long term storage on qFMRP by retrospectively assaying 74 aged newborn DBS that had been stored for 7-84 months that included normal and full mutation individuals. These analyses were performed on 3 mm DBS disks. To identify the alleles associated with the lowest FMRP levels in the fresh DBS, we analyzed the DNA in the samples that were more than two standard deviations below the mean. Analysis of the fresh newborn DBS revealed a broad distribution of FMRP with a mean approximately 7-fold higher than that we previously reported for fresh DBS in normal adults and no samples whose FMRP level indicated FXS. DNA analysis of the lowest FMRP DBS showed that this was the low extreme of the normal range and included a female carrying a 165 CGG repeat premutation. In the retrospective study of aged newborn DBS, the FMRP mean of the normal samples was less than 30% of the mean of the fresh DBS. Despite the degraded signal from these aged DBS, qFMRP identified the FXS individuals. The assay showed that newborn DBS contain high levels of FMRP that will allow identification of males and potentially females, affected by FXS. The assay is also an effective screening tool for aged DBS stored for up to four years.

Diagnostic accuracy of detection and quantification of HBV-DNA and HCV-RNA using dried blood spot (DBS) samples - a systematic review and meta-analysis.

Science.gov (United States)

Lange, Berit; Roberts, Teri; Cohn, Jennifer; Greenman, Jamie; Camp, Johannes; Ishizaki, Azumi; Messac, Luke; Tuaillon, Edouard; van de Perre, Philippe; Pichler, Christine; Denkinger, Claudia M; Easterbrook, Philippa

2017-11-01

The detection and quantification of hepatitis B (HBV) DNA and hepatitis C (HCV) RNA in whole blood collected on dried blood spots (DBS) may facilitate access to diagnosis and treatment of HBV and HCV infection in resource-poor settings. We evaluated the diagnostic performance of DBS compared to venous blood samples for detection and quantification of HBV-DNA and HCV-RNA in two systematic reviews and meta-analyses on the diagnostic accuracy of HBV DNA and HCV RNA from DBS compared to venous blood samples. We searched MEDLINE, Embase, Global Health, Web of Science, LILAC and Cochrane library for studies that assessed diagnostic accuracy with DBS. Heterogeneity was assessed and where appropriate pooled estimates of sensitivity and specificity were generated using bivariate analyses with maximum likelihood estimates and 95% confidence intervals. We also conducted a narrative review on the impact of varying storage conditions or different cut-offs for detection from studies that undertook this in a subset of samples. The QUADAS-2 tool was used to assess risk of bias. In the quantitative synthesis for diagnostic accuracy of HBV-DNA using DBS, 521 citations were identified, and 12 studies met the inclusion criteria. Overall quality of studies was rated as low. The pooled estimate of sensitivity and specificity for HBV-DNA was 95% (95% CI: 83-99) and 99% (95% CI: 53-100), respectively. In the two studies that reported on cut-offs and limit of detection (LoD) - one reported a sensitivity of 98% for a cut-off of ≥2000 IU/ml and another reported a LoD of 914 IU/ml using a commercial assay. Varying storage conditions for individual samples did not result in a significant variation of results. In the synthesis for diagnostic accuracy of HCV-RNA using DBS, 15 studies met the inclusion criteria, and this included six additional studies to a previously published review. The pooled sensitivity and specificity was 98% (95% CI:95-99) and 98% (95% CI:95-99.0), respectively

Radial line-scans as representative sampling strategy in dried-droplet laser ablation of liquid samples deposited on pre-cut filter paper disks.

Science.gov (United States)

Nischkauer, Winfried; Vanhaecke, Frank; Bernacchi, Sébastien; Herwig, Christoph; Limbeck, Andreas

2014-11-01

Nebulising liquid samples and using the aerosol thus obtained for further analysis is the standard method in many current analytical techniques, also with inductively coupled plasma (ICP)-based devices. With such a set-up, quantification via external calibration is usually straightforward for samples with aqueous or close-to-aqueous matrix composition. However, there is a variety of more complex samples. Such samples can be found in medical, biological, technological and industrial contexts and can range from body fluids, like blood or urine, to fuel additives or fermentation broths. Specialized nebulizer systems or careful digestion and dilution are required to tackle such demanding sample matrices. One alternative approach is to convert the liquid into a dried solid and to use laser ablation for sample introduction. Up to now, this approach required the application of internal standards or matrix-adjusted calibration due to matrix effects. In this contribution, we show a way to circumvent these matrix effects while using simple external calibration for quantification. The principle of representative sampling that we propose uses radial line-scans across the dried residue. This compensates for centro-symmetric inhomogeneities typically observed in dried spots. The effectiveness of the proposed sampling strategy is exemplified via the determination of phosphorus in biochemical fermentation media. However, the universal viability of the presented measurement protocol is postulated. Detection limits using laser ablation-ICP-optical emission spectrometry were in the order of 40 μg mL - 1 with a reproducibility of 10 % relative standard deviation (n = 4, concentration = 10 times the quantification limit). The reported sensitivity is fit-for-purpose in the biochemical context described here, but could be improved using ICP-mass spectrometry, if future analytical tasks would require it. Trueness of the proposed method was investigated by cross-validation with

Radial line-scans as representative sampling strategy in dried-droplet laser ablation of liquid samples deposited on pre-cut filter paper disks

Science.gov (United States)

Nischkauer, Winfried; Vanhaecke, Frank; Bernacchi, Sébastien; Herwig, Christoph; Limbeck, Andreas

2014-11-01

Nebulising liquid samples and using the aerosol thus obtained for further analysis is the standard method in many current analytical techniques, also with inductively coupled plasma (ICP)-based devices. With such a set-up, quantification via external calibration is usually straightforward for samples with aqueous or close-to-aqueous matrix composition. However, there is a variety of more complex samples. Such samples can be found in medical, biological, technological and industrial contexts and can range from body fluids, like blood or urine, to fuel additives or fermentation broths. Specialized nebulizer systems or careful digestion and dilution are required to tackle such demanding sample matrices. One alternative approach is to convert the liquid into a dried solid and to use laser ablation for sample introduction. Up to now, this approach required the application of internal standards or matrix-adjusted calibration due to matrix effects. In this contribution, we show a way to circumvent these matrix effects while using simple external calibration for quantification. The principle of representative sampling that we propose uses radial line-scans across the dried residue. This compensates for centro-symmetric inhomogeneities typically observed in dried spots. The effectiveness of the proposed sampling strategy is exemplified via the determination of phosphorus in biochemical fermentation media. However, the universal viability of the presented measurement protocol is postulated. Detection limits using laser ablation-ICP-optical emission spectrometry were in the order of 40 μg mL- 1 with a reproducibility of 10 % relative standard deviation (n = 4, concentration = 10 times the quantification limit). The reported sensitivity is fit-for-purpose in the biochemical context described here, but could be improved using ICP-mass spectrometry, if future analytical tasks would require it. Trueness of the proposed method was investigated by cross-validation with

Mass Spectrometry Method to Measure Membrane Proteins in Dried Blood Spots for the Detection of Blood Doping Practices in Sport.

Science.gov (United States)

Cox, Holly D; Eichner, Daniel

2017-09-19

The dried blood spot (DBS) matrix has significant utility for applications in the field where venous blood collection and timely shipment of labile blood samples is difficult. Unfortunately, protein measurement in DBS is hindered by high abundance proteins and matrix interference that increases with hematocrit. We developed a DBS method to enrich for membrane proteins and remove soluble proteins and matrix interference. Following a wash in a series of buffers, the membrane proteins are digested with trypsin and quantitated by parallel reaction monitoring mass spectrometry methods. The DBS method was applied to the quantification of four cell-specific cluster of differentiation (CD) proteins used to count cells by flow cytometry, band 3 (CD233), CD71, CD45, and CD41. We demonstrate that the DBS method counts low abundance cell types such as immature reticulocytes as well as high abundance cell types such as red blood cells, white blood cells, and platelets. When tested in 82 individuals, counts obtained by the DBS method demonstrated good agreement with flow cytometry and automated hematology analyzers. Importantly, the method allows longitudinal monitoring of CD protein concentration and calculation of interindividual variation which is difficult by other methods. Interindividual variation of band 3 and CD45 was low, 6 and 8%, respectively, while variation of CD41 and CD71 was higher, 18 and 78%, respectively. Longitudinal measurement of CD71 concentration in DBS over an 8-week period demonstrated intraindividual variation 17.1-38.7%. Thus, the method may allow stable longitudinal measurement of blood parameters currently monitored to detect blood doping practices.

A simple method for measurement of cerebral blood flow using 123I-IMP SPECT with calibrated standard input function by one point blood sampling. Validation of calibration by one point venous blood sampling as a substitute for arterial blood sampling

International Nuclear Information System (INIS)

Ito, Hiroshi; Akaizawa, Takashi; Goto, Ryoui

1994-01-01

In a simplified method for measurement of cerebral blood flow using one 123 I-IMP SPECT scan and one point arterial blood sampling (Autoradiography method), input function is obtained by calibrating a standard input function by one point arterial blood sampling. A purpose of this study is validation of calibration by one point venous blood sampling as a substitute for one point arterial blood sampling. After intravenous infusion of 123 I-IMP, frequent arterial and venous blood sampling were simultaneously performed on 12 patients of CNS disease without any heart and lung disease and 5 normal volunteers. The radioactivity ratio of venous whole blood which obtained from cutaneous cubital vein to arterial whole blood were 0.76±0.08, 0.80±0.05, 0.81±0.06, 0.83±0.11 at 10, 20, 30, 50 min after 123 I-IMP infusion, respectively. The venous blood radioactivities were always 20% lower than those of arterial blood radioactivity during 50 min. However, the ratio which obtained from cutaneous dorsal hand vein to artery were 0.93±0.02, 0.94±0.05, 0.98±0.04, 0.98±0.03, at 10, 20, 30, 50 min after 123 I-IMP infusion, respectively. The venous blood radioactivity was consistent with artery. These indicate that arterio-venous difference of radioactivity in a peripheral cutaneous vein like a dorsal hand vein is minimal due to arteriovenous shunt in palm. Therefore, a substitution by blood sampling from cutaneous dorsal hand vein for artery will be possible. Optimized time for venous blood sampling evaluated by error analysis was 20 min after 123 I-IMP infusion, which is 10 min later than that of arterial blood sampling. (author)

On-chip sample preparation for complete blood count from raw blood.

Science.gov (United States)

Nguyen, John; Wei, Yuan; Zheng, Yi; Wang, Chen; Sun, Yu

2015-03-21

This paper describes a monolithic microfluidic device capable of on-chip sample preparation for both RBC and WBC measurements from whole blood. For the first time, on-chip sample processing (e.g. dilution, lysis, and filtration) and downstream single cell measurement were fully integrated to enable sample preparation and single cell analysis from whole blood on a single device. The device consists of two parallel sub-systems that perform sample processing and electrical measurements for measuring RBC and WBC parameters. The system provides a modular environment capable of handling solutions of various viscosities by adjusting the length of channels and precisely controlling mixing ratios, and features a new 'offset' filter configuration for increased duration of device operation. RBC concentration, mean corpuscular volume (MCV), cell distribution width, WBC concentration and differential are determined by electrical impedance measurement. Experimental characterization of over 100,000 cells from 10 patient blood samples validated the system's capability for performing on-chip raw blood processing and measurement.

Outbreak of hepatitis E virus infection in Darfur, Sudan: effectiveness of real-time reverse transcription-PCR analysis of dried blood spots.

Science.gov (United States)

Mérens, Audrey; Guérin, Philippe Jean; Guthmann, Jean-Paul; Nicand, Elisabeth

2009-06-01

Biological samples collected in refugee camps during an outbreak of hepatitis E were used to compare the accuracy of hepatitis E virus RNA amplification by real-time reverse transcription-PCR (RT-PCR) for sera and dried blood spots (concordance of 90.6%). Biological profiles (RT-PCR and serology) of asymptomatic individuals were also analyzed.

Sample preservation for determination of organic compounds: microwave versus freeze-drying

International Nuclear Information System (INIS)

Popp, M.; Lied, W.; Meyer, A.J.; Richter, A.; Schiller, P.; Schwitte, H.

1996-01-01

In search of a reliable drying method, which might be used even under field conditions, microwave drying was compared to freeze-drying of plant material. Leaves of Ananas comosus and Avicennia germinans as well as buds and phloem of Acer pseudoplatanus were used and checked for one or more of the following substances: sugars, sugar alcohols, organic and amino acids, total nitrogen, and glycinebetaine. With most samples good agreement was achieved between the two drying methods. Only in the case of the Ananas comosus leaves, which exhibited low pH and high water content, did appreciable differences occur in organic and amino acids. Besides that, sucrose was the compound most susceptible to alterations, which was especially evident when leaves of Sambucus nigra were dried in the two different compartments (condenser compartment, drying bell jar) of the freeze-dryer in use. For Ananas comosus leaf samples it was shown that microwaving can also be used prior to extraction of tissue sap. (author)

Thyroglobulin is a more sensitive indicator of iodine deficiency than thyrotropin: development and evaluation of dry blood spot assays for thyrotropin and thyroglobulin in iodine-deficient geographical areas.

Science.gov (United States)

Missler, U; Gutekunst, R; Wood, W G

1994-03-01

Immunometric assays were developed for thyrotropin and thyroglobulin using time-resolved fluorescence as the measurement signal. The assays were suitable for measurements in serum/plasma or in dry blood spots (3 mm diameter). Both assays have acceptable coefficients of variation for dry blood spots (intra-assay median CV dry blood spot samples could be transported without special precautions for up to 5-6 weeks without significant loss in immunoreactivity. This agrees with other findings. The results showed that serum thyroglobulin levels are a more sensitive indicator of iodine deficiency than thyrotropin; elevated thyroglobulin levels were found in 182/304 children in Zimbabwe compared with elevated thyrotropin level in 28/304 cases. 213/304 children had enlarged thyroid glands. The cut-off levels used here were 4.5 mU/l thyrotropin and 20 micrograms/l for thyroglobulin, both in whole blood. The assays proved useful for assessing the efficacy of iodine therapy, either by oral dosage or intramuscularly (iodised oil).(ABSTRACT TRUNCATED AT 250 WORDS)

Light assisted drying (LAD) for protein stabilization: optical characterization of samples

Science.gov (United States)

Young, Madison A.; McKinnon, Madison E.; Elliott, Gloria D.; Trammell, Susan R.

2018-02-01

Light-Assisted Drying (LAD) is a novel biopreservation technique which allows proteins to be immobilized in a dry, amorphous solid at room temperature. Indicator proteins are used in a variety of diagnostic assays ranging from highthroughput 96-well plates to new microfluidic devices. A challenge in the development of protein-based assays is preserving the structure of the protein during production and storage of the assay, as the structure of the protein is responsible for its functional activity. Freeze-drying or freezing are currently the standard for the preservation of proteins, but these methods are expensive and can be challenging in some environments due to a lack of available infrastructure. An inexpensive, simple processing method that enables supra-zero temperature storage of proteins used in assays is needed. Light-assisted drying offers a relatively inexpensive method for drying samples. Proteins suspended in a trehalose solution are dehydrated using near-infrared laser light. The laser radiation speeds drying and as water is removed the sugar forms a protective matrix. The goal of this study is optically characterize samples processed with LAD. We use polarized light imaging (PLI) to look at crystallization kinetics of samples and determine optimal humidity. PLI shows a 62.5% chance of crystallization during LAD processing and negligible crystallization during low RH storage.

Effect of Sample Storage Temperature and Time Delay on Blood Gases, Bicarbonate and pH in Human Arterial Blood Samples.

Science.gov (United States)

Mohammadhoseini, Elham; Safavi, Enayat; Seifi, Sepideh; Seifirad, Soroush; Firoozbakhsh, Shahram; Peiman, Soheil

2015-03-01

Results of arterial blood gas analysis can be biased by pre-analytical factors, such as time interval before analysis, temperature during storage and syringe type. To investigate the effects of samples storage temperature and time delay on blood gases, bicarbonate and PH results in human arterial blood samples. 2.5 mL arterial blood samples were drawn from 45 patients via an indwelling Intraarterial catheter. Each sample was divided into five equal samples and stored in multipurpose tuberculin plastic syringes. Blood gas analysis was performed on one of five samples as soon as possible. Four other samples were divided into two groups stored at 22°C and 0°C. Blood gas analyses were repeated at 30 and 60 minutes after sampling. PaO2 of the samples stored at 0°C was increased significantly after 60 minutes (P = 0.007). The PaCO2 of the samples kept for 30 and 60 minutes at 22°C was significantly higher than primary result (P = 0.04, P samples stored at 22°C, pH decreased significantly after 30 and 60 minutes (P = 0.017, P = 0.001). There were no significant differences in other results of samples stored at 0°C or 22°C after 30 or 60 minutes. In samples stored in plastic syringes, overestimation of PaO2 levels should be noted if samples cooled before analysis. In samples stored in plastic syringes, it is not necessary to store samples in iced water when analysis delayed up to one hour.

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

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Wang Q

2016-03-01

Full Text Available Qingjun Wang,1,2,* Tao Sun,3,* Yunfeng Cao,1,2,4,5 Peng Gao,2,4,6 Jun Dong,2,4 Yanhua Fang,2 Zhongze Fang,2 Xiaoyu Sun,2 Zhitu Zhu1,2 1Oncology Department 2, The First Affiliated Hospital of Liaoning Medical University, 2Personalized Treatment and Diagnosis Research Center, The First Affiliated Hospital of Liaoning Medical University and Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Jinzhou, 3Department of Internal Medicine 1, Cancer Hospital of China Medical University, Liaoning Cancer Hospital & Insititute, Shenyang, 4CAS Key Laboratory of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 5Key Laboratory of Contraceptives and Devices Research (NPFPC, Shanghai Engineer and Technology Research Center of Reproductive Health Drug and Devices, Shanghai Institute of Planned Parenthood Research, Shanghai, 6Clinical Laboratory, Dalian Sixth People’s Hospital, Dalian, People’s Republic of China *These authors contributed equally to this work 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

Validation and Application of a Dried Blood Spot Ceftriaxone Assay

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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 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

Use of finger-prick dried blood spots (fpDBS) and capillary electrophoresis for carbohydrate deficient transferrin (CDT) screening in forensic toxicology.

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Bertaso, Anna; Sorio, Daniela; Vandoros, Anthula; De Palo, Elio F; Bortolotti, Federica; Tagliaro, Franco

2016-10-01

Continued progress in chronic alcohol abuse investigation requires the development of less invasive procedures for screening purposes. The application of finger-prick and related dried blood spots (fpDBS) for carbohydrate deficient transferrin (CDT) detection appears suitable for this aim. Therefore, the goal of this project was to develop a screening method for CDT using fpDBS with CZE analysis. Blood samples prepared by finger-prick were placed on DBS cards and left to air dry; each dried fpDBS disc was shredded into small pieces and suspended in acid solution (60 μL of HCl 120 mmol/L). After centrifugation (10 min at 1500 × g), the collected sample was adjusted to pH 3.5. After an overnight incubation, the pH was neutralised and an iron rich solution was added. After 1 h, CZE analysis was carried out. A group of 47 individuals was studied. Parallel serum samples were collected from each investigated subject and the %CDT for each sample was measured using HPLC and CZE techniques. The fpDBS transferrin sialo isoform electropherograms were similar to those obtained with serum. Moreover, fpDBS CZE CDT percentage levels demonstrated significant statistical correlation with those obtained from serum for both HPLC and CZE %CDT (p < 0.01; r 2 = 0.8913 and 0.8976, respectively), with %CDT from 0.8 to 13.7% for fpDBS and from 0.7 to 12.7% for serum. The newly developed fpDBS procedure for CDT analysis provides a simple and inexpensive tool for use in population screening. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evaluation of Sex-Specific Gene Expression in Archived Dried Blood Spots (DBS

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Scott Jewell

2012-08-01

Full Text Available Screening newborns for treatable serious conditions is mandated in all US states and many other countries. After screening, Guthrie cards with residual blood (whole spots or portions of spots are typically stored at ambient temperature in many facilities. The potential of archived dried blood spots (DBS for at-birth molecular studies in epidemiological and clinical research is substantial. However, it is also challenging as analytes from DBS may be degraded due to preparation and storage conditions. We previously reported an improved assay for obtaining global RNA gene expression from blood spots. Here, we evaluated sex-specific gene expression and its preservation in DBS using oligonucleotide microarray technology. We found X inactivation-specific transcript (XIST, lysine-specific demethylase 5D (KDM5D (also known as selected cDNA on Y, homolog of mouse (SMCY, uncharacterized LOC729444 (LOC729444, and testis-specific transcript, Y-linked 21 (TTTY21 to be differentially-expressed by sex of the newborn. Our finding that trait-specific RNA gene expression is preserved in unfrozen DBS, demonstrates the technical feasibility of performing molecular genetic profiling using such samples. With millions of DBS potentially available for research, we see new opportunities in using newborn molecular gene expression to better understand molecular pathogenesis of perinatal diseases.

Quantification of total hexose on dry blood spot by tandem mass spectrometry.

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Gong, Zhenhua; Tian, Guoli; Huang, Qiwei; Wang, Yanmin; Ge, Qingwei

2012-12-01

Because hypoglycemia and hyperglycemia are harmful and not always associated with overt clinical signs, it is necessary to have methods available to screen for glucose levels to detect hypoglycemia and diabetes as early as possible. A new method for such screening and the clinical determination of blood total hexose on a dry blood spot (DBS) using tandem mass spectrometry (MS/MS) was developed. The serum glucose controls and blood were prepared as DBS and then extracted into a methanol solution containing isotope-labeled internal standards. The methanolic extraction was subjected to HPLC, followed by MS/MS in positive ion mode. Multiple-reaction monitoring of m/z 203.1→23 was used to detect hexose, and m/z 209.0→23 was used for 13C6-D-glucose. The recoveries of blood glucose by MS/MS were 90%-102% with an R(2) value of 0.999 after linear regression (pblood total hexose in neonates aged 3-7 days (6.41±1.46 mmol/L) was lower than that in neonates aged 8-30 days (6.66±1.38 mmol/L), and it was lower in neonates than in children aged 1-72 months (7.19±1.87 mmol/L). Quantification of total hexose on a dry blood spot by MS/MS is accurate, reliable and feasible for screening and clinical tests. Copyright © 2012 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

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

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

2018-06-01

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

Delay in blood sampling for routine newborn screening is associated with increased risk of schizophrenia.

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Nordentoft, Merete; Larsen, Janne Tidselbak; Pedersen, Carsten Bøcker; Sørensen, Holger Jelling; Hollegaard, Mads Villiam; Hougaard, David Michael; Mortensen, Preben Bo; Petersen, Liselotte

2015-03-01

The Danish Neonatal Screening Biobank, containing dried blood spot samples from all newborn in Denmark, is a unique source of data that can be utilized for analyses of genetic and environmental exposures related to schizophrenia and other mental disorders. In previous analyses, we have found that early and late blood sampling, compared to sampling at day 5, was associated with increased risk of schizophrenia. As delay in sampling of blood for neonatal screening cannot in itself influence the risk of schizophrenia, it must be seen as a proxy for unknown underlying causes responsible for this association. Therefore, we investigated whether the increased risk can be explained by other risk factors for schizophrenia. A case-control design was applied. A total of 846 cases with schizophrenia were selected from the Danish Psychiatric Case Register. One control was selected for each case, matched on sex and exact date of birth. Both early and late blood sampling was associated with increased risk for schizophrenia. Compared to blood sampling at day 5, sampling at days 0 to 4 after birth was associated with an incidence rate ratio (IRR) of 1.46 (95% CI 1.15-1.87) for development of schizophrenia, and sampling at days 6 to 9 and at days 10 to 53 was associated with an IRR of 1.5 (95% CI 1.13-1.98) and 3.00 (95% CI 1.59-5.67), respectively. After adjusting the estimates for place of birth, both parents' psychiatric illness, maternal and paternal age, parents' country of origin, child admission, and parental education and income, the estimates were slightly different. Thus, blood collection at 0-4days was associated with an IRR of 1.27 (95% CI 0.94-1.71), 6-9days 1.31 (95% CI 0.94-1.84) and 10+days 3.52 (95% CI 1.50 to 8.24). After adjusting risk estimates for well-known risk factors, delay in sampling of blood for neonatal screening was associated with unexplained increased risk of schizophrenia. Thus, a key finding is that age at test is a proxy for unobserved risk factors

Supplementation of Dried Mealworm ( larva on Growth Performance, Nutrient Digestibility and Blood Profiles in Weaning Pigs

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X. H. Jin

2016-07-01

Full Text Available 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.

[Comparison of the determination of cyclosporin-A in blood samples collected on filter paper and by the ordinary technique].

Science.gov (United States)

Azevedo, L S; Manrique, R; Sabbaga, E

1995-01-01

Monitoring cyclosporin-A (CsA) blood levels is of utmost importance for the rational use of this drug. Although many centers perform transplants, in Brazil there are few laboratories able to measure CsA blood levels. Therefore making blood samples reach the laboratory emerged as a problem. Collection of blood on filter paper has been a technique used for a long time in special cases. PURPOSE--To confirm the usefulness of measuring CsA blood levels in blood samples collected on filter paper and in the usual way. METHOD--We studied twenty renal cadaver kidney recipients who were receiving CsA, azathioprine and prednisone. Ninety five blood samples were collected and divided into two aliquots. One of them was sent routinely to one laboratory to perform whole blood CsA measurements. From the other aliquot, 20 microliters were pipetted on filter paper. When dried they were mailed to the other laboratory, where, after elution, CsA was measured. In both cases radioimmunoassay with polyclonal antibody was used. RESULTS--Linear correlation between both measurements revealed r = 0.81 with no statistical difference. CONCLUSION--The technique showed to be useful in clinical practice. In countries with continental size, as Brazil, it may be very helpful.

Summary Report for Evaluation of Compost Sample Drying Methods

National Research Council Canada - National Science Library

Frye, Russell

1994-01-01

.... Previous work in Support of these efforts developed a compost sample preparation scheme, consisting of air drying followed by milling, to reduce analytical variability in the heterogeneous compost matrix...

Liquid chromatography-tandem mass spectrometry method for simultaneous quantification of bisoprolol, ramiprilat, propranolol and midazolam in rat dried blood spots.

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Cvan Trobec, Katja; Trontelj, Jurij; Springer, Jochen; Lainscak, Mitja; Kerec Kos, Mojca

2014-05-01

Dried blood spot (DBS) sampling represents a suitable method for pharmacokinetic studies in rats, particularly if serial sampling is needed. To study the pharmacokinetics of drugs in a rat heart failure (HF) model, we developed and validated a method for the simultaneous determination of bisoprolol, ramiprilat, propranolol and midazolam in DBS samples. Bisoprolol and ramipril are widely used in the treatment of HF, and midazolam and propranolol are markers of hepatic metabolism, which can be altered in HF. A 20μL sample of rat blood was pipetted onto Whatman 903 Protein Saver Card and allowed to dry. The whole spot was excised and 300μL of solvent (methanol with 10% ultrapure water and 0.1% formic acid) was added. After mixing and incubating the sample in an ultrasonic bath, a mixture of isotopically labeled internal standards was added. After centrifugation, the extracts were cleaned on an Ostro™ plate and analyzed using liquid chromatography-tandem mass spectroscopy. The method was successfully validated. No significant interference was observed in the retention times of analytes or internal standards. The intraday and interday accuracy and precision were within a ±15% interval. The method was linear in the range 5-250μg/L and the lower limit of quantification was 5μg/L for all four analytes. The absolute matrix effect ranged from 98.7% for midazolam to 121% for ramiprilat. The recovery was lowest for ramiprilat and highest for propranolol. Samples were stable at all tested temperatures. The method has been used successfully in a real-time pharmacokinetic study in rats. Copyright © 2014 Elsevier B.V. All rights reserved.

Technical Stability and Biological Variability in MicroRNAs from Dried Blood Spots: A Lung Cancer Therapy-Monitoring Showcase.

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Kahraman, Mustafa; Laufer, Thomas; Backes, Christina; Schrörs, Hannah; Fehlmann, Tobias; Ludwig, Nicole; Kohlhaas, Jochen; Meese, Eckart; Wehler, Thomas; Bals, Robert; Keller, Andreas

2017-09-01

Different work flows have been proposed to use miRNAs as blood-borne biomarkers. In particular, the method used for collecting blood from patients can considerably influence the diagnostic results. We explored whether dried blood spots (DBSs) facilitate stable miRNA measurements and compared its technical stability with biological variability. First, we tested the stability of DBS samples by generating from 1 person 18 whole-genome-wide miRNA profiles of DBS samples that were exposed to different temperature and humidity conditions. Second, we investigated technical reproducibility by performing 7 replicates of DBS again from 1 person. Third, we investigated DBS samples from 53 patients with lung cancer undergoing different therapies. Across these 3 stages, 108 genome-wide miRNA profiles from DBS were generated and evaluated biostatistically. In the stability analysis, we observed that temperature and humidity had an overall limited influence on the miRNomes (average correlation between the different conditions of 0.993). Usage of a silica gel slightly diminished DBS' technical reproducibility. The 7 technical replicates had an average correlation of 0.996. The correlation with whole-blood PAXGene miRNomes of the same individual was remarkable (correlation of 0.88). Finally, evaluation of the samples from the 53 patients with lung cancer exposed to different therapies showed that the biological variations exceeded the technical variability significantly ( P work flow for profiling of whole miRNomes on the basis of samples collected from DBS. Biological variations exceeded technical variations significantly. DBS-based miRNA profiles will potentially further the translational character of miRNA biomarker studies. © 2017 American Association for Clinical Chemistry.

Multidrug resistant Salmonellae isolated from blood culture samples ...

African Journals Online (AJOL)

This study investigates the prevalence of R-plasmids in Salmonella sp. isolated from blood samples of suspected typhoid patients in Warri, Nigeria. A total of 136 blood samples were collected between May and December,2009 and screened for the presence of Salmonellae using standard blood culture techniques of which ...

Development and validation of dried matrix spot sampling for the quantitative determination of amyloid β peptides in cerebrospinal fluid.

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Delaby, Constance; Gabelle, Audrey; Meynier, Philippe; Loubiere, Vincent; Vialaret, Jérôme; Tiers, Laurent; Ducos, Jacques; Hirtz, Christophe; Lehmann, Sylvain

2014-05-01

The use of dried blood spots on filter paper is well documented as an affordable and practical alternative to classical venous sampling for various clinical needs. This technique has indeed many advantages in terms of collection, biological safety, storage, and shipment. Amyloid β (Aβ) peptides are useful cerebrospinal fluid (CSF) biomarkers for Alzheimer disease diagnosis. However, Aβ determination is hindered by preanalytical difficulties in terms of sample collection and stability in tubes. We compared the quantification of Aβ peptides (1-40, 1-42, and 1-38) by simplex and multiplex ELISA, following either a standard operator method (liquid direct quantification) or after spotting CSF onto dried matrix paper card. The use of dried matrix spot (DMS) overcame preanalytical problems and allowed the determination of Aβ concentrations that were highly commutable (Bland-Altman) with those obtained using CSF in classical tubes. Moreover, we found a positive and significant correlation (r2=0.83, Pearson coefficient p=0.0329) between the two approaches. This new DMS method for CSF represents an interesting alternative that increases the quality and efficiency in preanalytics. This should enable the better exploitation of Aβ analytes for Alzheimer's diagnosis.

Use of Dried Blood Spots to Elucidate Full-Length Transmitted/Founder HIV-1 Genomes

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Jesus F. Salazar-Gonzalez

2016-07-01

Full Text Available Background: Identification of HIV-1 genomes responsible for establishing clinical infection in newly infected individuals is fundamental to prevention and pathogenesis research. Processing, storage, and transportation of the clinical samples required to perform these virologic assays in resource-limited settings requires challenging venipuncture and cold chain logistics. Here, we validate the use of dried-blood spots (DBS as a simple and convenient alternative to collecting and storing frozen plasma. Methods: We performed parallel nucleic acid extraction, single genome amplification (SGA, next generation sequencing (NGS, and phylogenetic analyses on plasma and DBS. Results: We demonstrated the capacity to extract viral RNA from DBS and perform SGA to infer the complete nucleotide sequence of the transmitted/founder (TF HIV-1 envelope gene and full-length genome in two acutely infected individuals. Using both SGA and NGS methodologies, we showed that sequences generated from DBS and plasma display comparable phylogenetic patterns in both acute and chronic infection. SGA was successful on samples with a range of plasma viremia, including samples as low as 1,700 copies/ml and an estimated ~50 viral copies per blood spot. Further, we demonstrated reproducible efficiency in gp160 env sequencing in DBS stored at ambient temperature for up to three weeks or at -20ºC for up to five months. Conclusions: These findings support the use of DBS as a practical and cost-effective alternative to frozen plasma for clinical trials and translational research conducted in resource-limited settings.

Blood oxygen saturation determined by transmission spectrophotometry of hemolyzed blood samples

Science.gov (United States)

Malik, W. M.

1967-01-01

Use of the Lambert-Beer Transmission Law determines blood oxygen saturation of hemolyzed blood samples. This simplified method is based on the difference in optical absorption properties of hemoglobin and oxyhemoglobin.

Non-terminal blood sampling techniques in guinea pigs.

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Birck, Malene M; Tveden-Nyborg, Pernille; Lindblad, Maiken M; Lykkesfeldt, Jens

2014-10-11

Guinea pigs possess several biological similarities to humans and are validated experimental animal models(1-3). However, the use of guinea pigs currently represents a relatively narrow area of research and descriptive data on specific methodology is correspondingly scarce. The anatomical features of guinea pigs are slightly different from other rodent models, hence modulation of sampling techniques to accommodate for species-specific differences, e.g., compared to mice and rats, are necessary to obtain sufficient and high quality samples. As both long and short term in vivo studies often require repeated blood sampling the choice of technique should be well considered in order to reduce stress and discomfort in the animals but also to ensure survival as well as compliance with requirements of sample size and accessibility. Venous blood samples can be obtained at a number of sites in guinea pigs e.g., the saphenous and jugular veins, each technique containing both advantages and disadvantages(4,5). Here, we present four different blood sampling techniques for either conscious or anaesthetized guinea pigs. The procedures are all non-terminal procedures provided that sample volumes and number of samples do not exceed guidelines for blood collection in laboratory animals(6). All the described methods have been thoroughly tested and applied for repeated in vivo blood sampling in studies within our research facility.

Sensitive screening of abused drugs in dried blood samples using ultra-high-performance liquid chromatography-ion booster-quadrupole time-of-flight mass spectrometry.

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Chepyala, Divyabharathi; Tsai, I-Lin; Liao, Hsiao-Wei; Chen, Guan-Yuan; Chao, Hsi-Chun; Kuo, Ching-Hua

2017-03-31

An increased rate of drug abuse is a major social problem worldwide. The dried blood spot (DBS) sampling technique offers many advantages over using urine or whole blood sampling techniques. This study developed a simple and efficient ultra-high-performance liquid chromatography-ion booster-quadrupole time-of-flight mass spectrometry (UHPLC-IB-QTOF-MS) method for the analysis of abused drugs and their metabolites using DBS. Fifty-seven compounds covering the most commonly abused drugs, including amphetamines, opioids, cocaine, benzodiazepines, barbiturates, and many other new and emerging abused drugs, were selected as the target analytes of this study. An 80% acetonitrile solvent with a 5-min extraction by Geno grinder was used for sample extraction. A Poroshell column was used to provide efficient separation, and under optimal conditions, the analytical times were 15 and 5min in positive and negative ionization modes, respectively. Ionization parameters of both electrospray ionization source and ion booster (IB) source containing an extra heated zone were optimized to achieve the best ionization efficiency of the investigated abused drugs. In spite of their structural diversity, most of the abused drugs showed an enhanced mass response with the high temperature ionization from an extra heated zone of IB source. Compared to electrospray ionization, the ion booster (IB) greatly improved the detection sensitivity for 86% of the analytes by 1.5-14-fold and allowed the developed method to detect trace amounts of compounds on the DBS cards. The validation results showed that the coefficients of variation of intra-day and inter-day precision in terms of the signal intensity were lower than 19.65%. The extraction recovery of all analytes was between 67.21 and 115.14%. The limits of detection of all analytes were between 0.2 and 35.7ngmL -1 . The stability study indicated that 7% of compounds showed poor stability (below 50%) on the DBS cards after 6 months of storage at

Common criteria among States for storage and use of dried blood spot specimens after newborn screening

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Carlo Petrini

2012-06-01

Full Text Available Biological samples collected in biobanks are a resource with significant research potential. The Italian Joint Group cNB - cNBBSV (National committee of Bioethics - National committee for Biosecurity, Biotechnologies and Life Sciences published a document reporting recommendations on storage and use of dried blood spot (DBS and on the development of a National Network of Regional Newborn Screening Repositories for collection of residual DBS. Several ethical questions (about consent, possible use of genetic information, unanticipated possible usages for research purposes rise from residual newborn screening specimens collections. Moreover, legal and ethical controversies are accentuated by the conflicts between the interests of sample donors, biobank holders, researchers and the public. To overcome these difficulties the identification of a few criteria for storage and research usage of DBS is crucial.

Laser cutting eliminates nucleic acid cross-contamination in dried-blood-spot processing.

Science.gov (United States)

Murphy, Sean C; Daza, Glenda; Chang, Ming; Coombs, Robert

2012-12-01

Dried blood spots (DBS) are useful for molecular assays but are prone to false positives from cross-contamination. In our malaria DBS assay, cross-contamination was encountered despite cleaning techniques suitable for HIV-1. We therefore developed a contact-free laser cutting system that effectively eliminated cross-contamination during DBS processing.

Low-cost HIV-1 diagnosis and quantification in dried blood spots by real time PCR.

Science.gov (United States)

Mehta, Nishaki; Trzmielina, Sonia; Nonyane, Bareng A S; Eliot, Melissa N; Lin, Rongheng; Foulkes, Andrea S; McNeal, Kristina; Ammann, Arthur; Eulalievyolo, Vindu; Sullivan, John L; Luzuriaga, Katherine; Somasundaran, Mohan

2009-06-05

Rapid and cost-effective methods for HIV-1 diagnosis and viral load monitoring would greatly enhance the clinical management of HIV-1 infected adults and children in limited-resource settings. Recent recommendations to treat perinatally infected infants within the first year of life are feasible only if early diagnosis is routinely available. Dried blood spots (DBS) on filter paper are an easy and convenient way to collect and transport blood samples. A rapid and cost effective method to diagnose and quantify HIV-1 from DBS is urgently needed to facilitate early diagnosis of HIV-1 infection and monitoring of antiretroviral therapy. We have developed a real-time LightCycler (rtLC) PCR assay to detect and quantify HIV-1 from DBS. HIV-1 RNA extracted from DBS was amplified in a one-step, single-tube system using primers specific for long-terminal repeat sequences that are conserved across all HIV-1 clades. SYBR Green dye was used to quantify PCR amplicons and HIV-1 RNA copy numbers were determined from a standard curve generated using serially diluted known copies of HIV-1 RNA. This assay detected samples across clades, has a dynamic range of 5 log(10), and %CV real-time systems demonstrated similar performance. The accuracy, reliability, genotype inclusivity and affordability, along with the small volumes of blood required for the assay suggest that the rtLC DBS assay will be useful for early diagnosis and monitoring of pediatric HIV-1 infection in resource-limited settings.

Measurement of cerebral blood flow the blood sampling method using 99mTc-ECD. Simultaneous scintigram scanning of arterial blood samples and the brain with a gamma camera

International Nuclear Information System (INIS)

Hachiya, Takenori; Inugami, Atsushi; Iida, Hidehiro; Mizuta, Yoshihiko; Kawakami, Takeshi; Inoue, Minoru

1999-01-01

To measure regional cerebral blood flow (rCBF) by blood sampling using 99m Tc-ECD we devised a method of measuring the radioactive concentration in arterial blood sample with a gamma camera. In this method the head and a blood sample are placed within the same visual field to record the SPECT data of both specimens simultaneously. The results of an evaluation of the counting rate performance, applying the 30 hours decaying method using 99m Tc solution showed that this method is not comparable to the well-type scintillation counter and in clinical cases the active concentration in arterial blood sample remained well within the dynamic range. In addition, examination of the influence of scattered radiation from the brain by the dilution method showed that it was negligible at a distance of more than 7.5 cm between the brain and the arterial blood sample. In the present study we placed a head-shaped phantom next to the sample. The results of the examinations suggested that this method is suitable for clinical application, and because it does not require a well-type scintillation counter, it is expected to find wide application. (author)

DETERMINATION OF REFERENCE VALUES FOR TREC AND KREC IN DRY BLOOD SPOTS OF NEWBORNS FROM DIFFERENT GESTATION AGES IN SVERDLOVSK REGION

Directory of Open Access Journals (Sweden)

S. S. Deryabina

2018-01-01

Full Text Available As a preparatory stage for implementation of genetic testing for severe combined immunodeficiency under a neonatal screening program, a study was performed in Sverdlovsk Region which concerned quantitative determination of T and B cell neogenesis markers (TREC and KREC, respectively in blood of conditionally healthy newborns. Archived samples of dry blood spots collected in test-forms for routine neonatal screening were used as biological material for the study of full-term 26 girls and 26 boys who did not exhibit serious illnesses during first year of their life. In addition, we investigated potential effects of foetal gestational age upon the number of TREC and KREC in preterm infants. Blood samples from 55 preterm infants (23 to 36 gestational weeks were also examined. It was shown that the levels of TREC and KREC increased sequentially with the increased gestation terms, but the quantitative changes of markers showed different dynamics. In this respect, the recommended terms of blood sample collection for SCID screening is entirely consistent with timing of blood sampling for routine newborn screening. An alternative result was obtained with a complete absence of TREC or KREC in blood sample of a newborn, irrespectively of prematurity degree (at valid copy numbers of a control gene which should serve as an indication for immediate consulting of the child by immunologist and in-depth immunological examination, because it may be a first prognostic sign of a fatal disease. In order to obtain correct cut-off levels for TREC/KREC, additional studies are needed on a larger sample of newborns (1.000 to 5.000, followed by validation of the obtained reference boundaries in studies involving patients with different forms of primary immunodeficiencies. 

Detection of IL28B SNP DNA from buccal epithelial cells, small amounts of serum, and dried blood spots.

Directory of Open Access Journals (Sweden)

Philippe Halfon

Full Text Available BACKGROUND & AIMS: Point mutations in the coding region of the interleukin 28 gene (rs12979860 have recently been identified for predicting the outcome of treatment of hepatitis C virus infection. This polymorphism detection was based on whole blood DNA extraction. Alternatively, DNA for genetic diagnosis has been derived from buccal epithelial cells (BEC, dried blood spots (DBS, and genomic DNA from serum. The aim of the study was to investigate the reliability and accuracy of alternative routes of testing for single nucleotide polymorphism allele rs12979860CC. METHODS: Blood, plasma, and sera samples from 200 patients were extracted (400 µL. Buccal smears were tested using an FTA card. To simulate postal delay, we tested the influence of storage at ambient temperature on the different sources of DNA at five time points (baseline, 48 h, 6 days, 9 days, and 12 days. RESULTS: There was 100% concordance between blood, plasma, sera, and BEC, validating the use of DNA extracted from BEC collected on cytology brushes for genetic testing. Genetic variations in HPTR1 gene were detected using smear technique in blood smear (3620 copies as well as in buccal smears (5870 copies. These results are similar to those for whole blood diluted at 1/10. A minimum of 0.04 µL, 4 µL, and 40 µL was necessary to obtain exploitable results respectively for whole blood, sera, and plasma. No significant variation between each time point was observed for the different sources of DNA. IL28B SNPs analysis at these different time points showed the same results using the four sources of DNA. CONCLUSION: We demonstrated that genomic DNA extraction from buccal cells, small amounts of serum, and dried blood spots is an alternative to DNA extracted from peripheral blood cells and is helpful in retrospective and prospective studies for multiple genetic markers, specifically in hard-to-reach individuals.

Platelet-rich fibrin prepared from stored whole-blood samples.

Science.gov (United States)

Isobe, Kazushige; Suzuki, Masashi; Watanabe, Taisuke; Kitamura, Yutaka; Suzuki, Taiji; Kawabata, Hideo; Nakamura, Masayuki; Okudera, Toshimitsu; Okudera, Hajime; Uematsu, Kohya; Nakata, Koh; Tanaka, Takaaki; Kawase, Tomoyuki

2017-12-01

In regenerative therapy, self-clotted platelet concentrates, such as platelet-rich fibrin (PRF), are generally prepared on-site and are immediately used for treatment. If blood samples or prepared clots can be preserved for several days, their clinical applicability will expand. Here, we prepared PRF from stored whole-blood samples and examined their characteristics. Blood samples were collected from non-smoking, healthy male donors (aged 27-67 years, N = 6), and PRF clots were prepared immediately or after storage for 1-2 days. Fibrin fiber was examined by scanning electron microscopy. Bioactivity was evaluated by means of a bioassay system involving human periosteal cells, whereas PDGF-BB concentrations were determined by an enzyme-linked immunosorbent assay. Addition of optimal amounts of a 10% CaCl 2 solution restored the coagulative ability of whole-blood samples that contained an anticoagulant (acid citrate dextrose) and were stored for up to 2 days at ambient temperature. In PRF clots prepared from the stored whole-blood samples, the thickness and cross-links of fibrin fibers were almost identical to those of freshly prepared PRF clots. PDGF-BB concentrations in the PRF extract were significantly lower in stored whole-blood samples than in fresh samples; however, both extracts had similar stimulatory effects on periosteal-cell proliferation. Quality of PRF clots prepared from stored whole-blood samples is not reduced significantly and can be ensured for use in regenerative therapy. Therefore, the proposed method enables a more flexible treatment schedule and choice of a more suitable platelet concentrate immediately before treatment, not after blood collection.

[Development of a laboratory test on dried blood spots for facilitating early diagnosis of alpha-1-antitrypsin deficiency].

Science.gov (United States)

Balduyck, Malika; Chapuis Cellier, Colette; Roche, Denis; Odou, Marie-Françoise; Joly, Philippe; Madelain, Vincent; Vergne, Anita; Nouadje, Georges; Lafitte, Jean-Jacques; Porchet, Nicole; Beaune, Philippe; Zerimech, Farid

2014-01-01

Alpha- 1-antitrypsin (A1AT) deficiency is a hereditary autosomal codominant genetic disorder resulting in low circulating levels of A1AT and leading to lung and/or liver disease. It remains underdiagnosed and only 5 to 10% of PIZZ patients, the most common form of severe A1AT deficiency, would be actually identified in France. Facilitating early diagnosis of A1AT deficiency would allow a better management of this disease; therefore we have developed and standardized in three laboratories involved in this study, a diagnostic test on dried blood spots (DBS) including quantitative A1AT measurement, phenotyping by IEF electrophoresis and, if necessary, genotyping by SERPINA1 gene sequencing. We performed a quantitative assay on 90 DBS samples by immunoturbidimetric or immunonephelometric methods. We demonstrated that both methods were suitable for this type of sampling and the results obtained were highly correlated (R(2)>0.9) between the three laboratories: for a target value of 1.00 g/L, the results obtained from the three laboratories were between 1.00 and 1.02 g/L. Phenotyping and genotyping were performed under redefined operating conditions and adapted to the analysis of DBS samples. The results were comparable with those obtained for venous blood samples. Following this work, it becomes possible to provide pulmonologists with a reliable kit to perform a capillary blood sampling on filter paper which would allow a large-scale screening of A1AT deficiency in the population particularly affected by this genetic condition.

On-line liquid chromatography/tandem mass spectrometry simultaneous determination of opiates, cocainics and amphetamines in dried blood spots.

Science.gov (United States)

Saussereau, E; Lacroix, C; Gaulier, J M; Goulle, J P

2012-02-15

A novel approach has been developed for the illicit drugs quantitative determination using dried blood spots (DBS) on filter paper. The illicit drugs tested were opiates (morphine and its 3- and 6-glucuronide metabolites, codeine, 6-monoacetylmorphine), cocainics (ecgonine methylester, benzoylecgonine, cocaine, cocaethylene) and amphetamines (amphetamine, methamphetamine, MDA, MDMA, MDEA). The described method, requiring a small blood volume, is based on high performance liquid chromatography coupled to tandem mass spectrometry using on-line extraction. A Whatman card 903 was spotted with 30μL of whole blood and left overnight to dry at room temperature. A 3-mm diameter disk was removed using a manual punch, suspended in 150μL of water for 10min with ultrasonication, and then 100μL was injected in the on-line LC-MS/MS system. An Oasis HLB was used as an extraction column and a C18 Atlantis as an analytical column. The chromatographic cycle was performed with 20mM ammonium formate buffer (pH 2.8) (solvent A) and acetonitrile/solvent A (90:10, v/v) gradient in 16min. Detection was performed in positive electrospray ionization mode (ESI+) with a Quattro Micro (Waters). Recoveries of all analytes were up to 80%. DBS were stored in duplicate at 4°C and -20°C for up to 6 months. Illicit drugs seemed to be much more stabled at -20°C. Furthermore, it was tested whether analysis of DBS may be as reliable as that of whole blood investigating authentic samples; significant correlations were obtained. This DBS assay has potential as rapid, sensitive and inexpensive option for the illicit drugs determination in small blood volumes, which seems of great interest in suspected cases of driving under the influence of drugs. Copyright © 2011 Elsevier B.V. All rights reserved.

Determination of blood cell subtype concentrations from frozen whole blood samples using TruCount beads.

Science.gov (United States)

Langenskiöld, Cecilia; Mellgren, Karin; Abrahamsson, Jonas; Bemark, Mats

2016-06-24

In many studies it would be advantageous if blood samples could be collected and analyzed using flow cytometry at a later stage. Ideally, sample collection should involve little hands-on time, allow for long-term storage, and minimally influence the samples. Here we establish a flow cytometry antibody panel that can be used to determine granulocytes, monocytes, and lymphocyte subset concentrations in fresh and frozen whole blood using TruCount technology. The panel can be used on fresh whole-blood samples as well as whole-blood samples that have been frozen after mixing with 10% DMSO. Concentrations in frozen and fresh sample is highly correlated both when frozen within 4 h and the day after collection (r ≥ 0.98), and the estimated concentration in frozen samples was between 91 and 94% of that in fresh samples for all cell types. Using this method whole-blood samples can be frozen using a simple preparation method, and stored long-term before accurate determination of cell concentration. This allows for standardized analysis of the samples at a reference laboratory in multi-center studies. © 2016 International Clinical Cytometry Society. © 2016 International Clinical Cytometry Society.

Comparison of drying characteristic and uniformity of banana cubes dried by pulse-spouted microwave vacuum drying, freeze drying and microwave freeze drying.

Science.gov (United States)

Jiang, Hao; Zhang, Min; Mujumdar, Arun S; Lim, Rui-Xin

2014-07-01

To overcome the flaws of high energy consumption of freeze drying (FD) and the non-uniform drying of microwave freeze drying (MFD), pulse-spouted microwave vacuum drying (PSMVD) was developed. The results showed that the drying time can be dramatically shortened if microwave was used as the heating source. In this experiment, both MFD and PSMVD could shorten drying time by 50% as compared to the FD process. Depending on the heating method, MFD and PSMVD dried banana cubes showed trends of expansion while FD dried samples demonstrated trends of shrinkage. Shrinkage also brought intensive structure and highest fracturability of all three samples dried by different methods. The residual ascorbic acid content of PSMVD dried samples can be as high as in FD dried samples, which were superior to MFD dried samples. The tests confirmed that PSMVD could bring about better drying uniformity than MFD. Besides, compared with traditional MFD, PSMVD can provide better extrinsic feature, and can bring about improved nutritional features because of the higher residual ascorbic acid content. © 2013 Society of Chemical Industry.

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

Science.gov (United States)

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 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; pmercury levels were associated with dry eye symptoms in a nationally representative Korean population. PMID:27121705

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

Science.gov (United States)

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

Neonatal blood gas sampling methods | Goenka | South African ...

African Journals Online (AJOL)

There is little published guidance that systematically evaluates the different methods of neonatal blood gas sampling, where each method has its individual benefits and risks. This review critically surveys the available evidence to generate a comparison between arterial and capillary blood gas sampling, focusing on their ...

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

Science.gov (United States)

Chung, So-Hyang; Myong, Jun-Pyo

2016-04-27

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. 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 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). 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; pdry eye symptoms in a nationally representative Korean population. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

Combination syringe provides air-free blood samples

Science.gov (United States)

Pool, S. L.

1970-01-01

Standard syringe and spinal needle are combined in unique manner to secure air-free blood samples. Combination syringe obtains air free samples because air bubbles become insignificant when samples greater than 1 cc are drawn.

Stability of carboxyhemoglobin in stored and mailed blood samples.

Science.gov (United States)

Hampson, Neil B

2008-02-01

Elevated blood carboxyhemoglobin (COHb) levels are used to confirm a clinical diagnosis of exposure to carbon monoxide (CO) and, in some instances, assess severity of poisoning. However, many hospital laboratories cannot measure COHb because they do not have CO-oximeters. In such instances, blood samples are often sent to outside laboratories or with a transported patient for measurement at the receiving hospital. This study was conducted to assess the stability of COHb in stored and mailed blood samples anticoagulated with heparin. Adult human blood was drawn into standard sample tubes anticoagulated with sodium heparin. Carbon monoxide gas was infused to raise the COHb level to 25% to 35%. Samples were then refrigerated or stored at room temperature, and serial COHb determinations were performed for 28 days. Additional samples were measured after being mailed locally or across the United States and back. No significant changes in COHb levels were seen in samples stored either in refrigeration or at room temperature over a period of 28 days or in samples shipped without refrigeration locally or across the United States. Carboxyhemoglobin levels in whole blood samples anticoagulated with heparin are stable with or without refrigeration for up to 4 weeks. If COHb measurement capability is not available, such samples may be shipped or transported with patients with confidence that the COHb level will be stable when measured at a later time.

Blood donors and factors impacting the blood donation decision: motives for donating blood in Turkish sample.

Science.gov (United States)

Karacan, Eda; Cengiz Seval, Guldane; Aktan, Zeynep; Ayli, Meltem; Palabiyikoglu, Refia

2013-12-01

Donations in Turkey are insufficient to cover the high transfusion needs arising from large numbers of thalassemia and sickle cell anemia patients and increasing demands for blood due to advanced surgery and cancer treatment. The most acceptable means to get blood is voluntary blood donation and the blood donor system in Turkey mostly depends on a combination of voluntary and involuntary donors. The main aim of this study is to explore the motivations of Turkish voluntary blood donors toward blood donation and to determine predictors of blood donation motivation. A cross-sectional sample survey of active blood donors in Ankara, Turkey was conducted. The sample consisted of 189 male volunteer blood donor adults. Donors filled in a self-administered questionnaire including the measures of demographic information, empathetic concern, altruism, social responsibility and blood donation motivation questionnaire during donation. Factor analysis of Blood Donation Motivation Measure with varimax rotation revealed a three-factor solution named as "values and moral duty", "positive feelings and esteem" and "self-benefit and external reasons". The results with regression analyses showed that only social responsibility had an significant effect independent of age, income, and education on blood donation motivation. These result reflects that blood donation motivation not only linked to a high degree of altruistic reasons, but also to a combination of some self-regarding motives. Additionally, feelings of empathy or altruism may be less strong at the time the decision to help, other factors may have a larger influence on helping decisions. Copyright © 2013 Elsevier Ltd. All rights reserved.

Congener Production in Blood Samples During Preparation and Storage

DEFF Research Database (Denmark)

Felby, Søren; Nielsen, Erik

1995-01-01

Retsmedicin, congener production, preparation, head space GC, acetone, isobutanol, storage, blood samples, n-propanol, methanol, methylethylketone......Retsmedicin, congener production, preparation, head space GC, acetone, isobutanol, storage, blood samples, n-propanol, methanol, methylethylketone...

High plasma corticosterone levels persist during frequent automatic blood sampling in rats

DEFF Research Database (Denmark)

Abelson, Klas S P; Adem, Bashir; Royo, Felix

2005-01-01

Corticosterone levels in blood may be used as a marker of stress in rodents, provided that the blood sampling procedure itself is non-stressful. Automated blood sampling equipment (Accusampler) allows blood sampling without any interference with the animal and might be useful as a tool for an on......-line measurement of stress markers in blood. However, the impact of the blood sampling itself on the corticosterone levels in blood is unknown. The present study was designed to evaluate whether the frequency of blood sampling influences the plasma corticosterone levels in male and female rats. During anaesthesia...... the importance of considering the frequency of blood withdrawal during automated blood sampling. This parameter may have an impact on the experimental results when using blood corticosterone levels as a stress marker, but also during any in vivo study where blood is collected, since high corticosterone levels...

Soil sample moisture content as a function of time during oven drying for gamma-ray spectroscopic measurements

International Nuclear Information System (INIS)

Benke, R.R.; Kearfott, K.J.

1999-01-01

In routine gamma-ray spectroscopic analysis of collected soil samples, procedure often calls to remove soil moisture by oven drying overnight at a temperature of 100 deg. C . Oven drying not only minimizes the gamma-ray self-attenuation of soil samples due to the absence of water during the gamma-ray spectroscopic analysis, but also allows for a straightforward calculation of the specific activity of radionuclides in soil, historically based on the sample dry weight. Because radon exhalation is strongly dependent on moisture , knowledge of the oven-drying time dependence of the soil moisture content, combined with radon exhalation measurements during oven drying and at room temperature for varying soil moisture contents, would allow conclusions to be made on how the oven-drying radon exhalation rate depends on soil moisture content. Determinations of the oven-drying radon exhalation from soil samples allow corrections to be made for the immediate laboratory gamma-ray spectroscopy of radionuclides in the natural uranium decay chain. This paper presents the results of soil moisture content measurements during oven drying and suggests useful empirical fits to the moisture data

COMPARISON OF WHOLE BLOOD AND PLASMA GLUCOSE CONCENTRATIONS IN GREEN TURTLES ( CHELONIA MYDAS) DETERMINED USING A GLUCOMETER AND A DRY CHEMISTRY ANALYZER.

Science.gov (United States)

Perrault, Justin R; Bresette, Michael J; Mott, Cody R; Stacy, Nicole I

2018-01-01

:  We compared glucose concentrations in whole blood and plasma from green turtles ( Chelonia mydas) using a glucometer with plasma glucose analyzed by dry chemistry analyzer. Whole blood glucose (glucometer) and plasma glucose (dry chemistry) had the best agreement ( r s =0.85) and a small negative bias (-0.08 mmol/L).

High-resolution melting (HRM) analysis as a feasible method for detecting spinal muscular atrophy via dried blood spots.

Science.gov (United States)

Er, Tze-Kiong; Kan, Tzu-Min; Su, Yu-Fa; Liu, Ta-Chih; Chang, Jan-Gowth; Hung, Shih-Ya; Jong, Yuh-Jyh

2012-11-12

Spinal muscular atrophy (SMA) is a neurodegenerative disease with the leading genetic cause of infant mortality. More than 95% of patients with SMA have a homozygous disruption in the survival motor neuron1 (SMN1) gene, caused by mutation, deletion, or rearrangement. Recently, treatment in humans in the immediate postnatal period, prior to the development of weakness or very early in the course of the disease, may be effective. Therefore, our objective was to establish a feasible method for SMA screening. High-resolution melting (HRM) analysis is rapidly becoming the most important mutation-scanning methodology that allows mutation scanning and genotyping without the need for costly labeled oligonucleotides. In the current study, we aim to develop a method for identifying the substitution of single nucleotide in SMN1 exon 7 (c.840C>T) by HRM analysis. Genomic DNA was extracted from peripheral blood samples and dried blood spots obtained from 30 patients with SMA and 30 normal individuals. All results were previously confirmed by denaturing high-performance liquid chromatography (DHPLC). In order to identify the substitution of single nucleotide in SMN1 exon 7 (c.840C>T) by HRM analysis, a primer set was used in HRM analysis. At first, we failed to identify the substitution of single nucleotide in SMN1 exon 7 (c.840C>T) by HRM analysis because the homozygous CC and homozygous TT cannot be distinguished by HRM analysis. Therefore, all samples were mixed with a known SMN1/SMN2 copy number (SMN1/SMN2=0:3), which we may call driver. This strategy is used to differentiate between homozygous CC and homozygous TT. After mixing with driver, the melting profile of homozygous CC becomes heteroduplex; however, the homozygous TT remains the same in the normalized and temperature-shifted difference plots. HRM analysis can be successfully applied to screen SMA via DNA obtained from whole blood and dried blood spots. We strongly believe that HRM analysis, a high-throughput method

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

Science.gov (United States)

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.

Clinical effect observation of pranoprofen combined with deproteinized calf blood extract eye drops for moderate to severe dry eye

Directory of Open Access Journals (Sweden)

Jing-Hua Qiu

2018-04-01

Full Text Available AIM: To explore clinical effect of pranoprofen combined with deproteinized calf blood extract eye drops for moderate to severe dry eye. METHODS: A total of 84 patients(132 eyeswho received treatment at the Zhengzhou Second Hospital were selected from January 2016 to January 2017. According to random number table method they were divided into control group 42 cases(68 eyesand observation group 42 cases(64 eyes, the control group using polyvinyl alcohol eye drops with pranoprofen, observation group with pranoprofen with deproteinized extract of calf blood eye drops. Subjective and objective scores before and after treatment were recorded. RESULTS: There was no statistically significant difference on the four objective indicators of pretreatment FL, BUT, SⅠt, and vision between the two groups(P>0.05. Dry eye symptom scores of the two groups decreased after treatment, both with significantly different(PPPPCONCLUSION: The clinical effect of praprofen on the treatment of moderate to severe dry eye with the deproteinized calf blood extract is better.

The Influence of Sample Drying Procedures on Mercury Concentrations Analyzed in Soils

Czech Academy of Sciences Publication Activity Database

Hojdová, Maria; Rohovec, Jan; Chrastný, V.; Penížek, V.; Navrátil, Tomáš

2015-01-01

Roč. 94, č. 5 (2015), s. 570-576 ISSN 0007-4861 R&D Projects: GA ČR GP526/09/P404; GA ČR(CZ) GAP210/11/1369 Institutional support: RVO:67985831 Keywords : sample preparation * drying procedures * microbial activity * freeze-drying * contamination Subject RIV: DD - Geochemistry Impact factor: 1.191, year: 2015

Field study of dried blood spot specimens for HIV-1 drug resistance genotyping.

Science.gov (United States)

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

2014-08-01

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 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. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Blood Sample Transportation by Pneumatic Transportation Systems

DEFF Research Database (Denmark)

Nybo, Mads; Lund, Merete E; Titlestad, Kjell

2018-01-01

BACKGROUND: Pneumatic transportation systems (PTSs) are increasingly used for transportation of blood samples to the core laboratory. Many studies have investigated the impact of these systems on different types of analyses, but to elucidate whether PTSs in general are safe for transportation...... analysis, and the hemolysis index). CONCLUSIONS: Owing to their high degree of heterogeneity, the retrieved studies were unable to supply evidence for the safety of using PTSs for blood sample transportation. In consequence, laboratories need to measure and document the actual acceleration forces...

A femoral arteriovenous shunt facilitates arterial whole blood sampling in animals

International Nuclear Information System (INIS)

Weber, Bruno; Burger, Cyrill; Buck, Alfred; Biro, Peter

2002-01-01

In this study we evaluated on-line continuous blood sampling in a femoral arteriovenous (a-v) shunt for use in quantitative tracer studies using gamma-emitting radionuclides in animals. The shunt consisted of 40 cm polyethylene tubing (PE-50) guided through a coincidence probe. Two three-way valves allowed blood pressure measurements and tracer injection. Blood flow in the shunt and the impulse response function (IRF) were assessed using heparinized human blood mixed with fluorine-18 fluorodeoxyglucose (FDG). In vivo experiments were performed in eight male rats (300-350 g) anaesthetized with halothane. In three rats, manual blood sampling was performed in parallel with on-line sampling. In another five animals, the arterial whole blood activity was recorded on-line for 40 min. For the experiments 150-180 MBq FDG was injected over 35 s. Blood flow in the shunt was 23.6, 29.2 and 42.8 ml/h at 100, 120 and 160 mmHg, respectively. The IRF was characterized by minimal dispersion (1-2 s FWHM). Deconvolution of the measured arterial input curves with the IRF changed the measured curve only minimally. Whole blood radioactivity concentration derived from manual and on-line sampling were in excellent agreement. The curves derived from on-line sampling were of high statistical quality. In conclusion, a femoral a-v shunt allows multiple manipulations such as measurement of the arterial whole blood activity, continuous blood pressure monitoring, injection of the tracer and collection of blood samples if necessary. It is not associated with blood loss if the collection of blood samples is not required. It is more convenient to use than manual sampling, the peak of the input curve is never missed and the input curves are of high statistical quality. (orig.)

High-throughput miRNA profiling of human melanoma blood samples

Directory of Open Access Journals (Sweden)

Rass Knuth

2010-06-01

Full Text Available Abstract Background MicroRNA (miRNA signatures are not only found in cancer tissue but also in blood of cancer patients. Specifically, miRNA detection in blood offers the prospect of a non-invasive analysis tool. Methods Using a microarray based approach we screened almost 900 human miRNAs to detect miRNAs that are deregulated in their expression in blood cells of melanoma patients. We analyzed 55 blood samples, including 20 samples of healthy individuals, 24 samples of melanoma patients as test set, and 11 samples of melanoma patients as independent validation set. Results A hypothesis test based approch detected 51 differentially regulated miRNAs, including 21 miRNAs that were downregulated in blood cells of melanoma patients and 30 miRNAs that were upregulated in blood cells of melanoma patients as compared to blood cells of healthy controls. The tets set and the independent validation set of the melanoma samples showed a high correlation of fold changes (0.81. Applying hierarchical clustering and principal component analysis we found that blood samples of melanoma patients and healthy individuals can be well differentiated from each other based on miRNA expression analysis. Using a subset of 16 significant deregulated miRNAs, we were able to reach a classification accuracy of 97.4%, a specificity of 95% and a sensitivity of 98.9% by supervised analysis. MiRNA microarray data were validated by qRT-PCR. Conclusions Our study provides strong evidence for miRNA expression signatures of blood cells as useful biomarkers for melanoma.

The pathology of facial vein blood sampling in mice

DEFF Research Database (Denmark)

Hansen, Ket; Harslund, Jakob le Fèvre; Bollen, Peter

2014-01-01

vein blood sampling. Therefore, we investigated if this technique was associated with pathological changes of the jaw region. Methods: 43 NMRI mice were subjected to facial vein blood sampling by using the lancet method during 12 months, starting at the age of 8 weeks. The mice were restrained manually......, and the tissue of the jaw was evaluated. Results: In the 23 mice, from which blood samples had been taken 2 days previously, 5 mice had no signs of gross pathological changes, whereas 12 mice had signs of minimal local subcutaneous bleeding and 6 mice had moderate local subcutaneous bleeding. No additional gross...... pathological changes were observed. In the 23 mice, from which blood samples had been taken 4 weeks earlier, no hemorrhage or signs of scar tissue formation could be observed. Histological slides are currently being processed (HE staining) and will be evaluated and discussed....

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

Science.gov (United States)

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

INAA of human and animal whole blood samples by short term reactor irradiation. [Au, Br, Cd, Ce, Cl, Cr, Cu, Fe, Hg, K, La, Mn, Na, P, Sc, Zn

Energy Technology Data Exchange (ETDEWEB)

Samudralwar, D L; Garg, A N

1986-11-03

Instrumental neutron activation analysis was employed for the determination of 15 major, minor and trace elements in human and animal blood samples. Dry whole blood samples along with NBS and IAEA standards were irradiated for 5 min, 1 h, 5 h and 10 h with reactor thermal neutrons and counted using high resolution ..gamma..-spectrometry at successive intervals. Data for a new IAEA proposed CRM Mixed Human Diet (H-9) is reported. 28 references, 4 tables.

Clinical Validation of Simultaneous Analysis of Tacrolimus, Cyclosporine A, and Creatinine in Dried Blood Spots in Kidney Transplant Patients.

Science.gov (United States)

Veenhof, Herman; Koster, Remco A; Alffenaar, Jan-Willem C; Berger, Stefan P; Bakker, Stephan J L; Touw, Daan J

2017-07-01

Monitoring of creatinine and immunosuppressive drug concentrations, such as tacrolimus (TaC) and cyclosporin A (CsA), is important in the outpatient follow-up of kidney transplant recipients. Monitoring by dried blood spot (DBS) provides patients the opportunity to sample a drop of blood from a fingerprick at home, which can be sent to the laboratory by mail. We performed a clinical validation in which we compared measurements from whole-blood samples obtained by venapuncture with measurements from DBS samples simultaneously obtained by fingerprick. After exclusion of 10 DBS for poor quality, and 2 for other reasons, 199, 104, and 58 samples from a total of 172 patients were available for validation of creatinine, TaC and CsA, respectively. Validation was performed by means of Passing & Bablok regression, and bias was assessed by Bland-Altman analysis. For creatinine, we found y = 0.73x - 1.55 (95% confidence interval [95% CI] slope, 0.71-0.76), giving the conversion formula: (creatinine plasma concentration in μmol/L) = (creatinine concentration in DBS in μmol/L)/0.73, with a nonclinically relevant bias of -2.1 μmol/L (95% CI, -3.7 to -0.5 μmol/L). For TaC, we found y = 1.00x - 0.23 (95% CI slope, 0.91-1.08), with a nonclinically relevant bias of -0.28 μg/L (95% CI, -0.45 to -0.12 μg/L). For CsA, we found y = 0.99x - 1.86 (95% CI slope, 0.91-1.08) and no significant bias. Therefore, for neither TaC nor CsA, a conversion formula is required. DBS sampling for the simultaneous analysis of immunosuppressants and creatinine can replace conventional venous sampling in daily routine.

Automated blood-sample handling in the clinical laboratory.

Science.gov (United States)

Godolphin, W; Bodtker, K; Uyeno, D; Goh, L O

1990-09-01

The only significant advances in blood-taking in 25 years have been the disposable needle and evacuated blood-drawing tube. With the exception of a few isolated barcode experiments, most sample-tracking is performed through handwritten or computer-printed labels. Attempts to reduce the hazards of centrifugation have resulted in air-tight lids or chambers, the use of which is time-consuming and cumbersome. Most commonly used clinical analyzers require serum or plasma, distributed into specialized containers, unique to that analyzer. Aliquots for different tests are prepared by handpouring or pipetting. Moderate to large clinical laboratories perform so many different tests that even multi-analyzers performing multiple analyses on a single sample may account for only a portion of all tests ordered for a patient. Thus several aliquots of each specimen are usually required. We have developed a proprietary serial centrifuge and blood-collection tube suitable for incorporation into an automated or robotic sample-handling system. The system we propose is (a) safe--avoids or prevents biological danger to the many "handlers" of blood; (b) small--minimizes the amount of sample taken and space required to adapt to the needs of satellite and mobile testing, and direct interfacing with analyzers; (c) serial--permits each sample to be treated according to its own "merits," optimizes throughput, and facilitates flexible automation; and (d) smart--ensures quality results through monitoring and intelligent control of patient identification, sample characteristics, and separation process.

Potassium-based algorithm allows correction for the hematocrit bias in quantitative analysis of caffeine and its major metabolite in dried blood spots.

Science.gov (United States)

De Kesel, Pieter M M; Capiau, Sara; Stove, Veronique V; Lambert, Willy E; Stove, Christophe P

2014-10-01

Although dried blood spot (DBS) sampling is increasingly receiving interest as a potential alternative to traditional blood sampling, the impact of hematocrit (Hct) on DBS results is limiting its final breakthrough in routine bioanalysis. To predict the Hct of a given DBS, potassium (K(+)) proved to be a reliable marker. The aim of this study was to evaluate whether application of an algorithm, based upon predicted Hct or K(+) concentrations as such, allowed correction for the Hct bias. Using validated LC-MS/MS methods, caffeine, chosen as a model compound, was determined in whole blood and corresponding DBS samples with a broad Hct range (0.18-0.47). A reference subset (n = 50) was used to generate an algorithm based on K(+) concentrations in DBS. Application of the developed algorithm on an independent test set (n = 50) alleviated the assay bias, especially at lower Hct values. Before correction, differences between DBS and whole blood concentrations ranged from -29.1 to 21.1%. The mean difference, as obtained by Bland-Altman comparison, was -6.6% (95% confidence interval (CI), -9.7 to -3.4%). After application of the algorithm, differences between corrected and whole blood concentrations lay between -19.9 and 13.9% with a mean difference of -2.1% (95% CI, -4.5 to 0.3%). The same algorithm was applied to a separate compound, paraxanthine, which was determined in 103 samples (Hct range, 0.17-0.47), yielding similar results. In conclusion, a K(+)-based algorithm allows correction for the Hct bias in the quantitative analysis of caffeine and its metabolite paraxanthine.

Measurement of regional cerebral blood flow using one-point venous blood sampling and causality model. Evaluation by comparing with conventional continuous arterial blood sampling method

International Nuclear Information System (INIS)

Mimura, Hiroaki; Sone, Teruki; Takahashi, Yoshitake

2008-01-01

Optimal setting of the input function is essential for the measurement of regional cerebral blood flow (rCBF) based on the microsphere model using N-isopropyl-4-[ 123 I]iodoamphetamine ( 123 I-IMP), and usually the arterial 123 I-IMP concentration (integral value) in the initial 5 min is used for this purpose. We have developed a new convenient method in which 123 I-IMP concentration in arterial blood sample is estimated from that in venous blood sample. Brain perfusion single photon emission computed tomography (SPECT) with 123 I-IMP was performed in 110 cases of central nervous system disorders. The causality was analyzed between the various parameters of SPECT data and the ratio of octanol-extracted arterial radioactivity concentration during the first 5 min (Caoct) to octanol-extracted venous radioactivity concentration at 27 min after intravenous injection of 123 I-IMP (Cvoct). A high correlation was observed between the measured and estimated values of Caoct/Cvoct (r=0.856) when the following five parameters were included in the regression formula: radioactivity concentration in venous blood sampled at 27 min (Cv), Cvoct, Cvoct/Cv, and total brain radioactivity counts that were measured by a four-head gamma camera 5 min and 28 min after 123 I-IMP injection. Furthermore, the rCBF values obtained using the input parameters estimated by this method were also highly correlated with the rCBF values measured using the continuous arterial blood sampling method (r=0.912). These results suggest that this method would serve as the new, convenient and less invasive method of rCBF measurement in clinical setting. (author)

High-Throughput, Multiplex Genotyping Directly from Blood or Dried Blood Spot without DNA Extraction for the Screening of Multiple G6PD Gene Variants at Risk for Drug-Induced Hemolysis.

Science.gov (United States)

Tian, Xiaoyi; Zhou, Jun; Zhao, Ye; Cheng, Zhibin; Song, Wenqi; Sun, Yu; Sun, Xiaodong; Zheng, Zhi

2017-09-01

Clinical or epidemiologic screening of single-nucleotide polymorphism markers requires large-scale multiplexed genotyping. Available genotyping tools require DNA extraction and multiplex PCR, which may limit throughput and suffer amplification bias. Herein, a novel genotyping approach has been developed, multiplex extension and ligation-based probe amplification (MELPA), which eliminates DNA extraction and achieves uniform PCR amplification. MELPA lyses blood or dried blood spot and directly captures specific target DNA to 96-well plates using tailed probes. Subsequent enzymatic extension and ligation form target single-nucleotide polymorphism-spanning single-stranded templates, which are PCR-amplified using universal primers. Multiplexed genotyping by single-base primer extension is analyzed by mass spectrometry, with a call rate >97%. MELPA was compared with a commercial assay (iPLEX) for detecting 24 G6PD variants known to be at risk for primaquine-induced hemolysis. MELPA provided results that were more reliable than iPLEX, with higher throughput and lower cost. Genotyping archival blood from 106 malaria patients taking primaquine found 10 G6PD-deficient variants, including 1 patient with a hemizygous Mahidol mutation who had hemolysis. Preemptive G6PD genotyping of 438 dried blood spots from a malaria-endemic area identified three variants. MELPA also enabled pooled genotyping without diluting rare alleles, in which undesired common-allele background increased by sample pooling can be repressed by adding specific common allele blockers. Thus, MELPA represents a high-throughput, cost-effective approach to targeted genotyping at the population level. Copyright © 2017 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

Characterization of the disposition of fostamatinib in Japanese subjects including pharmacokinetic assessment in dry blood spots: results from two phase I clinical studies.

Science.gov (United States)

Martin, Paul; Cheung, S Y Amy; Yen, Mark; Han, David; Gillen, Michael

2016-01-01

The aims of the present study were to characterize the pharmacokinetics of fostamatinib in two phase I studies in healthy Japanese subjects after single- and multiple-dose administration, and to evaluate the utility of dried blood spot (DBS) sampling. In study A, 40 Japanese and 16 white subjects were randomized in a double-blind parallel group study consisting of seven cohorts, which received either placebo or a fostamatinib dose between 50 and 200 mg after single and multiple dosing. Pharmacokinetics of R406 (active metabolite of fostamatinib) in plasma and urine was assessed, and safety was intensively monitored. Study B was an open-label study that assessed fostamatinib 100 and 200 mg in 24 Japanese subjects. In addition to plasma and urine sampling (as for study A), pharmacokinetics was also assessed in blood. Mean maximum plasma concentration (C max) and area under total plasma concentration–time curve (AUC) increased with increasing dose in Japanese subjects. Steady state was achieved in 5–7 days for all doses. C max and AUC were both higher in Japanese subjects administered a 150-mg single dose than in white subjects. This difference was maintained for steady state exposure by day 10. Overall, R406 blood concentrations were consistent and ∼2.5-fold higher than in plasma. Minimal (blood cells, and DBS sampling was a useful method for assessing R406 pharmacokinetics.

Volumetric adsorptive microsampling-liquid chromatography tandem mass spectrometry assay for the simultaneous quantification of four antibiotics in human blood: Method development, validation and comparison with dried blood spot.

Science.gov (United States)

Barco, Sebastiano; Castagnola, Elio; Moscatelli, Andrea; Rudge, James; Tripodi, Gino; Cangemi, Giuliana

2017-10-25

In this paper we show the development and validation of a volumetric absorptive microsampling (VAMS™)-LC-MS/MS method for the simultaneous quantification of four antibiotics: piperacillin-tazobactam, meropenem, linezolid and ceftazidime in 10μL human blood. The novel VAMS-LC-MS/MS method has been compared with a dried blood spot (DBS)-based method in terms of impact of hematocrit (HCT) on accuracy, reproducibility, recovery and matrix effect. Antibiotics were extracted from VAMS and DBS by protein precipitation with methanol after a re-hydration step at 37°C for 10min. LC-MS/MS was carried out on a Thermo Scientific™ TSQ Quantum™ Access MAX triple quadrupole coupled to an Accela ™UHPLC system. The VAMS-LC-MS/MS method is selective, precise and reproducible. In contrast to DBS, it allows an accurate quantification without any HCT influence. It has been applied to samples derived from pediatric patients under therapy. VAMS is a valid alternative sampling strategy for the quantification of antibiotics and is valuable in support of clinical PK/PD studies and consequently therapeutic drug monitoring (TDM) in pediatrics. Copyright © 2017 Elsevier B.V. All rights reserved.

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

NARCIS (Netherlands)

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

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

Identifying the potential of changes to blood sample logistics using simulation

DEFF Research Database (Denmark)

Jørgensen, Pelle Morten Thomas; Jacobsen, Peter; Poulsen, Jørgen Hjelm

2013-01-01

of the simulation was to evaluate changes made to the transportation of blood samples between wards and the laboratory. The average- (AWT) and maximum waiting time (MWT) from a blood sample was drawn at the ward until it was received at the laboratory, and the distribution of arrivals of blood samples......, each of the scenarios was tested in terms of what amount of resources would give the optimal result. The simulations showed a big improvement potential in implementing a new technology/mean for transporting the blood samples. The pneumatic tube system showed the biggest potential lowering the AWT...

Air bubbles and hemolysis of blood samples during transport by pneumatic tube systems.

Science.gov (United States)

Mullins, Garrett R; Bruns, David E

2017-10-01

Transport of blood samples through pneumatic tube systems (PTSs) generates air bubbles in transported blood samples and, with increasing duration of transport, the appearance of hemolysis. We investigated the role of air-bubble formation in PTS-induced hemolysis. Air was introduced into blood samples for 0, 1, 3 or 5min to form air bubbles. Hemolysis in the blood was assessed by (H)-index, lactate dehydrogenase (LD) and potassium in plasma. In an effort to prevent PTS-induced hemolysis, blood sample tubes were completely filled, to prevent air bubble formation, and compared with partially filled samples after PTS transport. We also compared hemolysis in anticoagulated vs clotted blood subjected to PTS transport. As with transport through PTSs, the duration of air bubble formation in blood by a gentle stream of air predicted the extent of hemolysis as measured by H-index (pair space in a blood sample prevented bubble formation and fully protected the blood from PTS-induced hemolysis (ptransport and was partially protected from hemolysis vs anticoagulated blood as indicated by lower LD (ptransport. Prevention of air bubble formation in blood samples during PTS transport protects samples from hemolysis. Copyright © 2017 Elsevier B.V. All rights reserved.

Randomized comparison of vaginal self-sampling by standard vs. dry swabs for Human papillomavirus testing

International Nuclear Information System (INIS)

Eperon, Isabelle; Vassilakos, Pierre; Navarria, Isabelle; Menoud, Pierre-Alain; Gauthier, Aude; Pache, Jean-Claude; Boulvain, Michel; Untiet, Sarah; Petignat, Patrick

2013-01-01

To evaluate if human papillomavirus (HPV) self-sampling (Self-HPV) using a dry vaginal swab is a valid alternative for HPV testing. Women attending colposcopy clinic were recruited to collect two consecutive Self-HPV samples: a Self-HPV using a dry swab (S-DRY) and a Self-HPV using a standard wet transport medium (S-WET). These samples were analyzed for HPV using real time PCR (Roche Cobas). Participants were randomized to determine the order of the tests. Questionnaires assessing preferences and acceptability for both tests were conducted. Subsequently, women were invited for colposcopic examination; a physician collected a cervical sample (physician-sampling) with a broom-type device and placed it into a liquid-based cytology medium. Specimens were then processed for the production of cytology slides and a Hybrid Capture HPV DNA test (Qiagen) was performed from the residual liquid. Biopsies were performed if indicated. Unweighted kappa statistics (κ) and McNemar tests were used to measure the agreement among the sampling methods. A total of 120 women were randomized. Overall HPV prevalence was 68.7% (95% Confidence Interval (CI) 59.3–77.2) by S-WET, 54.4% (95% CI 44.8–63.9) by S-DRY and 53.8% (95% CI 43.8–63.7) by HC. Among paired samples (S-WET and S-DRY), the overall agreement was good (85.7%; 95% CI 77.8–91.6) and the κ was substantial (0.70; 95% CI 0.57-0.70). The proportion of positive type-specific HPV agreement was also good (77.3%; 95% CI 68.2-84.9). No differences in sensitivity for cervical intraepithelial neoplasia grade one (CIN1) or worse between the two Self-HPV tests were observed. Women reported the two Self-HPV tests as highly acceptable. Self-HPV using dry swab transfer does not appear to compromise specimen integrity. Further study in a large screening population is needed. ClinicalTrials.gov: http://clinicaltrials.gov/show/NCT01316120

Correlation of Serum and Dried Blood Spot Results for Quantitation of Schistosoma Circulating Anodic Antigen: a Proof of Principle

Science.gov (United States)

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

2015-01-01

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 for prolonged periods of time. This finding opens new avenues for future work among more than 700 million individuals living in areas worldwide in which schistosomes are endemic. PMID:26149541

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

Science.gov (United States)

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 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

Blood sampling and hemolysis affect concentration of plasma metabolites

DEFF Research Database (Denmark)

Theil, Peter Kappel; Pedersen, Lene Juul; Jensen, Margit Bak

2012-01-01

design and blood was collected after restraint via vein puncture 1, 4, 11, and 23 h after morning feeding. Plasma samples were categorized as without or with minor or major hemolysis [clear (n = 218), yellow (n = 97), or red (n = 37)] upon centrifugation. Plasma NEFA (P ...Two experiments were carried out to reveal and quantify plasma metabolites that are sensitive to hemolysis and animal stress due to the blood sampling procedure (vein puncture vs. catheter). In Exp. 1, 48 sows were fed 4 diets either once (0800 h) or twice daily (0800 h and 1500 h) in a crossover......, a subset of samples from 24 sows fed twice daily in Exp. 1 was combined with data obtained from 30 sows sampled using jugular vein catheters. All sows in Exp. 2 were fed twice daily (0800 h and 1500 h) and blood samples collected repeatedly 1, 4, 11, and 23 h after morning feeding (other conditions were...

Extensive monitoring through multiple blood samples in professional soccer players

DEFF Research Database (Denmark)

Heisterberg, Mette F; Fahrenkrug, Jan; Krustrup, Peter

2013-01-01

of the season. Leucocytes decreased with increased physical training. Lymphocytes decreased at the end of the season. VO2max decreased towards the end of the season whereas no significant changes were observed in the IE2 test.The regular blood samples from elite soccer players reveal significant changes......ABSTRACT: The aim of this study was to make a comprehensive gathering of consecutive detailed blood samples from professional soccer players, and to analyze different blood parameters in relation to seasonal changes in training and match exposure.Blood samples were collected five times during a six...... months period and analyzed for 37 variables in 27 professional soccer players from the best Danish league. Additionally, players were tested for body composition, VO2max and physical performance by the Yo-Yo intermittent endurance sub-max test (IE2).Multiple variations in blood parameters occurred during...

Automated Blood Sample Preparation Unit (ABSPU) for Portable Microfluidic Flow Cytometry.

Science.gov (United States)

Chaturvedi, Akhil; Gorthi, Sai Siva

2017-02-01

Portable microfluidic diagnostic devices, including flow cytometers, are being developed for point-of-care settings, especially in conjunction with inexpensive imaging devices such as mobile phone cameras. However, two pervasive drawbacks of these have been the lack of automated sample preparation processes and cells settling out of sample suspensions, leading to inaccurate results. We report an automated blood sample preparation unit (ABSPU) to prevent blood samples from settling in a reservoir during loading of samples in flow cytometers. This apparatus automates the preanalytical steps of dilution and staining of blood cells prior to microfluidic loading. It employs an assembly with a miniature vibration motor to drive turbulence in a sample reservoir. To validate performance of this system, we present experimental evidence demonstrating prevention of blood cell settling, cell integrity, and staining of cells prior to flow cytometric analysis. This setup is further integrated with a microfluidic imaging flow cytometer to investigate cell count variability. With no need for prior sample preparation, a drop of whole blood can be directly introduced to the setup without premixing with buffers manually. Our results show that integration of this assembly with microfluidic analysis provides a competent automation tool for low-cost point-of-care blood-based diagnostics.

Non-terminal blood sampling techniques in Guinea pigs

DEFF Research Database (Denmark)

Birck, Malene Muusfeldt; Tveden-Nyborg, Pernille; Lindblad, Maiken Marie

2014-01-01

Guinea pigs possess several biological similarities to humans and are validated experimental animal models(1-3). However, the use of guinea pigs currently represents a relatively narrow area of research and descriptive data on specific methodology is correspondingly scarce. The anatomical features...... of guinea pigs are slightly different from other rodent models, hence modulation of sampling techniques to accommodate for species-specific differences, e.g., compared to mice and rats, are necessary to obtain sufficient and high quality samples. As both long and short term in vivo studies often require...... repeated blood sampling the choice of technique should be well considered in order to reduce stress and discomfort in the animals but also to ensure survival as well as compliance with requirements of sample size and accessibility. Venous blood samples can be obtained at a number of sites in guinea pigs e...

Randomized Comparison of Two Vaginal Self-Sampling Methods for Human Papillomavirus Detection: Dry Swab versus FTA Cartridge.

Science.gov (United States)

Catarino, Rosa; Vassilakos, Pierre; Bilancioni, Aline; Vanden Eynde, Mathieu; Meyer-Hamme, Ulrike; Menoud, Pierre-Alain; Guerry, Frédéric; Petignat, Patrick

2015-01-01

Human papillomavirus (HPV) self-sampling (self-HPV) is valuable in cervical cancer screening. HPV testing is usually performed on physician-collected cervical smears stored in liquid-based medium. Dry filters and swabs are an alternative. We evaluated the adequacy of self-HPV using two dry storage and transport devices, the FTA cartridge and swab. A total of 130 women performed two consecutive self-HPV samples. Randomization determined which of the two tests was performed first: self-HPV using dry swabs (s-DRY) or vaginal specimen collection using a cytobrush applied to an FTA cartridge (s-FTA). After self-HPV, a physician collected a cervical sample using liquid-based medium (Dr-WET). HPV types were identified by real-time PCR. Agreement between collection methods was measured using the kappa statistic. HPV prevalence for high-risk types was 62.3% (95%CI: 53.7-70.2) detected by s-DRY, 56.2% (95%CI: 47.6-64.4) by Dr-WET, and 54.6% (95%CI: 46.1-62.9) by s-FTA. There was overall agreement of 70.8% between s-FTA and s-DRY samples (kappa = 0.34), and of 82.3% between self-HPV and Dr-WET samples (kappa = 0.56). Detection sensitivities for low-grade squamous intraepithelial lesion or worse (LSIL+) were: 64.0% (95%CI: 44.5-79.8) for s-FTA, 84.6% (95%CI: 66.5-93.9) for s-DRY, and 76.9% (95%CI: 58.0-89.0) for Dr-WET. The preferred self-collection method among patients was s-DRY (40.8% vs. 15.4%). Regarding costs, FTA card was five times more expensive than the swab (~5 US dollars (USD)/per card vs. ~1 USD/per swab). Self-HPV using dry swabs is sensitive for detecting LSIL+ and less expensive than s-FTA. International Standard Randomized Controlled Trial Number (ISRCTN): 43310942.

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

OpenAIRE

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

2013-01-01

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,...

Effect of order of draw of blood samples during phlebotomy on routine biochemistry results.

Science.gov (United States)

Sulaiman, Raashda A; Cornes, Michael P; Whitehead, Simon J; Othonos, Nadia; Ford, Clare; Gama, Rousseau

2011-11-01

To investigate whether incorrect order of draw of blood samples during phlebotomy causes in vitro potassium ethylenediaminetetraacetic acid EDTA (kEDTA) contamination of blood samples. Serum kEDTA, potassium, calcium, magnesium, alkaline phosphatase, zinc and iron concentrations were measured in blood samples drawn before and after collecting blood into kEDTA containing sample tubes by an experienced phlebotomist using the Sarstedt Safety Monovette system. EDTA was undetectable in all samples. The concentrations of other analytes were similar in blood samples drawn before and after collection of the EDTA blood sample. Order of draw of blood samples using the Sarstedt Safety Monovette system has no effect on serum biochemistry results, when samples are taken by an experienced phlebotomist.

Investigation into solar drying of potato: effect of sample geometry on drying kinetics and CO2 emissions mitigation.

Science.gov (United States)

Tripathy, P P

2015-03-01

Drying experiments have been performed with potato cylinders and slices using a laboratory scale designed natural convection mixed-mode solar dryer. The drying data were fitted to eight different mathematical models to predict the drying kinetics, and the validity of these models were evaluated statistically through coefficient of determination (R(2)), root mean square error (RMSE) and reduced chi-square (χ (2)). The present investigation showed that amongst all the mathematical models studied, the Modified Page model was in good agreement with the experimental drying data for both potato cylinders and slices. A mathematical framework has been proposed to estimate the performance of the food dryer in terms of net CO2 emissions mitigation potential along with unit cost of CO2 mitigation arising because of replacement of different fossil fuels by renewable solar energy. For each fossil fuel replaced, the gross annual amount of CO2 as well as net amount of annual CO2 emissions mitigation potential considering CO2 emissions embodied in the manufacture of mixed-mode solar dryer has been estimated. The CO2 mitigation potential and amount of fossil fuels saved while drying potato samples were found to be the maximum for coal followed by light diesel oil and natural gas. It was inferred from the present study that by the year 2020, 23 % of CO2 emissions can be mitigated by the use of mixed-mode solar dryer for drying of agricultural products.

Dried blood spot analysis of creatinine with LC-MS/MS in addition to immunosuppressants analysis

NARCIS (Netherlands)

Koster, Remco A.; Greijdanus, Ben; Alffenaar, Jan-Willem C.; Touw, Daan J.

In order to monitor creatinine levels or to adjust the dosage of renally excreted or nephrotoxic drugs, the analysis of creatinine in dried blood spots (DBS) could be a useful addition to DBS analysis. We developed a LC-MS/MS method for the analysis of creatinine in the same DBS extract that was

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

Science.gov (United States)

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.

Membrane materials for storing biological samples intended for comparative nanotoxicological testing

Science.gov (United States)

Metelkin, A.; Kuznetsov, D.; Kolesnikov, E.; Chuprunov, K.; Kondakov, S.; Osipov, A.; Samsonova, J.

2015-11-01

The study is aimed at identifying the samples of most promising membrane materials for storing dry specimens of biological fluids (Dried Blood Spots, DBS technology). Existing sampling systems using cellulose fiber filter paper have a number of drawbacks such as uneven distribution of the sample spot, dependence of the spot spreading area on the individual biosample properties, incomplete washing-off of the sample due to partially inconvertible sorption of blood components on cellulose fibers, etc. Samples of membrane materials based on cellulose, polymers and glass fiber with applied biosamples were studied using methods of scanning electron microscopy, FT-IR spectroscopy and surface-wetting measurement. It was discovered that cellulose-based membrane materials sorb components of biological fluids inside their structure, while membranes based on glass fiber display almost no interaction with the samples and biological fluid components dry to films in the membrane pores between the structural fibers. This characteristic, together with the fact that membrane materials based on glass fiber possess sufficient strength, high wetting properties and good storage capacity, attests them as promising material for dry samples of biological fluids storage systems.

Membrane materials for storing biological samples intended for comparative nanotoxicological testing

International Nuclear Information System (INIS)

Metelkin, A; Kuznetsov, D; Kolesnikov, E; Chuprunov, K; Kondakov, S; Osipov, A; Samsonova, J

2015-01-01

The study is aimed at identifying the samples of most promising membrane materials for storing dry specimens of biological fluids (Dried Blood Spots, DBS technology). Existing sampling systems using cellulose fiber filter paper have a number of drawbacks such as uneven distribution of the sample spot, dependence of the spot spreading area on the individual biosample properties, incomplete washing-off of the sample due to partially inconvertible sorption of blood components on cellulose fibers, etc. Samples of membrane materials based on cellulose, polymers and glass fiber with applied biosamples were studied using methods of scanning electron microscopy, FT-IR spectroscopy and surface-wetting measurement. It was discovered that cellulose-based membrane materials sorb components of biological fluids inside their structure, while membranes based on glass fiber display almost no interaction with the samples and biological fluid components dry to films in the membrane pores between the structural fibers. This characteristic, together with the fact that membrane materials based on glass fiber possess sufficient strength, high wetting properties and good storage capacity, attests them as promising material for dry samples of biological fluids storage systems. (paper)

Improving blood sample logistics using simulation

DEFF Research Database (Denmark)

Jørgensen, Pelle Morten Thomas; Jacobsen, Peter

2012-01-01

Using simulation as an approach to display and improve internal logistics and handling at hospitals has great potential. This research will show how a simulation model can be used to evaluate changes made to two different cases of transportation of blood samples at a hospital, by evaluating...

Simultaneous measurement of 25 inflammatory markers and neurotrophins in neonatal dried blood spots by immunoassay with xMAP technology

DEFF Research Database (Denmark)

Skogstrand, Kristin; Thorsen, Poul; Nørgaard-Pedersen, Bent

2005-01-01

BACKGROUND: Inflammatory reactions and other events in early life may be part of the etiology of late-onset diseases, including cerebral palsy, autism, and type 1 diabetes. Most neonatal screening programs for congenital disorders are based on analysis of dried blood spot samples (DBSS), and stored...... on flowmetric Luminex xMAP technology to measure inflammatory markers and neutrophins in DBSS. RESULTS: The high-capacity 25-plex multianalyte method measured 23 inflammatory and trophic cytokines, triggering receptor expressed on myeloid cells-1 (TREM-1), and C-reactive protein in two 3.2-mm punches from DBSS...... potential for high-capacity analysis of DBSS in epidemiologic case-control studies and, with further refinements, in neonatal screening....

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

Science.gov (United States)

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.

Validation of curve-fitting method for blood retention of 99mTc-GSA. Comparison with blood sampling method

International Nuclear Information System (INIS)

Ha-Kawa, Sang Kil; Suga, Yutaka; Kouda, Katsuyasu; Ikeda, Koshi; Tanaka, Yoshimasa

1997-01-01

We investigated a curve-fitting method for the rate of blood retention of 99m Tc-galactosyl serum albumin (GSA) as a substitute for the blood sampling method. Seven healthy volunteers and 27 patients with liver disease underwent 99m Tc-GSA scanning. After normalization of the y-intercept as 100 percent, a biexponential regression curve for the precordial time-activity curve provided the percent injected dose (%ID) of 99m Tc-GSA in the blood without blood sampling. The discrepancy between %ID obtained by the curve-fitting method and that by the multiple blood samples was minimal in normal volunteers 3.1±2.1% (mean±standard deviation, n=77 sampling). Slightly greater discrepancy was observed in patients with liver disease (7.5±6.1%, n=135 sampling). The %ID at 15 min after injection obtained from the fitted curve was significantly greater in patients with liver cirrhosis than in the controls (53.2±11.6%, n=13; vs. 31.9±2.8%, n=7, p 99m Tc-GSA and the plasma retention rate for indocyanine green (r=-0.869, p 99m Tc-GSA and could be a substitute for the blood sampling method. (author)

Detection of drugs in 275 alcohol-positive blood samples of Korean drivers.

Science.gov (United States)

Kim, Eunmi; Choe, Sanggil; Lee, Juseon; Jang, Moonhee; Choi, Hyeyoung; Chung, Heesun

2016-08-01

Since driving under the influence of drugs (DUID) is as dangerous as drink-driving, many countries regulate DUID by law. However, laws against the use of drugs while driving are not yet established in Korea. In order to investigate the type and frequency of drugs used by drivers in Korea, we analyzed controlled and non-controlled drugs in alcohol-positive blood samples. Total 275 blood samples were taken from Korean drivers, which were positive in roadside alcohol testing. The following analyses were performed: blood alcohol concentrations by GC; screening for controlled drugs by immunoassay and confirmation for positive samples by GC-MS. For the detection of DUID related drugs in blood samples, a total of 49 drugs were selected and were examined by GC-MS. For a rapid detection of these drugs, an automated identification software called "DrugMan" was used. Concentrations of alcohol in 275 blood samples ranged from 0.011 to 0.249% (average 0.119%). Six specimens showed positive results by immunoassay: one methamphetamine and five benzodiazepines I. By GC-MS confirmation, only benzodiazepines in four cases were identified, while methamphetamine and benzodiazepine in two cases were not detected from the presumptive positive blood samples. Using DrugMan, four drugs were detected; chlorpheniramine (5)*, diazepam (4), dextromethorphan (1) and doxylamine (1). In addition, ibuprofen (1), lidocaine (1) and topiramate (1) were also detected as general drugs in blood samples ('*' indicates frequency). The frequency of drug abuse by Korean drivers was relatively low and a total 14 cases were positive in 275 blood samples with a ratio of 5%. However it is necessary to analyze more samples including alcohol negative blood, and to expand the range of drug lists to get the detailed information. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Randomized Comparison of Two Vaginal Self-Sampling Methods for Human Papillomavirus Detection: Dry Swab versus FTA Cartridge.

Directory of Open Access Journals (Sweden)

Rosa Catarino

Full Text Available Human papillomavirus (HPV self-sampling (self-HPV is valuable in cervical cancer screening. HPV testing is usually performed on physician-collected cervical smears stored in liquid-based medium. Dry filters and swabs are an alternative. We evaluated the adequacy of self-HPV using two dry storage and transport devices, the FTA cartridge and swab.A total of 130 women performed two consecutive self-HPV samples. Randomization determined which of the two tests was performed first: self-HPV using dry swabs (s-DRY or vaginal specimen collection using a cytobrush applied to an FTA cartridge (s-FTA. After self-HPV, a physician collected a cervical sample using liquid-based medium (Dr-WET. HPV types were identified by real-time PCR. Agreement between collection methods was measured using the kappa statistic.HPV prevalence for high-risk types was 62.3% (95%CI: 53.7-70.2 detected by s-DRY, 56.2% (95%CI: 47.6-64.4 by Dr-WET, and 54.6% (95%CI: 46.1-62.9 by s-FTA. There was overall agreement of 70.8% between s-FTA and s-DRY samples (kappa = 0.34, and of 82.3% between self-HPV and Dr-WET samples (kappa = 0.56. Detection sensitivities for low-grade squamous intraepithelial lesion or worse (LSIL+ were: 64.0% (95%CI: 44.5-79.8 for s-FTA, 84.6% (95%CI: 66.5-93.9 for s-DRY, and 76.9% (95%CI: 58.0-89.0 for Dr-WET. The preferred self-collection method among patients was s-DRY (40.8% vs. 15.4%. Regarding costs, FTA card was five times more expensive than the swab (~5 US dollars (USD/per card vs. ~1 USD/per swab.Self-HPV using dry swabs is sensitive for detecting LSIL+ and less expensive than s-FTA.International Standard Randomized Controlled Trial Number (ISRCTN: 43310942.

An automated blood sampling system used in positron emission tomography

International Nuclear Information System (INIS)

Eriksson, L.; Bohm, C.; Kesselberg, M.

1988-01-01

Fast dynamic function studies with positron emission tomography (PET), has the potential to give accurate information of physiological functions of the brain. This capability can be realised if the positron camera system accurately quantitates the tracer uptake in the brain with sufficiently high efficiency and in sufficiently short time intervals. However, in addition, the tracer concentration in blood, as a function of time, must be accurately determined. This paper describes and evaluates an automated blood sampling system. Two different detector units are compared. The use of the automated blood sampling system is demonstrated in studies of cerebral blood flow, in studies of the blood-brain barrier transfer of amino acids and of the cerebral oxygen consumption. 5 refs.; 7 figs

Free radical scavenging capacity and antioxidant activity of methanolic and ethanolic extracts of plum (Prunus domestica L. in both fresh and dried samples

Directory of Open Access Journals (Sweden)

Amin Morabbi Najafabad

2014-09-01

Full Text Available Objectives: Consumption of fruits, such as plums and prunes, is useful in treating blood circulation disorder, measles, digestive disorder, and prevention of cancer, diabetes, and obesity. The paper presents a description of antioxidant and antiradical capacity of plum (Prunus domestica L. in both fresh and dried samples. Materials and Methods: Samples were mixed with methanol and ethanol (as solvents and were extracted on magnetic shaker, separately. The experiments were carried out to measure the Total Phenolic Content (TPC, Total Flavonoid Content (TFC, Total Antioxidant Capacity (TAC, Reducing Power Assay (RPA, Chain Breaking Activity (CBA, and quantity of Malondialdehyde (MDA, 2,2-Diphenyl-1-Picrylhydrazyl (DPPH,Nitric Oxide (NO,Hydrogen peroxide (H2O2 and superoxide(O2- radicals inhibition. Results: The results showed that the highest values for the TPC, TFC,TAC, RPA, CBA, DPPH, and NO were related to ethanolic extractsof dried sample which showed statistically significant differences (p2O2 and O2-were related to ethanolic extracts of fresh sample. The correlations data were analyzed among all parameters and the TPC and TFC had a significant correlation (r2=0.977. Moreover, it was found that methanol was more successful in extraction procedure than ethanol (p

Extensive monitoring through multiple blood samples in professional soccer players.

Science.gov (United States)

Heisterberg, Mette F; Fahrenkrug, Jan; Krustrup, Peter; Storskov, Anders; Kjær, Michael; Andersen, Jesper L

2013-05-01

The aim of this study was to make a comprehensive gathering of consecutive detailed blood samples from professional soccer players and to analyze different blood parameters in relation to seasonal changes in training and match exposure. Blood samples were collected 5 times during a 6-month period and analyzed for 37 variables in 27 professional soccer players from the best Danish league. Additionally, the players were tested for body composition, V[Combining Dot Above]O2max and physical performance by the Yo-Yo intermittent endurance submax test (IE2). Multiple variations in blood parameters occurred during the observation period, including a decrease in hemoglobin and an increase in hematocrit as the competitive season progressed. Iron and transferrin were stable, whereas ferritin showed a decrease at the end of the season. The immunoglobulin A (IgA) and IgM increased in the period with basal physical training and at the end of the season. Leucocytes decreased with increased physical training. Lymphocytes decreased at the end of the season. The V[Combining Dot Above]O2max decreased toward the end of the season, whereas no significant changes were observed in the IE2 test. The regular blood samples from elite soccer players reveal significant changes that may be related to changes in training pattern, match exposure, or length of the match season. Especially the end of the preparation season and at the end of the competitive season seem to be time points were the blood-derived values indicate that the players are under excessive physical strain and might be more subjected to a possible overreaching-overtraining conditions. We suggest that regular analyses of blood samples could be an important initiative to optimize training adaptation, training load, and game participation, but sampling has to be regular, and a database has to be built for each individual player.

The development and validation of dried blood spots for external quality assurance of syphilis serology

Directory of Open Access Journals (Sweden)

Smit Pieter W

2013-02-01

Full Text Available Abstract Background Syphilis causes up to 1,500,000 congenital syphilis cases annually. These could be prevented if all pregnant women were screened, and those with syphilis treated with a single dose of penicillin before 28 weeks gestation. In recent years, rapid point-of-care tests have allowed greater access to syphilis screening, especially in rural or remote areas, but the lack of quality assurance of rapid testing has been a concern. We determined the feasibility of using dried blood spots (DBS as specimens for quality assurance of syphilis serological assays. Methods We developed DBS extraction protocols for use with Treponema pallidum particle agglutination assay (TPPA, Treponema pallidum haemagglutination assay (TPHA and an enzyme immunoassay (EIA and compared the results with those using matching plasma samples from the same patient. Results Since DBS samples showed poor performance with TPHA and EIA (TPHA sensitivity was 50.5% (95% confidence interval: 39.9–61.2% and EIA specificity was 50.4% (95% CI: 43.7–57.1%, only the DBS TPPA was used in the final evaluation. DBS TPPA showed an sensitivity of 95.5% (95% CI: 91.3–98.0% and a specificity of 99.0% (95% CI: 98.1–99.5% compared to TPPA using plasma samples as a reference. Conclusion DBS samples can be recommended for use with TPPA, and may be of value for external quality assurance of point-of-care syphilis testing.

Plasma and breast milk pharmacokinetics of emtricitabine, tenofovir and lamivudine using dried blood and breast milk spots in nursing African mother-infant pairs.

Science.gov (United States)

Waitt, Catriona; Olagunju, Adeniyi; Nakalema, Shadia; Kyohaire, Isabella; Owen, Andrew; Lamorde, Mohammed; Khoo, Saye

2018-04-01

Breast milk transfer of first-line ART from mother to infant is not fully understood. To determine the concentrations of lamivudine, emtricitabine and tenofovir in maternal blood, breast milk and infant blood from breastfeeding mother-infant pairs. Intensive pharmacokinetic sampling of maternal dried blood spots (DBS), dried breast milk spots (DBMS) and infant DBS from 30 Ugandan and 29 Nigerian mothers receiving first-line ART and their infants was conducted. DBS and DBMS were collected pre-dose and at 5-6 timepoints up to 12 h following observed dosing. Infant DBS were sampled twice during this period. Lamivudine, emtricitabine and tenofovir were quantified using LC-MS/MS, with non-compartmental analysis to calculate key pharmacokinetic parameters. Peak concentrations in breast milk from women taking lamivudine and emtricitabine occurred later than in plasma (4-8 h compared with 2 h for lamivudine and 2-4 h for emtricitabine). Consequently, the milk-to-plasma (M:P) ratio of lamivudine taken once daily was 0.95 (0.82-1.15) for AUC0-12, whereas for AUC12-20 this was 3.04 (2.87-4.16). Lamivudine was detectable in 36% (14/39) of infants [median 17.7 (16.3-22.7) ng/mL]. For 200 mg of emtricitabine once daily, the median M:P ratio was 3.01 (2.06-3.38). Three infants (19%) had measurable emtricitabine [median 18.5 (17.6-20.8) ng/mL]. For 300 mg of tenofovir once daily, the median M:P ratio was 0.015 (0-0.03) and no infant had measurable tenofovir concentrations. Emtricitabine and lamivudine accumulate in breast milk and were detected in breastfeeding infants. In contrast, tenofovir penetrates the breast milk to a small degree, but is undetectable in breastfeeding infants.

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

Science.gov (United States)

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.

Preanalytic Factors Associated With Hemolysis in Emergency Department Blood Samples.

Science.gov (United States)

Phelan, Michael P; Reineks, Edmunds Z; Schold, Jesse D; Hustey, Frederic M; Chamberlin, Janelle; Procop, Gary W

2018-02-01

- Hemolysis of emergency department blood samples is a common occurrence and has a negative impact on health care delivery. - To determine the effect of preanalytic factors (straight stick, intravenous [IV] line, needle gauge, location of blood draw, syringe versus vacuum tube use, tourniquet time) on hemolysis in emergency department blood samples. - A single 65 000-visit emergency department's electronic health record was queried for emergency department potassium results and blood draw technique for all samples obtained in calendar year 2014, resulting in 54 531 potassium results. Hemolyzed potassium was measured by hemolysis index. Comparisons of hemolysis by sampling technique were conducted by χ 2 tests. - Overall hemolysis was 10.0% (5439 of 54 531). Hemolysis among samples obtained from straight stick was significantly less than among those obtained with IV line (5.4% [33 of 615] versus 10.2% [4821 of 47 266], P < .001). For IV-placed blood draws, antecubital location had a statistically significant lower overall hemolysis compared with other locations: 7.4% (2117 of 28 786) versus 14.6% (2622 of 17 960) ( P < .001). For blood drawn with a syringe compared with vacuum, hemolysis was 13.0% (92 of 705) and 11.0% (1820 of 16 590), respectively ( P = .09, not significant). For large-gauge IV blood draws versus smaller-gauge IV lines, a lower hemolysis was also observed (9.3% [3882 of 41 571] versus 16.7% [939 of 5633]) ( P < .001). For IV-drawn blood with tourniquet time less than 60 seconds, hemolysis was 10.3% (1362 of 13 162) versus 13.9% for more than 60 seconds (532 of 3832), P < .001. - This study confirmed previous findings that straight stick and antecubital location are significantly associated with reduced hemolysis and indicated that shorter tourniquet time and larger gauge for IV draws were significantly associated with lower hemolysis.

Fingerprick autologous blood: a novel treatment for dry eye syndrome.

Science.gov (United States)

Than, J; Balal, S; Wawrzynski, J; Nesaratnam, N; Saleh, G M; Moore, J; Patel, A; Shah, S; Sharma, B; Kumar, B; Smith, J; Sharma, A

2017-12-01

PurposeDry eye syndrome (DES) causes significant morbidity. Trials of blood-derived products in treatment of the condition show promising results. However, their production is expensive and time-consuming. We investigate fingerprick autologous blood (FAB) as an alternative low-cost, readily accessible treatment for DES.Patients and methodsProspective, non-comparative, interventional case series. In total, 29 eyes of 16 DES patients (2 males and 14 females) from two NHS sites in the United Kingdom. Patients instructed to clean a finger, prick with a blood lancet, and apply a drop of blood to the lower fornix of the affected eye(s), 4 times daily for 8 weeks then stop and review 4 weeks later. Follow-up visits occurred ~3 days, 2, 4, 8 weeks into therapy, and 4 weeks post-cessation. At each visit, visual acuity, corneal staining, Schirmer's test, tear break-up time (TBUT), and ocular comfort index (OCI) were measured, and photographs taken. Results were analysed using Student's paired t-test.ResultsAt 8 weeks, there was improvement in mean Oxford corneal staining grade (3.31 to 2.07 (P<0.0001)), TBUT (5.00 to 7.80 s (P<0.05)), visual acuity (0.08 to 0.01 LogMAR equivalent (P<0.05)), and OCI score (56.03 to 39.72 (P<0.0001)). There was no statistically significant change in Schirmer's test results. Four weeks post-cessation versus immediately after completion of FAB therapy, mean staining grade worsened from 2.07 to 2.86 (P<0.0001). OCI score worsened from 39.72 to 44.67 (P<0.05).ConclusionsIn our limited case series FAB appears to be a safe and effective treatment for DES.

Investigation of microwave assisted drying of samples and evaporation of aqueous solutions in trace element analysis

Energy Technology Data Exchange (ETDEWEB)

Maichin, B.; Knapp, G. [Technische Univ., Graz (Austria). Inst. fuer Analytische Chemie, Mikro- und Radiochemie; Kettisch, P. [Anton Paar GmbH, Graz (Austria)

2000-01-01

Investigations of microwave assisted drying of sample materials and microwave assisted evaporation of aqueous sample solutions and acidic digestion residues were accomplished by means of special rotors for the microwave digestion system MULTIWAVE. To check the results obtained by microwave assisted drying, the samples were also conventionally dried at 105 C in an oven. The following samples have been dried: 10 g each of meat, fish, apple, cucumber, potato, mustard, yogurt, clay and marl; 1 g each of certified reference material TORT 2 (lobster hepatopancreas), BCR 278 (mussel tissue) and BCR 422 (cod muscle); 500 g garden mould. Microwave assisted drying takes 40 min for organic samples and 30 min for inorganic material. Important is a slow increase of microwave power during the first 20 min. The results agree well with conventional drying at 105 C. Losses of As, Se and Hg have been investigated for 3 CRMs. Only Se shows losses in the range of 20%. Losses of As, Be, Cd, Co, Cr, Cu, Fe, Hg, Li, Mg, Mn, Mo, Ni, Pb, Sb, Se, Sr, Ti, Tl, V and Zn after evaporation of aqueous samples and acidic solutions after wet digestion, respectively, have been investigated. 50 mL aqueous solution was evaporated almost to dryness within 25 min. The recovery of Hg is 40-50%, of Se 90-95% and of the other elements 97-102%. 0.2 g each of TORT 2, BCR 278 and BCR 422 have been digested with 4 mL nitric acid and 1 mL hydrochloric acid by means of the microwave digestion system MULTIWAVE. The digestion residue was evaporated almost to dryness and dissolved again in 10 mL diluted nitric acid. In this case no element losses have been observed. The measured concentration of As, Cd, Cu, Fe, Mn, Hg, Pb, Mo, Ni, Se, Sr, V and Zn agree very well with the certified values. An important prerequisite for good recoveries is not to evaporate the solutions to complete dryness. (orig.)

Effects of Transport and Storage Conditions on Gene Expression in Blood Samples.

Science.gov (United States)

Malentacchi, Francesca; Pizzamiglio, Sara; Wyrich, Ralf; Verderio, Paolo; Ciniselli, Chiara; Pazzagli, Mario; Gelmini, Stefania

2016-04-01

Inappropriate handling of blood samples might induce or repress gene expression and/or lead to RNA degradation affecting downstream analysis. In particular, sample transport is a critical step for biobanking or multicenter studies because of uncontrolled variables (i.e., unstable temperature). We report the results of a pilot study implemented within the EC funded SPIDIA project, aimed to investigate the role of transport and storage of blood samples containing and not containing an RNA stabilizer. Blood was collected from a single donor both in EDTA and in PAXgene Blood RNA tubes. Half of the samples were sent to a second laboratory both at room temperature and at 4°C, whereas the remaining samples were stored at room temperature and at 4°C. Gene expression of selected genes (c-FOS, IL-1β, IL-8, and GAPDH) known to be induced or repressed by ex vivo blood handling and of blood-mRNA quality biomarkers identified and validated within the SPIDIA project, which allow for monitoring changes in unstabilized blood samples after collection and during transport and storage, were analyzed by RT-qPCR. If the shipment of blood in tubes not containing RNA stabilizer is not performed under a stable condition, gene profile studies can be affected by the effects of transport. Moreover, also controlled temperature shipment (4°C) can influence the expression of specific genes if blood is collected in tubes not containing a stabilizer. The use of dedicated biomarkers or time course experiments should be performed in order to verify potential bias on gene expression analysis due to sample shipment and storage conditions. Alternatively, the use of RNA stabilizer containing tubes can represent a reliable option to avoid ex vivo RNA changes.

The Effect of Storage and Extraction Methods on Amplification of Plasmodium falciparum DNA from Dried Blood Spots

NARCIS (Netherlands)

Schwartz, A.; Baidjoe, A.Y.; Rosenthal, P.J.; Dorsey, G.; Bousema, T.; Greenhouse, B.

2015-01-01

Extraction and amplification of DNA from dried blood spots (DBS) collected in field studies is commonly used for detection of Plasmodium falciparum. However, there have been few systematic efforts to determine the effects of storage and extraction methods on the sensitivity of DNA amplification. We

A technique for extracting blood samples from mice in fire toxicity tests

Science.gov (United States)

Bucci, T. J.; Hilado, C. J.; Lopez, M. T.

1976-01-01

The extraction of adequate blood samples from moribund and dead mice has been a problem because of the small quantity of blood in each animal and the short time available between the animals' death and coagulation of the blood. These difficulties are particularly critical in fire toxicity tests because removal of the test animals while observing proper safety precautions for personnel is time-consuming. Techniques for extracting blood samples from mice were evaluated, and a technique was developed to obtain up to 0.8 ml of blood from a single mouse after death. The technique involves rapid exposure and cutting of the posterior vena cava and accumulation of blood in the peritoneal space. Blood samples of 0.5 ml or more from individual mice have been consistently obtained as much as 16 minutes after apparent death. Results of carboxyhemoglobin analyses of blood appeared reproducible and consistent with carbon monoxide concentrations in the exposure chamber.

Thermal activation of catalytic microjets in blood samples using microfluidic chips.

Science.gov (United States)

Soler, Lluís; Martínez-Cisneros, Cynthia; Swiersy, Anka; Sánchez, Samuel; Schmidt, Oliver G

2013-11-21

We demonstrate that catalytic microjet engines can out-swim high complex media composed of red blood cells and serum. Despite the challenge presented by the high viscosity of the solution at room temperature, the catalytic microjets can be activated at physiological temperature and, consequently, self-propel in diluted solutions of blood samples. We prove that these microjets self-propel in 10× diluted blood samples using microfluidic chips.

Thermal activation of catalytic microjets in blood samples using microfluidic chips†

Science.gov (United States)

Soler, Lluís; Martínez-Cisneros, Cynthia; Swiersy, Anka; Sánchez, Samuel; Schmidt, Oliver G.

2014-01-01

We demonstrate that catalytic microjet engines can out-swim high complex media composed of red blood cells and serum. Despite the challenge presented by the high viscosity of the solution at room temperature, the catalytic microjets can be activated at physiological temperature and, consequently, self-propel in diluted solutions of blood samples. We prove that these microjets self-propel in 10× diluted blood samples using microfluidic chips. PMID:24089195

The impact of freeze-drying infant fecal samples on measures of their bacterial community profiles and milk-derived oligosaccharide content

Directory of Open Access Journals (Sweden)

Zachery T. Lewis

2016-01-01

Full Text Available Infant fecal samples are commonly studied to investigate the impacts of breastfeeding on the development of the microbiota and subsequent health effects. Comparisons of infants living in different geographic regions and environmental contexts are needed to aid our understanding of evolutionarily-selected milk adaptations. However, the preservation of fecal samples from individuals in remote locales until they can be processed can be a challenge. Freeze-drying (lyophilization offers a cost-effective way to preserve some biological samples for transport and analysis at a later date. Currently, it is unknown what, if any, biases are introduced into various analyses by the freeze-drying process. Here, we investigated how freeze-drying affected analysis of two relevant and intertwined aspects of infant fecal samples, marker gene amplicon sequencing of the bacterial community and the fecal oligosaccharide profile (undigested human milk oligosaccharides. No differences were discovered between the fecal oligosaccharide profiles of wet and freeze-dried samples. The marker gene sequencing data showed an increase in proportional representation of Bacteriodes and a decrease in detection of bifidobacteria and members of class Bacilli after freeze-drying. This sample treatment bias may possibly be related to the cell morphology of these different taxa (Gram status. However, these effects did not overwhelm the natural variation among individuals, as the community data still strongly grouped by subject and not by freeze-drying status. We also found that compensating for sample concentration during freeze-drying, while not necessary, was also not detrimental. Freeze-drying may therefore be an acceptable method of sample preservation and mass reduction for some studies of microbial ecology and milk glycan analysis.

Na2EDTA anticoagulant impaired blood samples from the teleost Piaractus mesopotamicus

Directory of Open Access Journals (Sweden)

Thaís Heloisa Vaz Farias

2016-05-01

Full Text Available Abstract: The present study aimed to evaluate the effects of Na heparin and Na2EDTA on blood of Piaractus mesopotamicus (360.7±42.4g, 26.4±1.0cm. Twenty fishes were sampled in two experiment trials, ten for erythrocyte fragility analysis and ten for hematologic and plasma biochemical study. The blood collected by venous-caudal puncture was fractioned and stored in anticoagulants solution: Na2EDTA 10%, Na2EDTA 3%, Na heparin 5000 IU and Na heparin 100 IU. Plasmatic levels of calcium presented in the Na2EDTA stored samples were about 80% lower than both heparin groups. Blood samples of P. mesopotamicus stored with Na2EDTA demonstrated increase in the hematocrit and MCV, and decrease in MCHC. The dose-response effect was observed in this study. The results are reinforced by the higher levels of plasmatic protein and hemolysis presented in the Na2EDTA 10% stored blood, confirming the deleterious effect of this anticoagulant treatment on the quality of blood samples. Na2EDTA is not indicated to store P. mesopotamicus blood samples, but sodium heparin at 100 IU is the most recommended anticoagulant, since this treatment presented the lower rate of alterations in the stored blood.

Pediatric blood sample collection from a pre-existing peripheral intravenous (PIV) catheter.

Science.gov (United States)

Braniff, Heather; DeCarlo, Ann; Haskamp, Amy Corey; Broome, Marion E

2014-01-01

Aiming to minimize pain in a hospitalized child, the purpose of this observational study was to describe characteristics of blood samples collected from pre-existing peripheral intravenous (PIV) catheters in pediatric patients. One hundred and fifty blood samples were reviewed for number of unusable samples requiring a specimen to be re-drawn. Success of the blood draw and prevalence of the loss of the PIV following blood collection was also measured. Findings included one clotted specimen, success rate of 91.3%, and 1.3% of PIVs becoming non-functional after collection. Obtaining blood specimens from a pre-existing PIV should be considered in a pediatric patient. Copyright © 2014 Elsevier Inc. All rights reserved.

Calculation of Blood Dose in Patients Treated With 131I Using MIRD, Imaging, and Blood Sampling Methods.

Science.gov (United States)

Piruzan, Elham; Haghighatafshar, Mahdi; Faghihi, Reza; Entezarmahdi, Seyed Mohammad

2016-03-01

Radioiodine therapy is known as the most effective treatment of differentiated thyroid carcinoma (DTC) to ablate remnant thyroid tissue after surgery. In patients with DTC treated with radioiodine, internal radiation dosimetry of radioiodine is useful for radiation risk assessment. The aim of this study is to describe a method to estimate the absorbed dose to the blood using medical internal radiation dosimetry methods. In this study, 23 patients with DTC with different administrated activities, 3.7, 4.62, and 5.55 GBq after thyroidectomy, were randomly selected. Blood dosimetry of treated patients was performed with external whole body counting using a dual-head gamma camera imaging device and also with blood sample activity measurements using a dose calibrator. Absorbed dose to the blood was measured at 2, 6, 12, 24, 48, and 96 hours after the administration of radioiodine with the 2 methods. Based on the results of whole body counting and blood sample activity dose rate measurements, 96 hours after administration of 3.7, 4.62, and 5.55 GBq of radioiodine, absorbed doses to patients' blood were 0.65 ± 0.20, 0.67 ± 0.18, 0.79 ± 0.51 Gy, respectively. Increasing radioiodine activity from 3.7 to 5.55 GBq increased blood dose significantly, while there was no significant difference in blood dose between radioiodine dosages of 3.7 and 4.62 GBq. Our results revealed a significant correlation between the blood absorbed dose and blood sample activity and between the blood absorbed dose and whole body counts 24 to 48 hours after the administration of radioiodine.

Continuous quality control of the blood sampling procedure using a structured observation scheme

DEFF Research Database (Denmark)

Seemann, T. L.; Nybo, M.

2015-01-01

Background: An important preanalytical factor is the blood sampling procedure and its adherence to the guidelines, i.e. CLSI and ISO 15189, in order to ensure a consistent quality of the blood collection. Therefore, it is critically important to introduce quality control on this part of the process....... As suggested by the EFLM working group on the preanalytical phase we introduced continuous quality control of the blood sampling procedure using a structured observation scheme to monitor the quality of blood sampling performed on an everyday basis. Materials and methods: Based on our own routines the EFLM....... Conclusion: It is possible to establish a continuous quality control on blood sampling. It has been well accepted by the staff and we have already been able to identify critical areas in the sampling process. We find that continuous auditing increase focus on the quality of blood collection which ensures...

Bacterial Isolates from Blood Samples of Patients in University of ...

African Journals Online (AJOL)

Bacterial Isolates from Blood Samples of Patients in University of Benin Teaching Hospital Benin City, Edo State, Nigeria. ... The thioglychollate broth was sub cultured onto blood agar plate for anaerobic incubation, while the brain heart infusion broth was sub cultured onto chocolate, blood agar and McConkey agar for ...

The influence of protein free calf blood extract eye gel on dry eye after pterygium surgery

Directory of Open Access Journals (Sweden)

Cai-Ni Ji

2013-07-01

Full Text Available AIM: To investigate the influence of protein free calf blood extract eye gel on dry eye after pterygium surgery. METHODS: Thirty six patients(40 eyeswith primary nasal pterygium were enrolled in this study, which were divided into study group and control group randomly, with 20 eyes in each group. All patients received pterygium excision and limbal stem cell autograft surgery and tobramicin dexamethasone eye drops after surgery. Patients of the study group received protein free calf blood extract eye gel while those of the control group received 0.1% sodium hyaluronate eye drops furthermore. Ocular surface disease index(OSDIquestionnaire, tear film break-up time(BUTand Schirmer's Ⅰ test Ⅰ(SⅠtwere carried before and 3 months after surgery to evaluate the dry eye degree of the patients. RESULTS: There was no statistical difference between the age, gender and size of the pterygium of the study and control groups preoperatively. There was no statistical difference between the OSDI(2.33±1.02 vs 2.32±0.93, BUT(8.80±2.48 vs 8.35±2.28seconds and SⅠt(4.30±2.30 vs 4.40±2.44of the two groups preoperatively. There was statistical difference between the OSDI(1.45±0.47 vs 1.81±0.60, BUT(11.20±2.07 vs 9.50±2.40seconds and SⅠt(8.35±3.13 vs 6.35±2.18of the two groups 3 months postoperatively, which was also different from that of the preoperative data correspondingly. CONCLUSION: Protein free calf blood extract eye gel could reduce the dry eye after pterygium surgery.

Reliability of blood pressure parameters for dry weight estimation in hemodialysis patients.

Science.gov (United States)

Susantitaphong, Paweena; Laowaloet, Suthanit; Tiranathanagul, Khajohn; Chulakadabba, Adhisabandh; Katavetin, Pisut; Praditpornsilpa, Kearkiat; Tungsanga, Kriang; Eiam-Ong, Somchai

2013-02-01

Chronic volume overload resulting from interdialytic weight gain and inadequate fluid removal plays a significant role in poorly controlled high blood pressure. Although bioimpedance has been introduced as an accurate method for assessing hydration status, the instrument is not available in general hemodialysis (HEMO) centers. This study was conducted to explore the correlation between hydration status measured by bioimpedance and blood pressure parameters in chronic HEMO patients. Multifrequency bioimpedance analysis was used to determine pre- and post-dialysis hydration status in 32 stable HEMO patients. Extracellular water/total body water (ECW/TBW) determined by sum of segments from bioimpedance analysis was used as an index of hydration status. The mean age was 57.9 ± 16.4 years. The mean dry weight and body mass index were 57.7 ± 14.5 kg and 22.3 ± 4.7 kg/m(2), respectively. Pre-dialysis ECW/TBW was significantly correlated with only pulse pressure (r = 0.5, P = 0.003) whereas post-dialysis ECW/TBW had significant correlations with pulse pressure, systolic blood pressure, and diastolic blood pressure (r = 0.6, P = 0.001, r = 0.4, P = 0.04, r = -0.4, and P = 0.02, respectively). After dialysis, the mean values of ECW/TBW, systolic blood pressure, mean arterial pressure, and pulse pressure were significantly decreased. ECW/TBW was used to classify the patients into normohydration (≤ 0.4) and overhydration (>0.4) groups. Systolic blood pressure, mean arterial pressure, and pulse pressure significantly reduced after dialysis in the normohydration group but did not significantly change in the overhydration group. Pre-dialysis pulse pressure, post-dialysis pulse pressure, and post-dialysis systolic blood pressure in the overhydration group were significantly higher than normohydration group. Due to the simplicity and cost, blood pressure parameters, especially pulse pressure, might be a simple reference for clinicians to determine hydration status in HEMO

Simple, miniaturized blood plasma extraction method.

Science.gov (United States)

Kim, Jin-Hee; Woenker, Timothy; Adamec, Jiri; Regnier, Fred E

2013-12-03

A rapid plasma extraction technology that collects a 2.5 μL aliquot of plasma within three minutes from a finger-stick derived drop of blood was evaluated. The utility of the plasma extraction cards used was that a paper collection disc bearing plasma was produced that could be air-dried in fifteen minutes and placed in a mailing envelop for transport to an analytical laboratory. This circumvents the need for venipuncture and blood collection in specialized vials by a phlebotomist along with centrifugation and refrigerated storage. Plasma extraction was achieved by applying a blood drop to a membrane stack through which plasma was drawn by capillary action. During the course of plasma migration to a collection disc at the bottom of the membrane stack blood cells were removed by a combination of adsorption and filtration. After the collection disc filled with an aliquot of plasma the upper membranes were stripped from the collection card and the collection disc was air-dried. Intercard differences in the volume of plasma collected varied approximately 1% while volume variations of less than 2% were seen with hematocrit levels ranging from 20% to 71%. Dried samples bearing metabolites and proteins were then extracted from the disc and analyzed. 25-Hydroxy vitamin D was quantified by LC-MS/MS analysis following derivatization with a secosteroid signal enhancing tag that imparted a permanent positive charge to the vitamin and reduced the limit of quantification (LOQ) to 1 pg of collected vitamin on the disc; comparable to values observed with liquid-liquid extraction (LLE) of a venipuncture sample. A similar study using conventional proteomics methods and spectral counting for quantification was conducted with yeast enolase added to serum as an internal standard. The LOQ with extracted serum samples for enolase was 1 μM, linear from 1 to 40 μM, the highest concentration examined. In all respects protein quantification with extracted serum samples was comparable to

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.

Science.gov (United States)

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.

Radionuclide ratios in wet and dry deposition samples from June 1976 through December 1977

International Nuclear Information System (INIS)

Gavini, M.B.

1978-01-01

238 Pu, 239 Pu and 137 Cs in rain and dry fallout and 90 Sr in rain samples were measured at Woods Hole, Massachusetts, from June 1976 through December 1977. The dry fallout was estimated to be about 7.8% of the total deposition of 239 Pu and 137 Cs. 239 Pu/ 137 Cs ratios, almost constant at about 0.011 in rain or dry fallout, February through December 1977, suggested that fractionation between the refractory and volatile radionuclides is insignificant in stratospheric fallout. This supports the idea of regional homogeneity of radionuclide ratios in fallout. (Auth.)

Whole blood solubilization and discoloration before LSC of yttrium samples

International Nuclear Information System (INIS)

Lima, Marina F.; Leonardo, Lucio

2009-01-01

Liquid scintillation counting of whole blood and animal tissues samples could be severely impaired owing to quenching by their compounds. The objective of this previous study is preparing one protocol of 90 Y measurement to apply in biodistribution and dosimetry studies of radiopharmaceuticals labeled with this isotope and other beta emitters in in vivo and ex vivo samples. The first parameters considered to choose a method were: the largest blood sample per collection (80μl-90μl), attending the collection limit of less than 7.5% of total circulating blood volume for in vivo samples. Other parameters were the use of EDTA and cyclohexydine as solubilization and lytic agents, HNO 3 and H 2 O 2 as mineralizing agents and NH 4 OH as neutralization agent. One samples batch was tested in a water bath under the lower temperature to prevent the volume lose of the ionic phase. Other samples batch was mineralized over a hot-plate at 120 deg C following the currently largest sample amounts processing procedure in our laboratory by using HNO 3 and H 2 O 2 . Results show the contribution of the blood fragments as quenching in the region A ( 3 and/or NH 4 OH in the hot-plate digestion. As expected, the measurements in the three spectral regions show the proteins and colored fragments were completely removed by the hot-plate digestion. The rate between efficiency and 90 Sr- 90 Y concentration had not significant differences in the range between 120 Bq and 1200 Bq. (author)

Determination of proguanil and metabolites in small sample volumes of whole blood stored on filter paper by high-performance liquid chromatography.

Science.gov (United States)

Kolawole, J A; Taylor, R B; Moody, R R

1995-12-01

A method is reported for the determination of proguanil and its two metabolites cycloguanil and 4-chlorophenylbiguanide in whole blood and plasma samples obtained by thumbprick and stored dry on filter paper. The sample preparation involves liquid extraction from the filter paper and subsequent solid-phase extraction using C8 Bond-Elut cartridges. Separation and quantification is by a previously reported ion-pairing high-performance liquid chromatographic system with ODS Hypersil as stationary phase and an 50:50 acetonitrile-pH 2 phosphate buffer mobile phase containing 200 mM sodium dodecylsulphate as ion-pairing agent. The analytical characteristics of the method are reported. Representative concentrations are shown as a function of time from a human subject after ingestion of a single 200-mg dose of proguanil hydrochloride. Typical ranges of concentration detected by the proposed method in human subjects were proguanil 12-900 ng/ml, cycloguanil 16-44 ng/ml and 4-chlorophenylbiguanide 1.5-10 ng/ml in whole blood.

Astronaut Joseph Kerwin takes blood sample from Astronaut Charles Conrad

Science.gov (United States)

1973-01-01

Scientist-Astronaut Joseph P. Kerwin (right), Skylab 2 science pilot and a doctor of medicine, takes a blood sample from Astronaut Charles Conrad Jr., Sylab 2 commander, as seen in this reproduction taken from a color television transmission made by a TV camera aboard the Skylab 1 and 2 space station cluster in Earth orbit. The blood sampling was part of the Skylab Hematology and Immunology Experiment M110 series.

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

Science.gov (United States)

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

2013-09-01

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 (r(2)>0.99) and 27.4-20,000 ng/ml (r(2)>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. Copyright © 2013 Elsevier B.V. All rights reserved.

Physiological And Blood Biochemical Responses To Dried Live Yeast Plus Vitamin E As A Dietary Supplement To Bovine Baladi Calves Under Hot Summer Conditions

International Nuclear Information System (INIS)

ABDALLA, E.B.; EL-MASRY, K.A.; TEAMA, F.E.; EMARA, S.S.

2009-01-01

The experiment was designed to study the effect of supplemented dried live yeast (DLY) + vitamin E to the diet of growing calves under hot summer conditions in Egypt. Six bovine Baladi calves with 115 kg initial body weight and 8-10 months old were used during two periods. In the first period, the calves were offered the concentrated basal diet only for one month and considered as a control period. In the second period, the calves were fed the same basal diet which supplemented with 15 g dried live yeast (Saccharomyces cerevisiae) + 600 IU vitamin E (alpha- tocopherol) per calf daily for one month and considered as a treated period. Body weight was recorded at the beginning and the end of each period, and daily gain was calculated for each animal. Blood samples were collected from each animal at the end of each period to determine some blood biochemical parameters and T 3 and T 4 concentrations as well as some immunological indices.The results showed that supplementation of DLY + 600 IU vitamin E to the diet of calves reduced significantly (P 3 and T 4 levels and improved feed efficiency and daily gain. It is concluded that supplementation of growing calves with 15 g DLY + 600 IU vitamin E / calf / day under Egyptian hot summer conditions reduced the effect of heat stress as shown by a decline in RT and modified most blood constituents and thyroid function which leads to an improvement in growing calves

Human blood RNA stabilization in samples collected and transported for a large biobank

Science.gov (United States)

2012-01-01

Background The Norwegian Mother and Child Cohort Study (MoBa) is a nation-wide population-based pregnancy cohort initiated in 1999, comprising more than 108.000 pregnancies recruited between 1999 and 2008. In this study we evaluated the feasibility of integrating RNA analyses into existing MoBa protocols. We compared two different blood RNA collection tube systems – the PAXgene™ Blood RNA system and the Tempus™ Blood RNA system - and assessed the effects of suboptimal blood volumes in collection tubes and of transportation of blood samples by standard mail. Endpoints to characterize the samples were RNA quality and yield, and the RNA transcript stability of selected genes. Findings High-quality RNA could be extracted from blood samples stabilized with both PAXgene and Tempus tubes. The RNA yields obtained from the blood samples collected in Tempus tubes were consistently higher than from PAXgene tubes. Higher RNA yields were obtained from cord blood (3 – 4 times) compared to adult blood with both types of tubes. Transportation of samples by standard mail had moderate effects on RNA quality and RNA transcript stability; the overall RNA quality of the transported samples was high. Some unexplained changes in gene expression were noted, which seemed to correlate with suboptimal blood volumes collected in the tubes. Temperature variations during transportation may also be of some importance. Conclusions Our results strongly suggest that special collection tubes are necessary for RNA stabilization and they should be used for establishing new biobanks. We also show that the 50,000 samples collected in the MoBa biobank provide RNA of high quality and in sufficient amounts to allow gene expression analyses for studying the association of disease with altered patterns of gene expression. PMID:22988904

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

Science.gov (United States)

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

Reflections on Dry Eye Syndrome Treatment: Therapeutic Role of Blood Products

Directory of Open Access Journals (Sweden)

Victor J. Drew

2018-02-01

Full Text Available Dry eye syndrome (DES is a multifactorial, frequent, pathology characterized by deficient tear production or increased evaporation of tears and associated with ocular surface alteration and inflammation. It mostly affects, but not exclusively, older individuals and leads to varying degrees of discomfort and decreased quality of life. Although the typical treatments of DES rely on using artificial tears, polyunsaturated fatty acids, integrin antagonists, anti-inflammatory agents, or on performing punctal occlusion, recently, standardized blood-derived serum eye drops (SED are generating much interest as a new physiological treatment option. The scientific rationale in using SED for treating or releasing the symptoms of DES is thought to lie in its composition in multiple factors that resembles that of tears and contributes to the healing and protection of the ocular surface. This manuscript seeks to provide relevant background information on the management of DES, and on the increasing role that various types of SED or platelet lysates, from autologous or allogeneic origins, are playing in the improved therapeutic management of this pathology. The increasing role played by blood establishments in producing better-standardized SED is also addressed.

Sampling methods to the statistical control of the production of blood components.

Science.gov (United States)

Pereira, Paulo; Seghatchian, Jerard; Caldeira, Beatriz; Santos, Paula; Castro, Rosa; Fernandes, Teresa; Xavier, Sandra; de Sousa, Gracinda; de Almeida E Sousa, João Paulo

2017-12-01

The control of blood components specifications is a requirement generalized in Europe by the European Commission Directives and in the US by the AABB standards. The use of a statistical process control methodology is recommended in the related literature, including the EDQM guideline. The control reliability is dependent of the sampling. However, a correct sampling methodology seems not to be systematically applied. Commonly, the sampling is intended to comply uniquely with the 1% specification to the produced blood components. Nevertheless, on a purely statistical viewpoint, this model could be argued not to be related to a consistent sampling technique. This could be a severe limitation to detect abnormal patterns and to assure that the production has a non-significant probability of producing nonconforming components. This article discusses what is happening in blood establishments. Three statistical methodologies are proposed: simple random sampling, sampling based on the proportion of a finite population, and sampling based on the inspection level. The empirical results demonstrate that these models are practicable in blood establishments contributing to the robustness of sampling and related statistical process control decisions for the purpose they are suggested for. Copyright © 2017 Elsevier Ltd. All rights reserved.

Determination of trace elements and screening of metalloproteins in human blood and tissues

International Nuclear Information System (INIS)

Prohaska, K.

2003-02-01

Sequential and simultaneous atomic spectrometric detection was applied for the determination of metals in human whole blood, blood fractions and joint tissues. For this purpose ICP-OES (inductively coupled plasma - optical emission spectrometry) and GFAAS (graphite furnace - atomic absorption spectrometry) were optimized. The nebulizing technique of the ICP-OES causes a high consumption of the sample (2.4 mL /min uptake rate). Using the simultaneous modus of measurement it is possible to determine up to 20 elements in the samples of interest. Using GFAAS the elements can be detected sequentially only, the method is more time consuming in comparison. For the direct sample injection into the graphite tube only 20 aeL are needed. Since 1 mL sample has to be filled in the sample vessel of the autosampler, up to six elements can be determined in this volume. The most important advantage of the simultaneous multi-element detection is the low sample consumption, which is essential for analysis of biological samples. Normally only small amounts of human blood or tissues can be collected, and the concentrations of analytes are usually very low. Therefore in many cases a sample preparation is of advantage, which enables a pre-concentration of the analyte. The sample digestion was optimized with respect to the possible pre-concentration of the analytes. Ashing of the freeze-dried blood enabled a six fold higher concentration in the measuring solution compared with wet digestion of blood. For the ICP-OES sample introduction system two different nebulizers were tested in the complex matrix of the digested blood samples. A Meinhard and the microconcentric nebulizer were compared according to their analytical performance. The matrix of the samples caused LODs three to five times higher than in the aqueous standards. The application of the Meinhard nebulizer enabled sufficiently low LODs in the matrix for some elements of interest (Ca, Cu, Fe, Mg, P, S, Zn in freeze-dried blood

Time and temperature dependent analytical stability of dry-collected Evalyn HPV self-sampling brush for cervical cancer screening

DEFF Research Database (Denmark)

Ejegod, Ditte Møller; Pedersen, Helle; Alzua, Garazi Peña

2018-01-01

As a new initiative, HPV self-sampling to non-attenders using the dry Evalyn self-sampling brush is offered in the Capital Region of Denmark. The use of a dry brush is largely uncharted territory in terms of analytical stability. In this study we aim to provide evidence on the analytical quality...

Radioimmunoassay of trypsin in dried blood importance for the neonatal detection of cystic fibrosis

International Nuclear Information System (INIS)

Travert, G.; Mustin, C.; Fernandez, Y.

1981-01-01

The demonstration of very high levels of immunoreactive trypsin in the blood of newborn infants with cystic fibrosis has provided a new way of detecting the disease soon after birth. A radioimmunoassay of trypsin in the eluate of blood dried on filter paper has now been developed. The sensitivity and accuracy of the method, as well as the good correlation observed between the values obtained and those of the conventional plasma assay, indicate that it is reliable and well adapted to the newborn. The new assay can easily be inserted into the present system of neonatal disease detection. A preliminary assessment of more than 5000 tests enables the authors to report an early diagnosis of proven cystic fibrosis and to discuss an essential aspect of mass-detection methods: the indicence of false-positive results [fr

Ambulatory blood pressure and blood lipids in a multiethnic sample of healthy adults.

Science.gov (United States)

James, Gary D; Van Berge-Landry, Helene M; Morrison, Lynn A; Reza, Angela M; Nicolaisen, Nicola M; Bindon, James R; Brown, Daniel E

2013-01-01

Elevated blood pressure (BP), elevated serum cholesterol, and aberrant lipoprotein fractions (low levels of high-density lipoprotein (HDL) and high levels of low-density lipoprotein fractions and triglycerides) have all been used as measures that assess the "metabolic syndrome" and more recently in indexes of allostatic load, which are designed to assess the degree of integrated metabolic pathology. While there are ample data regarding the interrelationships of these measures in various pathophysiological settings, there are limited data regarding the interrelationship of ambulatory BP (ABP) and blood lipids in healthy subjects. The present study evaluates ABP-blood lipid relationships in a multiethnic sample of healthy adults. The subjects were 37 men (age = 40.9 ± 10.7 years) and 42 women (age = 35.8 ± 10.4 years) who were employed as hotel workers in Hawaii. Each wore an ABP monitor for one midweek workday and had pressures averaged in three daily microenvironments (work, home, and during sleep). They also had fasting blood samples taken for lipid profiling. Multivariate analysis of covariance shows that there was a strong inverse relationship between HDL and both systolic (P act as a group in healthy adults but that higher HDL is associated with lower BP. This latter finding is consistent with research that shows that HDL promotes vasodilation via its effect on endothelial nitric oxide synthase. Copyright © 2013 Wiley Periodicals, Inc.

Identification of clinical biomarkers for pre-analytical quality control of blood samples.

Science.gov (United States)

Kang, Hyun Ju; Jeon, Soon Young; Park, Jae-Sun; Yun, Ji Young; Kil, Han Na; Hong, Won Kyung; Lee, Mee-Hee; Kim, Jun-Woo; Jeon, Jae-Pil; Han, Bok Ghee

2013-04-01

Pre-analytical conditions are key factors in maintaining the high quality of biospecimens. They are necessary for accurate reproducibility of experiments in the field of biomarker discovery as well as achieving optimal specificity of laboratory tests for clinical diagnosis. In research at the National Biobank of Korea, we evaluated the impact of pre-analytical conditions on the stability of biobanked blood samples by measuring biochemical analytes commonly used in clinical laboratory tests. We measured 10 routine laboratory analytes in serum and plasma samples from healthy donors (n = 50) with a chemistry autoanalyzer (Hitachi 7600-110). The analyte measurements were made at different time courses based on delay of blood fractionation, freezing delay of fractionated serum and plasma samples, and at different cycles (0, 1, 3, 6, 9) of freeze-thawing. Statistically significant changes from the reference sample mean were determined using the repeated-measures ANOVA and the significant change limit (SCL). The serum levels of GGT and LDH were changed significantly depending on both the time interval between blood collection and fractionation and the time interval between fractionation and freezing of serum and plasma samples. The glucose level was most sensitive only to the elapsed time between blood collection and centrifugation for blood fractionation. Based on these findings, a simple formula (glucose decrease by 1.387 mg/dL per hour) was derived to estimate the length of time delay after blood collection. In addition, AST, BUN, GGT, and LDH showed sensitive responses to repeated freeze-thaw cycles of serum and plasma samples. These results suggest that GGT and LDH measurements can be used as quality control markers for certain pre-analytical conditions (eg, delayed processing or repeated freeze-thawing) of blood samples which are either directly used in the laboratory tests or stored for future research in the biobank.

GEMAS: Colours of dry and moist agricultural soil samples of Europe

Science.gov (United States)

Klug, Martin; Fabian, Karl; Reimann, Clemens

2016-04-01

High resolution HDR colour images of all Ap samples from the GEMAS survey were acquired using a GeoTek Linescan camera. Three measurements of dry and wet samples with increasing exposure time and increasing illumination settings produced a set of colour images at 50μm resolution. Automated image processing was used to calibrate the six images per sample with respect to the synchronously measured X-Rite colorchecker chart. The calibrated images were then fit to Munsell soil colours that were measured in the same way. The results provide overview maps of dry and moist European soil colours. Because colour is closely linked to iron mineralogy, carbonate, silicate and organic carbon content the results can be correlated to magnetic, mineralogical, and geochemical properties. In combination with the full GEMAS chemical and physical measurements, this yields a valuable data set for calibration and interpretation of visible satellite colour data with respect to chemical composition and geological background, soil moisture, and soil degradation. This data set will help to develop new methods for world-wide characterization and monitoring of agricultural soils which is essential for quantifying geologic and human impact on the critical zone environment. It furthermore enables the scientific community and governmental authorities to monitor consequences of climatic change, to plan and administrate economic and ecological land use, and to use the data set for forensic applications.

Are They Bloody Guilty? Blood Doping with Simulated Samples

Science.gov (United States)

Stuart, Parker E.; Lees, Kelsey D.; Milanick, Mark A.

2014-01-01

In this practice-based lab, students are provided with four Olympic athlete profiles and simulated blood and urine samples to test for illegal substances and blood-doping practices. Throughout the course of the lab, students design and conduct a testing procedure and use their results to determine which athletes won their medals fairly. All of the…

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

Science.gov (United States)

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 blood and the overall method was accurate and precise for the determination of DBS isotopic enrichments. Copyright © 2016 Elsevier B.V. All rights reserved.

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

Science.gov (United States)

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 Milk yield, FCM, fat yield, lactose yield and total solid yield was significantly higher ( P < 0.05) in cooled group of Murrah buffaloes.

A novel tandem mass spectrometry method for first-line screening of mainly beta-thalassemia from dried blood spots.

Science.gov (United States)

Yu, Chaowen; Huang, Shuodan; Wang, Ming; Zhang, Juan; Liu, Hao; Yuan, Zhaojian; Wang, Xingbin; He, Xiaoyan; Wang, Jie; Zou, Lin

2017-02-10

Traditional methods for thalassemia screening are time-consuming and easily affected by cell hemolysis or hemoglobin degradation in stored blood samples. Tandem mass spectrometry (MS/MS) proved to be an effective technology for sickle cell disorders (SCD) screening. Here, we developed a novel MS/MS method for β-thalassemia screening from dried blood spots (DBS). Stable isotopic-labeled peptides were used as internal standards for quantification and calculation of the α:β-globin ratios. We used the α:β-globin ratio cutoffs to differentiate between normal individuals and patients with thalassemia. About 781 patients and 300 normal individuals were analyzed. The α:β-globin ratios showed significant difference between normal and β-thalassemia patients (Pthalassemia mutation. In the parallel study, all cases screened for suspected thalassemia from six hundred DBS samples by using this MS/MS method were successfully confirmed by genotyping. The intra-assay and inter-assay CVs of the ratios ranged from 2.4% to 3.9% and 4.7% to 7.1%, and there was no significant sample carryover or matrix effect for this MS/MS method. Combined with SCD screening, this MS/MS method could be used as a first-line screening assay for both structural and expression abnormalities of human hemoglobin. Traditional methods for thalassemia screening were depending on the structural integrity of tetramers and could be affected by hemolysis and degradation of whole blood samples, especially when stored. We used proteospecific peptides produced by the tryptic digestion of each globin to evaluate the production ratio between α- and β-globin chains, which turned out to be quite stable even when stored for more than two months. Though most of the peptides were specific to α-globin or β-globin, we only chose four most informative peptides and its stable isotopic-labeled peptides as internal standards for analysis, which could obtain a high accuracy. Currently, we are the first to address the

EFFECT OF ADDING THE INTERNAL STANDARD TO BLOOD SAMPLES, PRIOR TO THE PREPARATION OF BLOOD SPOTS FOR ACYLCARNITINE ANALYSIS

OpenAIRE

Osorio, José Henry; Pourfarzam, Morteza

2010-01-01

Background: some general factors can influence when determining acylcarnitines through tandem mass spectrometry. Objective: to study the effect of adding the internal standard to blood samples before the preparation of filter paper cards compared with the addition of internal standard after having the filter paper cards prepared for determining acylcarnitines in blood for tandem mass spectrometry. Methodology: two groups of blood samples were prepared: group one without adding internal standa...

A method for estimating radioactive cesium concentrations in cattle blood using urine samples.

Science.gov (United States)

Sato, Itaru; Yamagishi, Ryoma; Sasaki, Jun; Satoh, Hiroshi; Miura, Kiyoshi; Kikuchi, Kaoru; Otani, Kumiko; Okada, Keiji

2017-12-01

In the region contaminated by the Fukushima nuclear accident, radioactive contamination of live cattle should be checked before slaughter. In this study, we establish a precise method for estimating radioactive cesium concentrations in cattle blood using urine samples. Blood and urine samples were collected from a total of 71 cattle on two farms in the 'difficult-to-return zone'. Urine 137 Cs, specific gravity, electrical conductivity, pH, sodium, potassium, calcium, and creatinine were measured and various estimation methods for blood 137 Cs were tested. The average error rate of the estimation was 54.2% without correction. Correcting for urine creatinine, specific gravity, electrical conductivity, or potassium improved the precision of the estimation. Correcting for specific gravity using the following formula gave the most precise estimate (average error rate = 16.9%): [blood 137 Cs] = [urinary 137 Cs]/([specific gravity] - 1)/329. Urine samples are faster to measure than blood samples because urine can be obtained in larger quantities and has a higher 137 Cs concentration than blood. These advantages of urine and the estimation precision demonstrated in our study, indicate that estimation of blood 137 Cs using urine samples is a practical means of monitoring radioactive contamination in live cattle. © 2017 Japanese Society of Animal Science.

Use of Capillary Blood Samples Leads to Higher Parasitemia Estimates and Higher Diagnostic Sensitivity of Microscopic and Molecular Diagnostics of Malaria than Venous Blood Samples.

Science.gov (United States)

Mischlinger, Johannes; Pitzinger, Paul; Veletzky, Luzia; Groger, Mirjam; Zoleko-Manego, Rella; Adegnika, Ayola A; Agnandji, Selidji T; Lell, Bertrand; Kremsner, Peter G; Tannich, Egbert; Mombo-Ngoma, Ghyslain; Mordmüller, Benjamin; Ramharter, Michael

2018-05-25

Diagnosis of malaria is usually based on samples of peripheral blood. However, it is unclear whether capillary (CAP) or venous (VEN) blood samples provide better diagnostic performance. Quantitative differences of parasitemia between CAP and VEN blood and diagnostic performance characteristics were investigated. Patients were recruited between September 2015 and February 2016 in Gabon. Light microscopy and qPCR quantified parasitemia of paired CAP and VEN samples, whose preparation followed the exact same methodology. CAP and VEN performance characteristics using microscopy were evaluated against a qPCR gold-standard. Microscopy revealed a median (IQR) parasites/L of 495 (853,243) in CAP and 429 (524,074) in VEN samples manifesting in a +16.6% (p=0.04) higher CAPparasitemia compared with VENparasitemia. Concordantly, qPCR demonstrated that -0.278 (p=0.006) cycles were required for signal detection in CAP samples. CAPsensitivity of microscopy relative to the gold-standard was 81.5% (77.485.6%) versus VENsensitivity of 73.4% (68.878.1%), while CAPspecificity and VENspecificity were 91%. CAPsensitivity and VENsensitivity dropped to 63.3% and 45.9%, respectively for a sub-population of low-level parasitemias while specificities were 92%. CAP sampling leads to higher parasitemias compared to VEN sampling and improves diagnostic sensitivity. These findings may have important implications for routine diagnostics, research and elimination campaigns of malaria.

Comparative research of effectiveness of cellulose and fiberglass porous membrane carriers for bio sampling in veterinary and food industry monitoring

Science.gov (United States)

Gusev, Alexander; Vasyukova, Inna; Zakharova, Olga; Altabaeva, Yuliya; Saushkin, Nikolai; Samsonova, Jeanne; Kondakov, Sergey; Osipov, Alexander; Snegin, Eduard

2017-11-01

The aim of proposed research is to study the applicability of fiberglass porous membrane materials in a new strip format for dried blood storage in food industry monitoring. A comparative analysis of cellulosic and fiberglass porous membrane materials was carried out to obtain dried samples of serum or blood and the possibility of further species-specific analysis. Blood samples of Sus scrofa were used to study the comparative effectiveness of cellulose and fiberglass porous membrane carriers for long-term biomaterial storage allowing for further DNA detection by real-time polymerase chain reaction (PCR) method. Scanning electron microscopy of various membranes - native and with blood samples - indicate a fundamental difference in the form of dried samples. Membranes based on cellulosic materials sorb the components of the biological fluid on the surface of the fibers of their structure, partially penetrating the cellulose fibers, while in the case of glass fiber membranes the components of the biological fluid dry out as films in the pores of the membrane between the structural filaments. This fundamental difference in the retention mechanisms affects the rate of dissolution of the components of dry samples and contributes to an increase in the efficiency of the desorption process of the sample before subsequent analysis. Detecting of pig DNA in every analyzed sample under the performed Real-time PCR as well as good state of the biomaterial preservation on the glass fiber membranes was clearly demonstrated. Good biomaterials preservation has been revealed on the test cards for 4 days as well as for 1 hour.

Plasma and breast milk pharmacokinetics of emtricitabine, tenofovir and lamivudine using dried blood and breast milk spots in nursing African mother–infant pairs

Science.gov (United States)

Waitt, Catriona; Olagunju, Adeniyi; Nakalema, Shadia; Kyohaire, Isabella; Owen, Andrew; Lamorde, Mohammed; Khoo, Saye

2018-01-01

Abstract Background Breast milk transfer of first-line ART from mother to infant is not fully understood. Objectives To determine the concentrations of lamivudine, emtricitabine and tenofovir in maternal blood, breast milk and infant blood from breastfeeding mother–infant pairs. Methods Intensive pharmacokinetic sampling of maternal dried blood spots (DBS), dried breast milk spots (DBMS) and infant DBS from 30 Ugandan and 29 Nigerian mothers receiving first-line ART and their infants was conducted. DBS and DBMS were collected pre-dose and at 5–6 timepoints up to 12 h following observed dosing. Infant DBS were sampled twice during this period. Lamivudine, emtricitabine and tenofovir were quantified using LC-MS/MS, with non-compartmental analysis to calculate key pharmacokinetic parameters. Results Peak concentrations in breast milk from women taking lamivudine and emtricitabine occurred later than in plasma (4–8 h compared with 2 h for lamivudine and 2–4 h for emtricitabine). Consequently, the milk-to-plasma (M:P) ratio of lamivudine taken once daily was 0.95 (0.82–1.15) for AUC0–12, whereas for AUC12–20 this was 3.04 (2.87–4.16). Lamivudine was detectable in 36% (14/39) of infants [median 17.7 (16.3–22.7) ng/mL]. For 200 mg of emtricitabine once daily, the median M:P ratio was 3.01 (2.06–3.38). Three infants (19%) had measurable emtricitabine [median 18.5 (17.6–20.8) ng/mL]. For 300 mg of tenofovir once daily, the median M:P ratio was 0.015 (0–0.03) and no infant had measurable tenofovir concentrations. Conclusions Emtricitabine and lamivudine accumulate in breast milk and were detected in breastfeeding infants. In contrast, tenofovir penetrates the breast milk to a small degree, but is undetectable in breastfeeding infants. PMID:29309634

Adaptive control of theophylline therapy: importance of blood sampling times.

Science.gov (United States)

D'Argenio, D Z; Khakmahd, K

1983-10-01

A two-observation protocol for estimating theophylline clearance during a constant-rate intravenous infusion is used to examine the importance of blood sampling schedules with regard to the information content of resulting concentration data. Guided by a theory for calculating maximally informative sample times, population simulations are used to assess the effect of specific sampling times on the precision of resulting clearance estimates and subsequent predictions of theophylline plasma concentrations. The simulations incorporated noise terms for intersubject variability, dosing errors, sample collection errors, and assay error. Clearance was estimated using Chiou's method, least squares, and a Bayesian estimation procedure. The results of these simulations suggest that clinically significant estimation and prediction errors may result when using the above two-point protocol for estimating theophylline clearance if the time separating the two blood samples is less than one population mean elimination half-life.

A Comparative Study of the use of Dried Blood on Filter Papers and ...

African Journals Online (AJOL)

A study was conducted to compare the suitability of blood collected on filter papers with corresponding fluid serum samples in serodiagnosis of bovine anaplasmosis using DOT-ELISA, RCAT and Western imunobtotting. Serum was obtained from blood collected by jugular venipuncture of288 heads of cattle and stored at ...

A simple method for regional cerebral blood flow measurement by one-point arterial blood sampling and 123I-IMP microsphere model (part 2). A study of time correction of one-point blood sample count

International Nuclear Information System (INIS)

Masuda, Yasuhiko; Makino, Kenichi; Gotoh, Satoshi

1999-01-01

In our previous paper regarding determination of the regional cerebral blood flow (rCBF) using the 123 I-IMP microsphere model, we reported that the accuracy of determination of the integrated value of the input function from one-point arterial blood sampling can be increased by performing correction using the 5 min: 29 min ratio for the whole-brain count. However, failure to carry out the arterial blood collection at exactly 5 minutes after 123 I-IMP injection causes errors with this method, and there is thus a time limitation. We have now revised out method so that the one-point arterial blood sampling can be performed at any time during the interval between 5 minutes and 20 minutes after 123 I-IMP injection, with addition of a correction step for the sampling time. This revised method permits more accurate estimation of the integral of the input functions. This method was then applied to 174 experimental subjects: one-point blood samples collected at random times between 5 and 20 minutes, and the estimated values for the continuous arterial octanol extraction count (COC) were determined. The mean error rate between the COC and the actual measured continuous arterial octanol extraction count (OC) was 3.6%, and the standard deviation was 12.7%. Accordingly, in 70% of the cases, the rCBF was able to be estimated within an error rate of 13%, while estimation was possible in 95% of the cases within an error rate of 25%. This improved method is a simple technique for determination of the rCBF by 123 I-IMP microsphere model and one-point arterial blood sampling which no longer shows a time limitation and does not require any octanol extraction step. (author)

ABO and D typing and alloantibody screening in marrow samples: relevance to intraosseous blood transfusion.

Science.gov (United States)

Bäckman, Sari; Ångerman-Haasmaa, Susanne; Jousi, Milla; Siitonen, Sanna; Salmela, Katja

2018-03-01

Blood transfusion through the intraosseous route is gaining popularity in emergency medicine. Pretransfusion peripheral blood (PB) samples are usually not available in these patients, leading to discrepancies in blood group typing and a possible delay in transferring to group-specific blood products. The aim of this study was to assess the feasibility of ABO and D typing and red blood cell alloantibody screening in marrow (BM) samples. Direct and reverse ABO typing, D typing, and a two-cell alloantibody screen were performed in EDTA-anticoagulated BM samples with standard manual column agglutination techniques. EDTA-anticoagulated PB samples were used as controls. The mean age of the study subjects (n = 71) was 47 years (range, 1-82 years). All ABO groups and both D+ and D- types were represented. In all subjects, concordant results were observed for all analyses in BM and PB samples. In 15 (21%) of the samples, a discrepancy of one reaction strength step (1+) was observed in at least one of the analyses (Cohen's weighted κ = 0.993); this did not affect interpretation of the results. Blood group typing and alloantibody screening are feasible in BM samples, providing proof-of-concept that intraosseous samples for blood group serologic analyses can be collected from emergency patients before intraosseous blood transfusion. This will enable a timely transfer to group-specific blood products and enable conservation of the valuable universal-donor blood products. © 2018 AABB.

Elemental distribution and sample integrity comparison of freeze-dried and frozen-hydrated biological tissue samples with nuclear microprobe

Energy Technology Data Exchange (ETDEWEB)

Vavpetič, P., E-mail: primoz.vavpetic@ijs.si [Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); Vogel-Mikuš, K. [Biotechnical Faculty, Department of Biology, University of Ljubljana, Jamnikarjeva 101, SI-1000 Ljubljana (Slovenia); Jeromel, L. [Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); Ogrinc Potočnik, N. [Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); FOM-Institute AMOLF, Science Park 104, 1098 XG Amsterdam (Netherlands); Pongrac, P. [Biotechnical Faculty, Department of Biology, University of Ljubljana, Jamnikarjeva 101, SI-1000 Ljubljana (Slovenia); Department of Plant Physiology, University of Bayreuth, Universitätstr. 30, 95447 Bayreuth (Germany); Drobne, D.; Pipan Tkalec, Ž.; Novak, S.; Kos, M.; Koren, Š.; Regvar, M. [Biotechnical Faculty, Department of Biology, University of Ljubljana, Jamnikarjeva 101, SI-1000 Ljubljana (Slovenia); Pelicon, P. [Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia)

2015-04-01

The analysis of biological samples in frozen-hydrated state with micro-PIXE technique at Jožef Stefan Institute (JSI) nuclear microprobe has matured to a point that enables us to measure and examine frozen tissue samples routinely as a standard research method. Cryotome-cut slice of frozen-hydrated biological sample is mounted between two thin foils and positioned on the sample holder. The temperature of the cold stage in the measuring chamber is kept below 130 K throughout the insertion of the samples and the proton beam exposure. Matrix composition of frozen-hydrated tissue is consisted mostly of ice. Sample deterioration during proton beam exposure is monitored during the experiment, as both Elastic Backscattering Spectrometry (EBS) and Scanning Transmission Ion Microscopy (STIM) in on–off axis geometry are recorded together with the events in two PIXE detectors and backscattered ions from the chopper in a single list-mode file. The aim of this experiment was to determine differences and similarities between two kinds of biological sample preparation techniques for micro-PIXE analysis, namely freeze-drying and frozen-hydrated sample preparation in order to evaluate the improvements in the elemental localisation of the latter technique if any. In the presented work, a standard micro-PIXE configuration for tissue mapping at JSI was used with five detection systems operating in parallel, with proton beam cross section of 1.0 × 1.0 μm{sup 2} and a beam current of 100 pA. The comparison of the resulting elemental distributions measured at the biological tissue prepared in the frozen-hydrated and in the freeze-dried state revealed differences in elemental distribution of particular elements at the cellular level due to the morphology alteration in particular tissue compartments induced either by water removal in the lyophilisation process or by unsatisfactory preparation of samples for cutting and mounting during the shock-freezing phase of sample preparation.

Determination of pistachio drying behaviour and conditions in a solar drying system

Energy Technology Data Exchange (ETDEWEB)

Midilli, Adnan [Karadeniz Technical Univ., Mechanical Engineering Dept., Trabzon (Turkey)

2001-07-01

The main object of this study is to determine the drying behaviour and conditions of shelled and unshelled pistachio samples using both solar assisted and open sun drying. For each drying experiment, 100 g each of unshelled and shelled pistachio were used. The least-squares method was applied to find the drying curve equation of pistachio. During the experiments, shelf temperatures, weight loss of pistachio, moisture content of air, and distribution of solar radiation were measured; and presented depending on the drying time. Also, the mass shrinkage ratios of shelled and unshelled pistachio samples were determined, and the experimental uncertainty ratio was calculated as 15-16.5 per cent based on the experimental results. It was deduced that the shelled and unshelled pistachio samples in the solar assisted forced convection dryer were perfectly dried at temperatures of 50{+-}10degC in the time period of 6 h. Whereas, the samples in the open sun drying were not sufficiently dried at temperatures of 28{+-}4degC in the same time period. Hence, it is suggested that the pistachio samples with approximately 29.0 per cent of moisture are dried in the solar assisted convection dryer at 50{+-}10degC of temperature in the time period of approximately 6 h in order to protect from the negative climatic and environmental effects. However, it is not desirable to dry the pistachio samples in the open sun because of greater drying time, dirt, dust and harmful insects. (Author)

Determination of drying characteristics and quality properties of eggplant in different drying conditions

Directory of Open Access Journals (Sweden)

Gözde Bayraktaroglu Urun

2015-12-01

Full Text Available Drying is the most traditional process used for preserving eggplant a long time. The aim of this study was to determining drying characteristics and quality properties of eggplant dried by sun drying, hot air convective drying and infrared assisted convective drying. Convective drying and infrared assisted convective were carried out in a convective dryer at three different temperatures(40°, 50°, 60°C and air velocity at 5 m/s.The increasing of temperatures during the drying of eggplant led to a significant reduction of the drying time. However loss of nutrition was observed in eggplant samples dried at higher temperature.The biggest change in colour parameters was observed in samples dried with sun drying.So it was thought that sun drying had a negative effect on quality properties of eggplant samples.

Randomized Comparison of Two Vaginal Self-Sampling Methods for Human Papillomavirus Detection: Dry Swab versus FTA Cartridge

OpenAIRE

Catarino, Rosa; Vassilakos, Pierre; Bilancioni, Aline; Vanden Eynde, Mathieu; Meyer-Hamme, Ulrike; Menoud, Pierre-Alain; Guerry, Fr?d?ric; Petignat, Patrick

2015-01-01

Background Human papillomavirus (HPV) self-sampling (self-HPV) is valuable in cervical cancer screening. HPV testing is usually performed on physician-collected cervical smears stored in liquid-based medium. Dry filters and swabs are an alternative. We evaluated the adequacy of self-HPV using two dry storage and transport devices, the FTA cartridge and swab. Methods A total of 130 women performed two consecutive self-HPV samples. Randomization determined which of the two tests was performed f...

Single injection 51Cr EDTA plasma clearance determination in children using capillary blood samples

International Nuclear Information System (INIS)

Broechner-Mortensen, J.; Christoffersen, J.

1977-01-01

The reliability of a determination of the total 51 Cr EDTA plasma clearance (e) (and with it the glomerular filtration rate), by a simplified single injection method (injected dose: 4.5 μCi per kg b.w.) using capillary blood samples (0.2 ml), was investigated in twenty children. Clearance values determined from capillary blood samples did not differ significantly from those measured simultaneously from venous blood samples, the mean ratio+-SD being 1.02+-0.06(n = 10). The reproducibility (total day-to-day variation) of E determined from capillary blood samples was 6.7% in children with decreased renal function (n = 3) and 6.9% in children with normal renal function (n = 7). The present data indicate that the use of capillary blood samples is an accurate and very precise approach for determination of E in children. (Auth.)

Development of a novel single tube nested PCR for enhanced detection of cytomegalovirus DNA from dried blood spots.

Science.gov (United States)

Atkinson, C; Emery, V C; Griffiths, P D

2014-02-01

Newborn screening for congenital cytomegalovirus (CCMV) using dried blood spots (DBS) has been proposed because many developed countries have DBS screening programmes in place for other diseases. The aim of this study was to develop a rapid, single tube nested polymerase chain reaction (PCR) method for enhanced detection of CMV from DBS compared to existing (single target) real time PCRs. The new method was compared with existing real time PCRs for sensitivity and specificity. Overall sensitivity of the single target PCR assays in both asymptomatic and symptomatic infants with laboratory confirmed congenital CMV was 69% (CMV PCR or culture positive before day 21 of life). In contrast, the single tube nested assay had an increased sensitivity of 81% with100% specificity. Overall the assay detected CMV from a DBS equivalent to an original blood sample which contained 500IU/ml. In conclusion this single tube nested methodology allows simultaneous amplification and detection of CMV DNA in 1.5h removing the associated contamination risk of a two step nested PCR. Owing to its increased sensitivity, it has the potential to be used as a screening assay and ultimately allow early identification and intervention for children with congenital CMV. Copyright © 2013 Elsevier B.V. All rights reserved.

A prospective clinical trial to compare the performance of dried blood spots prenatal screening for Down's syndrome with conventional non-invasive testing technology.

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Hu, Huiying; Jiang, Yulin; Zhang, Minghui; Liu, Shanying; Hao, Na; Zhou, Jing; Liu, Juntao; Zhang, Xiaojin; Ma, Liangkun

2017-03-01

To evaluate, side by side, the efficiency of dried blood spots (DBSs) against serum screening for Down's syndrome, and then, to construct a two-tier strategy by topping up the fetal cell-free DNA (cfDNA) secondary screening over the high-risk women marked by the primary blood testing to build a practical screening tactic to identify fetal Down's syndrome. One thousand eight hundred and thirty-seven low-risk Chinese women, with singleton pregnancy, were enrolled for the study. Alpha-fetoprotein and free beta human chorionic gonadotropin were measured for the serum as well as for the parallel DBS samples. Partial high-risk pregnant women identified by primary blood testing (n = 38) were also subject to the secondary cfDNA screening. Diagnostic amniocentesis was utilized to confirm the screening results. The true positive rate for Down's syndrome detection was 100% for both blood screening methods; however, the false-positive rate was 3.0% for DBS and 4.0% for serum screening, respectively. DBS correlated well with serum screening on Down's syndrome detection. Three out of 38 primary high-risk women displayed chromosomal abnormalities by cfDNA analysis, which were confirmed by amniocentesis. Either the true detection rate or the false-positive rate for Down's syndrome between DBS and the serum test is comparable. In addition, blood primary screening aligned with secondary cfDNA analysis, a "before and after" two-tier screening strategy, can massively decrease the false-positive rate, which, then, dramatically reduces the demand for invasive diagnostic operation. Impact statement Children born with Down's syndrome display a wide range of mental and physical disability. Currently, there is no effective treatment to ease the burden and anxiety of the Down's syndrome family and the surrounding society. This study is to evaluate the efficiency of dried blood spots against serum screening for Down's syndrome and to construct a two-tier strategy by topping up the fetal

A content validated questionnaire for assessment of self reported venous blood sampling practices.

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Bölenius, Karin; Brulin, Christine; Grankvist, Kjell; Lindkvist, Marie; Söderberg, Johan

2012-01-19

Venous blood sampling is a common procedure in health care. It is strictly regulated by national and international guidelines. Deviations from guidelines due to human mistakes can cause patient harm. Validated questionnaires for health care personnel can be used to assess preventable "near misses"--i.e. potential errors and nonconformities during venous blood sampling practices that could transform into adverse events. However, no validated questionnaire that assesses nonconformities in venous blood sampling has previously been presented. The aim was to test a recently developed questionnaire in self reported venous blood sampling practices for validity and reliability. We developed a questionnaire to assess deviations from best practices during venous blood sampling. The questionnaire contained questions about patient identification, test request management, test tube labeling, test tube handling, information search procedures and frequencies of error reporting. For content validity, the questionnaire was confirmed by experts on questionnaires and venous blood sampling. For reliability, test-retest statistics were used on the questionnaire answered twice. The final venous blood sampling questionnaire included 19 questions out of which 9 had in total 34 underlying items. It was found to have content validity. The test-retest analysis demonstrated that the items were generally stable. In total, 82% of the items fulfilled the reliability acceptance criteria. The questionnaire could be used for assessment of "near miss" practices that could jeopardize patient safety and gives several benefits instead of assessing rare adverse events only. The higher frequencies of "near miss" practices allows for quantitative analysis of the effect of corrective interventions and to benchmark preanalytical quality not only at the laboratory/hospital level but also at the health care unit/hospital ward.

Detection of cytomegalovirus DNA in dried blood spots of Minnesota infants who do not pass newborn hearing screening.

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Choi, K Yeon; Schimmenti, Lisa A; Jurek, Anne M; Sharon, Bazak; Daly, Kathy; Khan, Cindy; McCann, Mark; Schleiss, Mark R

2009-12-01

Up to 15% of infants with asymptomatic congenital cytomegalovirus (CMV) infection will experience some degree of sensorineural hearing loss. Many infants who fail newborn hearing screening (NHS) are likely to have congenital CMV infection, but may escape definitive virologic identification because diagnostic evaluation may not commence until several weeks or months of age, making differentiation between congenital and postnatal CMV infection difficult. Early diagnosis linking virologic identification of congenital CMV infection to infants failing NHS may improve diagnostic precision and enhance opportunities for therapeutic intervention. The goal of this study was to compare newborn dried blood spots from Minnesota infants who had failed NHS, and were designated for referral, with control infants who passed NHS, for the presence of CMV DNA by real-time PCR, using hybridization probes for the CMV gene UL54. Of 479 infants with a failed NHS (bilateral failure), 13 had CMV DNA present in the blood spot (2.7%). This compared with only 2/479 positive results from a control group of infants who passed the NHS (0.4%; P = 0.007, Fisher exact test). Comparisons of the glycoprotein B (gB) genotype as well as direct DNA sequencing of selected positives revealed that PCR positive samples represented unique clinical isolates. The mean viral load among the 15 positive samples was 1.6 x 10(3) genomes/microgram of total DNA. Newborn bloodspot CMV screening by real-time PCR may be a useful and rapid adjunct to functional NHS and may enable more rapid etiologic diagnosis of sensorineural hearing loss in newborns.

Long-term facial artery catheter implantation for serial arterial blood sampling and invasive arterial blood pressure measurement in horses.

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Dias, Deborah Penteado Martins; Teixeira, Luisa Gouvêa; Canola, Paulo Aléscio; Albernaz, Raquel Mincarelli; Marques, José Antônio; Neto, José Corrêa de Lacerda

2012-06-01

The purpose of this investigation was to evaluate surgical catheter implantation in the facial artery of horses and the long-term maintenance of such arteries using heparin and ascorbic acid as filling solution. Nine horses were implanted with a polyurethane catheter. The catheters were flushed with a heparin/ascorbic acid solution every 8h and remained patent for 25 days. Arterial blood samples were collected twice a day, and one exercise test that included serial blood samples and arterial pressure recordings was performed on a treadmill. Polyurethane catheters surgically implanted in the facial artery can be kept patent by filling with a heparin/ascorbic acid solution and provide convenient invasive arterial access in horses which is suitable for use for serial blood sampling and blood pressure recordings, even during exercise on treadmill. Copyright © 2011 Elsevier Ltd. All rights reserved.

Relative Abundance of Proteins in Blood Plasma Samples from Patients with Chronic Cerebral Ischemia.

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Kaysheva, Anna L; Kopylov, Artur T; Ponomarenko, Elena A; Kiseleva, Olga I; Teryaeva, Nadezhda B; Potapov, Alexander A; Izotov, Alexander А; Morozov, Sergei G; Kudryavtseva, Valeria Yu; Archakov, Alexander I

2018-03-01

A comparative protein profile analysis of 17 blood plasma samples from patients with ischemia and 20 samples from healthy volunteers was carried out using ultra-high resolution mass spectrometry. The analysis of measurements was performed using the proteomics search engine OMSSA. Normalized spectrum abundance factor (NSAF) in the biological samples was assessed using SearchGUI. The findings of mass spectrometry analysis of the protein composition of blood plasma samples demonstrate that the depleted samples are quite similar in protein composition and relative abundance of proteins. By comparing them with the control samples, we have found a small group of 44 proteins characteristic of the blood plasma samples from patients with chronic cerebral ischemia. These proteins contribute to the processes of homeostasis maintenance, including innate immune response unfolding, the response of a body to stress, and contribution to the blood clotting cascade.

Perfluoroalkyl Acid Concentrations in Blood Samples Subjected to Transportation and Processing Delay.

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Bach, Cathrine Carlsen; Henriksen, Tine Brink; Bossi, Rossana; Bech, Bodil Hammer; Fuglsang, Jens; Olsen, Jørn; Nohr, Ellen Aagaard

2015-01-01

In studies of perfluoroalkyl acids, the validity and comparability of measured concentrations may be affected by differences in the handling of biospecimens. We aimed to investigate whether measured plasma levels of perfluoroalkyl acids differed between blood samples subjected to delay and transportation prior to processing and samples with immediate processing and freezing. Pregnant women recruited at Aarhus University Hospital, Denmark, (n = 88) provided paired blood samples. For each pair of samples, one was immediately processed and plasma was frozen, and the other was delayed and transported as whole blood before processing and freezing of plasma (similar to the Danish National Birth Cohort). We measured 12 perfluoroalkyl acids and present results for compounds with more than 50% of samples above the lower limit of quantification. For samples taken in the winter, relative differences between the paired samples ranged between -77 and +38% for individual perfluoroalkyl acids. In most cases concentrations were lower in the delayed and transported samples, e.g. the relative difference was -29% (95% confidence interval -30; -27) for perfluorooctane sulfonate. For perfluorooctanoate there was no difference between the two setups [corresponding estimate 1% (0, 3)]. Differences were negligible in the summer for all compounds. Transport of blood samples and processing delay, similar to conditions applied in some large, population-based studies, may affect measured perfluoroalkyl acid concentrations, mainly when outdoor temperatures are low. Attention to processing conditions is needed in studies of perfluoroalkyl acid exposure in humans.

Identifying the potential of changes to blood sample logistics using simulation.

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Jørgensen, Pelle; Jacobsen, Peter; Poulsen, Jørgen Hjelm

2013-01-01

Using simulation as an approach to display and improve internal logistics at hospitals has great potential. This study shows how a simulation model displaying the morning blood-taking round at a Danish public hospital can be developed and utilized with the aim of improving the logistics. The focus of the simulation was to evaluate changes made to the transportation of blood samples between wards and the laboratory. The average- (AWT) and maximum waiting time (MWT) from a blood sample was drawn at the ward until it was received at the laboratory, and the distribution of arrivals of blood samples in the laboratory were used as the evaluation criteria. Four different scenarios were tested and compared with the current approach: (1) Using AGVs (mobile robots), (2) using a pneumatic tube system, (3) using porters that are called upon, or (4) using porters that come to the wards every 45 minutes. Furthermore, each of the scenarios was tested in terms of what amount of resources would give the optimal result. The simulations showed a big improvement potential in implementing a new technology/mean for transporting the blood samples. The pneumatic tube system showed the biggest potential lowering the AWT and MWT with approx. 36% and 18%, respectively. Additionally, all of the scenarios had a more even distribution of arrivals except for porters coming to the wards every 45 min. As a consequence of the results obtained in the study, the hospital decided to implement a pneumatic tube system.

Genetic screening of spinal muscular atrophy using a real-time modified COP-PCR technique with dried blood-spot DNA.

Science.gov (United States)

Ar Rochmah, Mawaddah; Harahap, Nur Imma Fatimah; Niba, Emma Tabe Eko; Nakanishi, Kenta; Awano, Hiroyuki; Morioka, Ichiro; Iijima, Kazumoto; Saito, Toshio; Saito, Kayoko; Lai, Poh San; Takeshima, Yasuhiro; Takeuchi, Atsuko; Bouike, Yoshihiro; Okamoto, Maya; Nishio, Hisahide; Shinohara, Masakazu

2017-10-01

Spinal muscular atrophy (SMA) is a common neuromuscular disorder caused by mutations in SMN1. More than 95% of SMA patients carry homozygous SMN1 deletion. SMA is the leading genetic cause of infant death, and has been considered an incurable disease. However, a recent clinical trial with an antisense oligonucleotide drug has shown encouraging clinical efficacy. Thus, early and accurate detection of SMN1 deletion may improve prognosis of many infantile SMA patients. A total of 88 DNA samples (37 SMA patients, 12 carriers and 39 controls) from dried blood spots (DBS) on filter paper were analyzed. All participants had previously been screened for SMN genes by PCR restriction fragment length polymorphism (PCR-RFLP) using DNA extracted from freshly collected blood. DNA was extracted from DBS that had been stored at room temperature (20-25°C) for 1week to 5years. To ensure sufficient quality and quantity of DNA samples, target sequences were pre-amplified by conventional PCR. Real-time modified competitive oligonucleotide priming-PCR (mCOP-PCR) with the pre-amplified PCR products was performed for the gene-specific amplification of SMN1 and SMN2 exon 7. Compared with PCR-RFLP using DNA from freshly collected blood, results from real-time mCOP-PCR using DBS-DNA for detection of SMN1 exon 7 deletion showed a sensitivity of 1.00 (CI [0.87, 1.00])] and specificity of 1.00 (CI [0.90, 1.00]), respectively. We combined DNA extraction from DBS on filter paper, pre-amplification of target DNA, and real-time mCOP-PCR to specifically detect SMN1 and SMN2 genes, thereby establishing a rapid, accurate, and high-throughput system for detecting SMN1-deletion with practical applications for newborn screening. Copyright © 2017 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.

Drying characteristics of zucchini and empirical modeling of its drying process

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Naciye Kutlu

2017-10-01

Full Text Available The aim of the study was to dry zucchini (Cucurbita pepo by two different methods (convective hot-air (CHD and microwave-assisted drying (MWD. The effect of air temperature (60, 70 and 80°C, microwave (MW power (180, 360, 540 W and sample thickness (5 and 10 mm on some drying characteristics of zucchini were investigated. Thirteen mathematical models available in the literature were fitted to the experimental moisture ratio data. The coefficients of the models were determined by non-linear regression analysis. It was determined that the model that fits the moisture ratio data the best varies at different drying conditions. Increasing drying temperature and MW power and reducing sample thickness improved the drying rate and drying time. Drying in microwave has reduced the drying time by 52-64% for zucchini. It was found that the effective moisture diffusivities increased with increasing temperature and MW power. MWD samples had better rehydration ratios compared to ones dried only in tray drier for 5 mm thickness.  

Payload specialist Reinhard Furrer show evidence of previous blood sampling

Science.gov (United States)

1985-01-01

Payload specialist Reinhard Furrer shows evidence of previous blood sampling while Wubbo J. Ockels, Dutch payload specialist (only partially visible), extends his right arm after a sample has been taken. Both men show bruises on their arms.

A content validated questionnaire for assessment of self reported venous blood sampling practices

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Bölenius Karin

2012-01-01

Full Text Available Abstract Background Venous blood sampling is a common procedure in health care. It is strictly regulated by national and international guidelines. Deviations from guidelines due to human mistakes can cause patient harm. Validated questionnaires for health care personnel can be used to assess preventable "near misses"--i.e. potential errors and nonconformities during venous blood sampling practices that could transform into adverse events. However, no validated questionnaire that assesses nonconformities in venous blood sampling has previously been presented. The aim was to test a recently developed questionnaire in self reported venous blood sampling practices for validity and reliability. Findings We developed a questionnaire to assess deviations from best practices during venous blood sampling. The questionnaire contained questions about patient identification, test request management, test tube labeling, test tube handling, information search procedures and frequencies of error reporting. For content validity, the questionnaire was confirmed by experts on questionnaires and venous blood sampling. For reliability, test-retest statistics were used on the questionnaire answered twice. The final venous blood sampling questionnaire included 19 questions out of which 9 had in total 34 underlying items. It was found to have content validity. The test-retest analysis demonstrated that the items were generally stable. In total, 82% of the items fulfilled the reliability acceptance criteria. Conclusions The questionnaire could be used for assessment of "near miss" practices that could jeopardize patient safety and gives several benefits instead of assessing rare adverse events only. The higher frequencies of "near miss" practices allows for quantitative analysis of the effect of corrective interventions and to benchmark preanalytical quality not only at the laboratory/hospital level but also at the health care unit/hospital ward.

Intraosseous blood samples for point-of-care analysis: agreement between intraosseous and arterial analyses.

Science.gov (United States)

Jousi, Milla; Saikko, Simo; Nurmi, Jouni

2017-09-11

Point-of-care (POC) testing is highly useful when treating critically ill patients. In case of difficult vascular access, the intraosseous (IO) route is commonly used, and blood is aspirated to confirm the correct position of the IO-needle. Thus, IO blood samples could be easily accessed for POC analyses in emergency situations. The aim of this study was to determine whether IO values agree sufficiently with arterial values to be used for clinical decision making. Two samples of IO blood were drawn from 31 healthy volunteers and compared with arterial samples. The samples were analysed for sodium, potassium, ionized calcium, glucose, haemoglobin, haematocrit, pH, blood gases, base excess, bicarbonate, and lactate using the i-STAT® POC device. Agreement and reliability were estimated by using the Bland-Altman method and intraclass correlation coefficient calculations. Good agreement was evident between the IO and arterial samples for pH, glucose, and lactate. Potassium levels were clearly higher in the IO samples than those from arterial blood. Base excess and bicarbonate were slightly higher, and sodium and ionised calcium values were slightly lower, in the IO samples compared with the arterial values. The blood gases in the IO samples were between arterial and venous values. Haemoglobin and haematocrit showed remarkable variation in agreement. POC diagnostics of IO blood can be a useful tool to guide treatment in critical emergency care. Seeking out the reversible causes of cardiac arrest or assessing the severity of shock are examples of situations in which obtaining vascular access and blood samples can be difficult, though information about the electrolytes, acid-base balance, and lactate could guide clinical decision making. The analysis of IO samples should though be limited to situations in which no other option is available, and the results should be interpreted with caution, because there is not yet enough scientific evidence regarding the agreement of IO

[DNA quantification of blood samples pre-treated with pyramidon].

Science.gov (United States)

Zhu, Chuan-Hong; Zheng, Dao-Li; Ni, Rao-Zhi; Wang, Hai-Sheng; Ning, Ping; Fang, Hui; Liu, Yan

2014-06-01

To study DNA quantification and STR typing of samples pre-treated with pyramidon. The blood samples of ten unrelated individuals were anticoagulated in EDTA. The blood stains were made on the filter paper. The experimental groups were divided into six groups in accordance with the storage time, 30 min, 1 h, 3 h, 6 h, 12 h and 24h after pre-treated with pyramidon. DNA was extracted by three methods: magnetic bead-based extraction, QIAcube DNA purification method and Chelex-100 method. The quantification of DNA was made by fluorescent quantitative PCR. STR typing was detected by PCR-STR fluorescent technology. In the same DNA extraction method, the sample DNA decreased gradually with times after pre-treatment with pyramidon. In the same storage time, the DNA quantification in different extraction methods had significant differences. Sixteen loci DNA typing were detected in 90.56% of samples. Pyramidon pre-treatment could cause DNA degradation, but effective STR typing can be achieved within 24 h. The magnetic bead-based extraction is the best method for STR profiling and DNA extraction.

IDENTIFICATION AND AUTHENTICATION OF DRY SAMPLES OF SOME MEDICINAL PLANTS USING LEAF EPIDERMAL FEATURES AS MARKER

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Abdullahi Alanamu ABDULRAHAMAN

2016-06-01

Full Text Available Herbal medicine is the oldest and still the most widely used system of medicine in the world today and they are made exclusively from plants. However, most of these medicines or drugs are adulterated due to lack of proper identification of the plant samples. Method of checking adulteration of drug plants is the main focus of this study. The identification and authentication of dry samples of some medicinal plants were carried out using anatomical features. Twenty-five (25 plants materials were collected in Ibadan and Ilorin, Nigeria. The plants studied include Azardiracta indica, Newboudia leavis, Polyalthia longifolia, Cymbopogon citratus, Anarcardium occidentalis, Nicotiana tobbaccum, Jatropha curcas, Chromoleana odorata, Mangifera indica, Terminalia catappa, Ocimum gratisimum, Morus messosygia, Morinda lucida, Psidium guajava, Vitellaria paradoxa, Annona senegalensis, Vernonia amygdalina, Gliricidium sepium, Ravoulvia vomitora, Telferia occindentalis Citrus aurantifolia, C. limon, C. paradisi and C. sinensis. Leaf epidermal anatomy of these selected plants showed no major variations in stomatal complex types, frequency, size and shape of stomatal cells, epidermal cell wall and trichomes between fresh and dry samples. The variations that occur were between different species but not within species. Leaf epidermal anatomy, therefore, proved to be a significant tool for resolution of taxonomic confusion of dried samples of these plants.

Validation and clinical application of a method to quantify nevirapine in dried blood spots and dried breast-milk spots.

Science.gov (United States)

Olagunju, Adeniyi; Amara, Alieu; Waitt, Catriona; Else, Laura; Penchala, Sujan D; Bolaji, Oluseye; Soyinka, Julius; Siccardi, Marco; Back, David; Owen, Andrew; Khoo, Saye

2015-10-01

The validation and clinical application of an LC-MS/MS method for the quantification of nevirapine in dried blood spots (DBS) and dried breast-milk spots (DBMS) are presented. DBS and DBMS were prepared from 50 and 30 μL of nevirapine-spiked whole blood and human breast milk, respectively. Chromatographic separation was achieved on a reverse-phase C18 column with 0.1% formic acid in water/acetonitrile using a solvent gradient programme at a flow rate of 400 μL/min, and detection was by a TSQ Quantum Access triple quadrupole mass spectrometer. The clinical application was evaluated in HIV-positive nursing mothers and their breastfed infants. The assay was validated over the concentration range 50-10,000 ng/mL. Accuracy ranged from 93.3% to 113.4% and precision ranged from 1.9% to 12.0%. The mean (percentage coefficient of variation) recovery of nevirapine from DBS and DBMS was ≥ 70.7% (≤ 8.2) and the matrix effect was ≤ 1.04 (≤ 6.1). Nevirapine was stable in DBS and DBMS for ≥ 15 months at room temperature and -80°C. Mean (SD) AUC0-12, Cmax and Cmin in maternal plasma versus breast milk were 57,808 ng · h/mL (24,315) versus 55,817 ng · h/mL (22,368), 6140 ng/mL (2605) versus 5231 ng/mL (2215) and 4334 ng/mL (1880) versus 4342 ng/mL (2245), respectively. The milk-to-plasma concentration ratio over the dosing interval was 0.94 (0.15). Infant plasma concentrations 2 and 8 h after maternal dosing were 580.6 ng/mL (464.7-1607) and 584.1 ng/mL (381.5-1570), respectively. These methods further extend opportunities for conducting clinical pharmacokinetic studies in nursing mother-infant pairs, especially in resource-limited settings. © The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Characteristics of a new fully programmable blood sampling device for monitoring blood radioactivity during PET

International Nuclear Information System (INIS)

Boellaard, R.; Lingen, A. van; Balen, S.C.M. van; Hoving, B.G.; Lammertsma, A.A.

2001-01-01

The first performance tests of a new fully programmable blood sampling device for monitoring blood radioactivity during positron emission tomography (PET) are described. Blood is withdrawn through 1-mm internal diameter tubing using an infusion pump which can be operated at rates varying from 0 to 600 ml/h. Activity in blood is measured by a 6-cm-thick bismuth germanate crystal connected to a photomultiplier tube and multichannel analyser (MCA) which are positioned within 6 cm lead shielding. Positioning of the tubing is an exact and simple procedure. The minimal readout time of the MCA is 1 s. Two independent energy windows can be set. Operation of the pump and MCA is fully controlled by a PC, i.e. sampling time, interval time and pump rate can be varied at any time during the PET scan by user-defined scripts. A number of characteristics of the new system were studied, such as sensitivity, dead time, linearity, effect of background radiation and pump rate as a function of input pressure. In addition, dispersion was measured as a function of pump rate. Finally, first clinical results were compared with manual samples. The sensitivity equalled 0.7 and 0.2 cps/Bq for 511- and 1022-keV 30% energy windows, respectively, and the system dead time was 500 ns. The system remained linear within 2% with activity concentrations up to 2.5 MBq/cc. Short-term reproducibility was better than 3% for a 1-h period. Long-term reproducibility was about 5% (1SD), which was mainly caused by variation in the diameter of the tubing. If the device was positioned in such a way that maximum shielding was directed towards the patient, the effects of background radiation from the patient on the measured activity concentration for clinically relevant conditions was minimal ( -1 were observed for pump rates higher than 300 ml/h, indicating that the system dispersion is small. Clinical data showed an excellent agreement to within 3% (1SD) between the results obtained with the new system and

The prevalence of toxoplasmosis in Imam Reza Hospital blood bank samples, Tehran, Iran.

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Shaddel, M; Mirzaii Dizgah, I; Sharif, F

2014-10-01

The prevalence of toxoplasma gondii (T.g) infection in blood donors has been poorly studied. The aim of this study was to assess the prevalence of acute and chronic toxoplasmosis in blood products. A total of 223 blood products (101 fresh frozen plasma (FFP) and 122 packed cells (PC)) in Imam Reza hospital blood bank, Tehran, Iran were tested for specific T.g antibodies (IgG and IgM) by ELISA method. Positive IgG anti-T.g samples were further tested for IgM anti-T.g. A positive IgG test with the negative and positive IgM test was interpreted as a chronic and acute toxoplasmosis respectively. Of 223 samples 38.6% and 0.45% were positive for IgG anti-T.g and IgM anti-T.g levels respectively. Therefore, one and 85 samples were involved acute and chronic toxoplasmosis respectively. Twenty-six of fresh frozen plasma samples were positive for IgG anti-T.g and one of them was positive for IgM anti-T.g. Sixty packed cell samples were positive for IgG anti-T.g. Our study showed that there were chronic and acute toxoplasmosis in blood products and the prevalence of toxoplasmosis especially chronic form was high. Therefore screening of blood for T.g antibodies may be considered. Copyright © 2014 Elsevier Ltd. All rights reserved.

[Optimization of trehalose loading in red blood cells before freeze-drying].

Science.gov (United States)

Zhuang, Yuan; Liu, Jing-Han; Ouyang, Xi-Lin; Chen, Lin-Feng; Che, Ji

2007-04-01

The key points for better protection of trehalose in freeze-drying red blood cells (RBCs) are to resolve non-osmosis of trehalose to red blood cells and to make cytoplasmic trehalose to reach effective concentration. This study was aimed to investigate the regularity of loading RBCs with trehalose, screen out optimal loading condition and evaluate the effect of trehalose on physico-chemical parameters of RBCs during the period of loading. The cytoplasmic trehalose concentration in red blood cells, free hemoglobin and ATP level were determined at different incubation temperatures (4, 22 and 37 degrees C), different trehaolse concentrations (0, 200, 400, 600, 800 and 1000 mmol/L) and different incubation times (2, 4, 6, 8 and 10 hours), the cytoplasmic trehalose, free hemoglobin (FHb), hemoglobin (Hb) and mean corpuscular volume (MCV) in fresh RBCs and RBCs stored for 72 hours at 4 degrees C were compared, when loading condition was ensured. The results showed that with increase of incubation temperature, time and extracellular trehalose concentration, the loading of trehalose in RBCs also increased. Under the optimal loading condition, cytoplasmic trehalose concentration and free hemoglobin level of fresh RBCs and RBCs stored for 72 hours at 4 degrees C were 65.505 +/- 6.314 mmol/L, 66.2 +/- 5.002 mmol/L and 6.567 +/- 2.568 g/L, 16.168 +/- 3.922 g/L respectively. It is concluded that the most optimal condition of loading trehalose is that fresh RBCs incubate in 800 mmol/L trehalose solution for 8 hours at 37 degrees C. This condition can result in a efficient cytoplasmic trehalose concentration. The study provides an important basis for long-term preservation of RBCs.

Total phenolics, antioxidant capacity, colour and drying characteristics of date fruit dried with different methods

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Gökçen İZLİ

2016-01-01

Full Text Available Abstract Date slices were dried with the three drying methods convective (60, 70 and 80 °C, microwave (120 W and freeze drying to determine drying characteristics and to compare the dried fruit quality. All colour parameters changed depending on the drying method and colours closest to the fresh sample were obtained with freeze drying. It is interesting to note that the total phenolic content and antioxidant capacity in each sample rose when looked at in relation to the fresh sample. In particular, microwave-dried samples were recorded as having the highest total phenolic content and the highest antioxidant capacity. To explain the drying kinetics of the date slices, nine thin-layer drying models were also attempted. Based on statistical tests, the model developed by Midilli et al. model was found to be the best model for convective and microwave drying, but the Two Term model was the best for freeze drying. This study shows that microwave drying can produce high quality date slices with the additional advantage of reduced drying times compared to convective and freeze drying.

Elimination of heparin interference during microarray processing of fresh and biobank-archived blood samples.

Science.gov (United States)

Hebels, Dennie G A J; van Herwijnen, Marcel H M; Brauers, Karen J J; de Kok, Theo M C M; Chalkiadaki, Georgia; Kyrtopoulos, Soterios A; Kleinjans, Jos C S

2014-07-01

In the context of environmental health research, biobank blood samples have recently been identified as suitable for high-throughput omics analyses enabling the identification of new biomarkers of exposure and disease. However, blood samples containing the anti-coagulant heparin could complicate transcriptomic analysis because heparin may inhibit RNA polymerase causing inefficient cRNA synthesis and fluorophore labelling. We investigated the inhibitory effect of heparin and the influence of storage conditions (0 or 3 hr bench times, storage at room temperature or -80°C) on fluorophore labelling in heparinized fresh human buffy coat and whole blood biobank samples during the mRNA work-up protocol for microarray analysis. Subsequently, we removed heparin by lithium chloride (LiCl) treatment and performed a quality control analysis of LiCl-treated biobank sample microarrays to prove their suitability for downstream data analysis. Both fresh and biobank samples experienced varying degrees of heparin-induced inhibition of fluorophore labelling, making most samples unusable for microarray analysis. RNA derived from EDTA and citrate blood was not inhibited. No effect of bench time was observed but room temperature storage gave slightly better results. Strong correlations were observed between original blood sample RNA yield and the amount of synthesized cRNA. LiCl treatment restored sample quality to normal standards in both fresh and biobank samples and the previously identified correlations disappeared. Microarrays hybridized with LiCl-treated biobank samples were of excellent quality with no identifiable influence of heparin. We conclude that, to obtain high quality results, in most cases heparin removal is essential in blood-derived RNA samples intended for microarray analysis. Copyright © 2014 Wiley Periodicals, Inc.

Proinflammatory gene polymorphisms are potentially associated with Korean non-Sjogren dry eye patients

OpenAIRE

Na, Kyung-Sun; Mok, Jee-Won; Kim, Ja Yeon; Joo, Choun-Ki

2011-01-01

Purpose To determine whether proinflammatory cytokine genes were potential susceptibility candidate genes for Korean patients with non-Sjogren dry eye, we investigated the association of the interleukin 1 beta (IL1B), interleukin 6 (IL6), and interleukin 6 receptor (IL6R) variations with this disease in Korean patients. Methods Genomic DNA was extracted from blood samples of unrelated non-Sjogren dry eye patients and healthy control individuals who visited the Eye Center and Health Promotion ...

The impact of different blood sampling methods on laboratory rats under different types of anaesthesia

DEFF Research Database (Denmark)

Toft, Martin Fitzner; Petersen, Mikke Haxø; Dragsted, Nils

2006-01-01

for rats sampled from the tail vein, which showed fluctuations in body temperature in excess of 30 h after sampling. Increases in heart rate and blood pressure within the first hours after sampling indicated that periorbital puncture was the method that had the largest acute impact on the rats......Rats with implanted telemetry transponders were blood sampled by jugular puncture, periorbital puncture or tail vein puncture, or sampled by jugular puncture in carbon dioxide (CO?), isoflurane or without anaesthesia in a crossover design. Heart rate, blood pressure and body temperature were...... registered for three days after sampling. Initially blood pressure increased, but shortly after sampling it decreased, which led to increased heart rate. Sampling induced rapid fluctuations in body temperature, and an increase in body temperature. Generally, rats recovered from sampling within 2-3 h, except...

Use of filter paper blood samples for rabies antibody detection in foxes and raccoon dogs.

Science.gov (United States)

Wasniewski, Marine; Barrat, Jacques; Combes, Benoit; Guiot, Anne Laure; Cliquet, Florence

2014-08-01

The effectiveness of oral rabies vaccination in wildlife is usually evaluated by the detection of rabies antibodies. However, the assessment of rabies antibodies has several technical difficulties in the field, such as the collection, storage, transport and titration of blood samples, often of poor quality. The objective of this study was to assess the feasibility of collecting blood on a filter paper (FP) coupled with enzyme-linked immunosorbent assay (ELISA) titration of rabies antibodies in raccoon dogs and red foxes. The FP blood sampling method was found highly specific and repeatable in both species. Overall, results obtained with the FP sampling method were highly concordant with the conventional (venipuncture) sampling methods. Blood eluates from FP samples from foxes and raccoon dogs tested using ELISA showed concordance values of 92% and 95%, respectively, with serum samples tested using the seroneutralisation test and values of 95% and 91%, respectively, when the ELISA was used on both types of sample. The use of FP blood sampling coupled with the titration of rabies antibodies by ELISA provides a reliable alternative to conventional blood sampling and serum testing by seroneutralisation. This simple procedure is particularly attractive and cost-effective for assessing the effectiveness of oral rabies vaccination in field conditions. Copyright © 2014. Published by Elsevier B.V.

Analysis of liquid samples using dried-droplet laser ablation inductively coupled plasma mass spectrometry

Energy Technology Data Exchange (ETDEWEB)

Do, Trong-Mui; Hsieh, Hui-Fang; Chang, Wei-Ciang [Department of Biomedical Engineering and Environmental Sciences, National Tsing Hua University, Hsinchu, 30013 Taiwan (China); Chang, E.-E. [Department of Biochemistry, Taipei Medical University, Taipei City, 11031 Taiwan (China); Wang, Chu-Fang, E-mail: cfwang@mx.nthu.edu.tw [Department of Biomedical Engineering and Environmental Sciences, National Tsing Hua University, Hsinchu, 30013 Taiwan (China); Institute of Nuclear Engineering and Science, National Tsing Hua University, Hsinchu 30013, Taiwan (China)

2011-08-15

In this study we developed a dried-droplet method for laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). The proposed method provides accurate and precise results when building calibration curves and determining elements of interest in real liquid samples. After placing just 1 {mu}L of a liquid standard solution or a real sample onto the filter surface and then converting the solution into a very small, thin dry spot, the sample could be applied as an analytical subject for LA. To demonstrate the feasibility of this proposed method, we used LA-ICP-MS and conventional ICP-MS to determine the levels of 13 elements (Li, V, Mn, Co, Ni, Cu, Zn, As, Mo, Cd, Sb, Tl, and Pb) in five water samples. The correlation coefficients obtained from the various calibration curves ranged from 0.9920 ({sup 205}Tl) to 0.9998 ({sup 51}V), sufficient to allow the determination of a wide range of elements in the samples. We also investigated the effects of Methylene Blue (MB) and the NaCl concentration on the elemental analyses. MB could be used as an indicator during the ablation process; its presence in the samples only negligibly influenced the intensities of the signals of most of the tested elements. Notably, high NaCl contents led to signal suppression for some of the elements. In comparison with the established sample introduction by nebulization, our developed technique abrogates the need for time-consuming sample preparation and reduces the possibility of sample contamination.

Blood Sampling in Newborns: A Systematic Review of YouTube Videos.

Science.gov (United States)

Bueno, Mariana; Nishi, Érika Tihemi; Costa, Taine; Freire, Laís Machado; Harrison, Denise

Objective of this study was to conduct a systematic review of YouTube videos showing neonatal blood sampling, and to evaluate pain management and comforting interventions used. Selected videos were consumer- or professional-produced videos showing human newborns undergoing heel lancing or venipuncture for blood sampling, videos showing the entire blood sampling procedure (from the first attempt or puncture to the time of application of a cotton ball or bandage), publication date prior to October 2014, Portuguese titles, available audio. Search terms included "neonate," "newborn," "neonatal screening," and "blood collection." Two reviewers independently screened the videos and extracted the following data. A total of 13 140 videos were retrieved, of which 1354 were further evaluated, and 68 were included. Videos were mostly consumer produced (97%). Heel lancing was performed in 62 (91%). Forty-nine infants (72%) were held by an adult during the procedure. Median pain score immediately after puncture was 4 (interquartile range [IQR] = 0-5), and median length of cry throughout the procedure was 61 seconds (IQR = 88). Breastfeeding (3%) and swaddling (1.5%) were rarely implemented. Posted YouTube videos in Portuguese of newborns undergoing blood collection demonstrate minimal use of pain treatment, and maximal distress during procedures. Knowledge translation strategies are needed to implement effective measures for neonatal pain relief and comfort.

Direct and long-term detection of gene doping in conventional blood samples.

Science.gov (United States)

Beiter, T; Zimmermann, M; Fragasso, A; Hudemann, J; Niess, A M; Bitzer, M; Lauer, U M; Simon, P

2011-03-01

The misuse of somatic gene therapy for the purpose of enhancing athletic performance is perceived as a coming threat to the world of sports and categorized as 'gene doping'. This article describes a direct detection approach for gene doping that gives a clear yes-or-no answer based on the presence or absence of transgenic DNA in peripheral blood samples. By exploiting a priming strategy to specifically amplify intronless DNA sequences, we developed PCR protocols allowing the detection of very small amounts of transgenic DNA in genomic DNA samples to screen for six prime candidate genes. Our detection strategy was verified in a mouse model, giving positive signals from minute amounts (20 μl) of blood samples for up to 56 days following intramuscular adeno-associated virus-mediated gene transfer, one of the most likely candidate vector systems to be misused for gene doping. To make our detection strategy amenable for routine testing, we implemented a robust sample preparation and processing protocol that allows cost-efficient analysis of small human blood volumes (200 μl) with high specificity and reproducibility. The practicability and reliability of our detection strategy was validated by a screening approach including 327 blood samples taken from professional and recreational athletes under field conditions.

Influence of sample quality on phenylalanine and 17-hydroxyprogesterone levels in neonatal screening

OpenAIRE

del Río Fabre, Lesley; González, Ernesto C; Frómeta, Amarilys; Castells, Elisa M; Tejeda, Yileidis

2010-01-01

Quality of the sample and phenylalanine and 17-hydroxyprogesterone levels in neonatal screening. In the programs for neonatal screening many different analytes are quantified from dried blood on filter paper cards. Several factors affect the quality of the samples invalidating their employment in the laboratory: inadequate collection procedures, quality of the filter paper, the drying, storage and transportation under extreme environmental conditions. This article aims to show how the quality...

Dataset of longitudinal analysis of tear cytokine levels, CD4, CD8 counts and HIV viral load in dry eye patients with HIV infection

Directory of Open Access Journals (Sweden)

Praveen Kumar Balne

2017-04-01

Full Text Available The data presented in this article shows the longitudinal analysis of tear fluid cytokine profiles, blood CD4 and CD8 counts and HIV viral load in 34 dry eye patients with HIV infection during the HAART therapy. Clinical samples were collected from HIV patients with dry eye disease at the time of presentation to the clinic (visit 1, three months (visit 2 and 6 months (visit 3 after the presentation. At each time point tear samples were evaluated for 41 cytokines using Luminex bead based multiplex assay and blood samples were tested for HIV viral load and CD4 and CD8 counts.

Multispectral Imaging Analysis of Circulating Tumor Cells in Negatively Enriched Peripheral Blood Samples.

Science.gov (United States)

Miller, Brandon; Lustberg, Maryam; Summers, Thomas A; Chalmers, Jeffrey J

2017-01-01

A variety of biomarkers are present on cells in peripheral blood of patients with a variety of disorders, including solid tumor malignancies. While rare, characterization of these cells for specific protein levels with the advanced technology proposed, will lead to future validation studies of blood samples as "liquid biopsies" for the evaluation of disease status and therapeutic response. While circulating tumor cells (CTCs) have been isolated in the blood samples of patients with solid tumors, the exact role of CTCs as clinically useful predictive markers is still debated. Current commercial technology has significant bias in that a positive selection technology is used that preassumes specific cell surface markers (such as EpCAM) are present on CTCs. However, CTCs with low EpCAM expression have been experimentally demonstrated to be more likely to be missed by this method. In contrast, this application uses a previously developed, technology that performs a purely negative enrichment methodology on peripheral blood, yielding highly enriched blood samples that contain CTCs as well as other, undefined cell types. The focus of this contribution is the use of multispectral imaging of epifluorescent, microscopic images of these enriched cells in order to help develop clinically relevant liquid biopsies from peripheral blood samples.

Survivability of Existing Peripheral Intravenous Access Following Blood Sampling in a Pediatric Population.

Science.gov (United States)

O'Neil, Sheree W; Friesen, Mary Ann; Stanger, Debra; Trickey, Amber Williams

2018-03-07

Although pediatric patients report venipuncture as their most feared experience during hospitalization, blood sampling from peripheral intravenous accesses (PIVs) is not standard of care. Blood sampling from PIVs has long been considered by healthcare personnel to harm the access. In an effort to minimize painful procedures, pediatric nursing staff conducted a prospective, observational study to determine if blood sampling using existing PIVs resulted in the loss of the access. The ability to obtain the sample from the PIV was measured along with patient and PIV characteristics. Specimen collection using 100 existing PIVs was attempted on pediatric inpatients. Each PIV was observed for functionality, infiltration, occlusion, and dislodgement following collection and again in 4h. Frequencies of PIV loss and successful blood sampling were calculated. Patient age, PIV gauge, access site, and PIV age were evaluated for associations with successful sampling using chi-square tests, Fisher's exact tests, and logistic regression. PIV survivability was reported at 99%. The ability to obtain a complete specimen was reported at 76% and found to be significantly related to PIV age and site. Size of PIV and patient's age were not significantly related to successful sampling. Encouraging rates of PIV survivability and collectability suggest blood sampling from PIVs to be a valuable technique to minimize painful and distressful procedures. Nursing practice was changed in this pediatric department. Patients and families are saved the pain and distress of venipuncture. Nurses reported saving time and personal distress by avoiding the venipuncture procedure. Copyright © 2018 Elsevier Inc. All rights reserved.

Sample preparation prior to the LC-MS-based metabolomics/metabonomics of blood-derived samples.

Science.gov (United States)

Gika, Helen; Theodoridis, Georgios

2011-07-01

Blood represents a very important biological fluid and has been the target of continuous and extensive research for diagnostic, or health and drug monitoring reasons. Recently, metabonomics/metabolomics have emerged as a new and promising 'omics' platform that shows potential in biomarker discovery, especially in areas such as disease diagnosis, assessment of drug efficacy or toxicity. Blood is collected in various establishments in conditions that are not standardized. Next, the samples are prepared and analyzed using different methodologies or tools. When targeted analysis of key molecules (e.g., a drug or its metabolite[s]) is the aim, enforcement of certain measures or additional analyses may correct and harmonize these discrepancies. In omics fields such as those performed by holistic analytical approaches, no such rules or tools are available. As a result, comparison or correlation of results or data fusion becomes impractical. However, it becomes evident that such obstacles should be overcome in the near future to allow for large-scale studies that involve the assaying of samples from hundreds of individuals. In this case the effect of sample handling and preparation becomes very serious, in order to avoid wasting months of work from experts and expensive instrument time. The present review aims to cover the different methodologies applied to the pretreatment of blood prior to LC-MS metabolomic/metabonomic studies. The article tries to critically compare the methods and highlight issues that need to be addressed.

Ehrlichia canis morulae and DNA detection in whole blood and spleen aspiration samples.

Science.gov (United States)

Faria, Joice Lara Maia; Dagnone, Ana Sílvia; Munhoz, Thiago Demarchi; João, Carolina Franchi; Pereira, Wanderson Adriano Biscola; Machado, Rosângela Zacarias; Tinucci-Costa, Mirela

2010-01-01

The aim of this study was to compare the detection of Ehrlichia canis morulae and DNA by nPCR in whole blood and spleen aspiration. The sample included 40 dogs showing thrombocytopenia associated to clinical signs suggestive of canine ehrlichiosis. Morulae detection showed that in 35 of the dogs studied, 17 had morulae in spleen tissue, and two in buffy coat smears. E. canis DNA was detected in 29/40 blood samples. We verified that morulae detection is more efficient in cytological preparations from spleen aspiration. On the other hand, nPCR on spleen and blood samples were equally efficient for disease diagnosis.

Suitability of small diagnostic peripheral-blood samples for cell-therapy studies.

Science.gov (United States)

Stephanou, Coralea; Papasavva, Panayiota; Zachariou, Myria; Patsali, Petros; Epitropou, Marilena; Ladas, Petros; Al-Abdulla, Ruba; Christou, Soteroulla; Antoniou, Michael N; Lederer, Carsten W; Kleanthous, Marina

2017-02-01

Primary hematopoietic stem and progenitor cells (HSPCs) are key components of cell-based therapies for blood disorders and are thus the authentic substrate for related research. We propose that ubiquitous small-volume diagnostic samples represent a readily available and as yet untapped resource of primary patient-derived cells for cell- and gene-therapy studies. In the present study we compare isolation and storage methods for HSPCs from normal and thalassemic small-volume blood samples, considering genotype, density-gradient versus lysis-based cell isolation and cryostorage media with different serum contents. Downstream analyses include viability, recovery, differentiation in semi-solid media and performance in liquid cultures and viral transductions. We demonstrate that HSPCs isolated either by ammonium-chloride potassium (ACK)-based lysis or by gradient isolation are suitable for functional analyses in clonogenic assays, high-level HSPC expansion and efficient lentiviral transduction. For cryostorage of cells, gradient isolation is superior to ACK lysis, and cryostorage in freezing media containing 50% fetal bovine serum demonstrated good results across all tested criteria. For assays on freshly isolated cells, ACK lysis performed similar to, and for thalassemic samples better than, gradient isolation, at a fraction of the cost and hands-on time. All isolation and storage methods show considerable variation within sample groups, but this is particularly acute for density gradient isolation of thalassemic samples. This study demonstrates the suitability of small-volume blood samples for storage and preclinical studies, opening up the research field of HSPC and gene therapy to any blood diagnostic laboratory with corresponding bioethics approval for experimental use of surplus material. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Paper membrane-based SERS platform for the determination of glucose in blood samples.

Science.gov (United States)

Torul, Hilal; Çiftçi, Hakan; Çetin, Demet; Suludere, Zekiye; Boyacı, Ismail Hakkı; Tamer, Uğur

2015-11-01

In this report, we present a paper membrane-based surface-enhanced Raman scattering (SERS) platform for the determination of blood glucose level using a nitrocellulose membrane as substrate paper, and the microfluidic channel was simply constructed by wax-printing method. The rod-shaped gold nanorod particles were modified with 4-mercaptophenylboronic acid (4-MBA) and 1-decanethiol (1-DT) molecules and used as embedded SERS probe for paper-based microfluidics. The SERS measurement area was simply constructed by dropping gold nanoparticles on nitrocellulose membrane, and the blood sample was dropped on the membrane hydrophilic channel. While the blood cells and proteins were held on nitrocellulose membrane, glucose molecules were moved through the channel toward the SERS measurement area. Scanning electron microscopy (SEM) was used to confirm the effective separation of blood matrix, and total analysis is completed in 5 min. In SERS measurements, the intensity of the band at 1070 cm(-1) which is attributed to B-OH vibration decreased depending on the rise in glucose concentration in the blood sample. The glucose concentration was found to be 5.43 ± 0.51 mM in the reference blood sample by using a calibration equation, and the certified value for glucose was 6.17 ± 0.11 mM. The recovery of the glucose in the reference blood sample was about 88 %. According to these results, the developed paper-based microfluidic SERS platform has been found to be suitable for use for the detection of glucose in blood samples without any pretreatment procedure. We believe that paper-based microfluidic systems may provide a wide field of usage for paper-based applications.

The effectiveness of cooling conditions on temperature of canine EDTA whole blood samples.

Science.gov (United States)

Tobias, Karen M; Serrano, Leslie; Sun, Xiaocun; Flatland, Bente

2016-01-01

Preanalytic factors such as time and temperature can have significant effects on laboratory test results. For example, ammonium concentration will increase 31% in blood samples stored at room temperature for 30 min before centrifugation. To reduce preanalytic error, blood samples may be placed in precooled tubes and chilled on ice or in ice water baths; however, the effectiveness of these modalities in cooling blood samples has not been formally evaluated. The purpose of this study was to evaluate the effectiveness of various cooling modalities on reducing temperature of EDTA whole blood samples. Pooled samples of canine EDTA whole blood were divided into two aliquots. Saline was added to one aliquot to produce a packed cell volume (PCV) of 40% and to the second aliquot to produce a PCV of 20% (simulated anemia). Thirty samples from each aliquot were warmed to 37.7 °C and cooled in 2 ml allotments under one of three conditions: in ice, in ice after transfer to a precooled tube, or in an ice water bath. Temperature of each sample was recorded at one minute intervals for 15 min. Within treatment conditions, sample PCV had no significant effect on cooling. Cooling in ice water was significantly faster than cooling in ice only or transferring the sample to a precooled tube and cooling it on ice. Mean temperature of samples cooled in ice water was significantly lower at 15 min than mean temperatures of those cooled in ice, whether or not the tube was precooled. By 4 min, samples cooled in an ice water bath had reached mean temperatures less than 4 °C (refrigeration temperature), while samples cooled in other conditions remained above 4.0 °C for at least 11 min. For samples with a PCV of 40%, precooling the tube had no significant effect on rate of cooling on ice. For samples with a PCV of 20%, transfer to a precooled tube resulted in a significantly faster rate of cooling than direct placement of the warmed tube onto ice. Canine EDTA whole blood samples cool most

Measurement of regional cerebral blood flow using one-point arterial blood sampling and microsphere model with 123I-IMP. Correction of one-point arterial sampling count by whole brain count ratio

International Nuclear Information System (INIS)

Makino, Kenichi; Masuda, Yasuhiko; Gotoh, Satoshi

1998-01-01

The experimental subjects were 189 patients with cerebrovascular disorders. 123 I-IMP, 222 MBq, was administered by intravenous infusion. Continuous arterial blood sampling was carried out for 5 minutes, and arterial blood was also sampled once at 5 minutes after 123 I-IMP administration. Then the whole blood count of the one-point arterial sampling was compared with the octanol-extracted count of the continuous arterial sampling. A positive correlation was found between the two values. The ratio of the continuous sampling octanol-extracted count (OC) to the one-point sampling whole blood count (TC5) was compared with the whole brain count ratio (5:29 ratio, Cn) using 1-minute planar SPECT images, centering on 5 and 29 minutes after 123 I-IMP administration. Correlation was found between the two values. The following relationship was shown from the correlation equation. OC/TC5=0.390969 x Cn-0.08924. Based on this correlation equation, we calculated the theoretical continuous arterial sampling octanol-extracted count (COC). COC=TC5 x (0.390969 x Cn-0.08924). There was good correlation between the value calculated with this equation and the actually measured value. The coefficient improved to r=0.94 from the r=0.87 obtained before using the 5:29 ratio for correction. For 23 of these 189 cases, another one-point arterial sampling was carried out at 6, 7, 8, 9 and 10 minutes after the administration of 123 I-IMP. The correlation coefficient was also improved for these other point samplings when this correction method using the 5:29 ratio was applied. It was concluded that it is possible to obtain highly accurate input functions, i.e., calculated continuous arterial sampling octanol-extracted counts, using one-point arterial sampling whole blood counts by performing correction using the 5:29 ratio. (K.H.)

Comparison of Measurements of Autoantibodies to Glutamic Acid Decarboxylase and Islet Antigen-2 in Whole Blood Eluates from Dried Blood Spots Using the RSR-Enzyme Linked Immunosorbent Assay Kits and In-House Radioimmunoassays

Directory of Open Access Journals (Sweden)

Anders Persson

2010-01-01

Full Text Available To evaluate the performance of dried blood spots (DBSs with subsequent analyses of glutamic acid decarboxylase (GADA and islet antigen-2 (IA-2A with the RSR-ELISAs, we selected 80 children newly diagnosed with type 1 diabetes and 120 healthy women. DBSs from patients and controls were used for RSR-ELISAs while patients samples were analysed also with in-house RIAs. The RSR-ELISA-GADA performed well with a specificity of 100%, albeit sensitivity (46% was lower compared to in RIA (56%; P=.008. No prozone effect was observed after dilution of discrepant samples. RSR-ELISA-IA-2A achieved specificity of 69% and sensitivity was lower (59% compared with RIA (66%; P<.001. Negative or low positive patients and control samples in the RSR-ELISA-IA-2A increased after dilution. Eluates from DBS can readily be used to analyse GADA with the RSR-ELISA, even if low levels of autoantibodies were not detected. Some factor could disturb RSR-ELISA-IA-2A analyses.

Usefulness of Dried Blood Spots (DBS) to perform hepatitis C virus genotyping in drug users in Senegal.

Science.gov (United States)

Ndiaye, O; Gozlan, J; Diop-Ndiaye, H; Sall, A S; Chapelain, S; Leprêtre, A; Maynart, M; Gueye, M; Lo, G; Thiam, M; Ba, I; Lacombe, K; Girard, P M; Mboup, S; Kane, C T

2017-03-01

The aim of this pilot study was to analyze the Hepatitis C Virus (HCV) genotypes circulating in Senegal among Drug User (DUs), using Dried Blood Spots (DBS) as RNA source for molecular assays. Heroin and/or cocaine users (n = 506) were recruited in Dakar from April to July 2011, using a Respondent Driven Sampling (RDS) method. DBS preparation consisted of five drops of whole blood from finger applied to a Whatman paper card. HCV infection was screened by the detection of anti-HCV antibodies, using a rapid immune-chromatographic test. HCV RNA was quantified on anti-HCV positive DBS, using the Abbott RealTime HCV® Genotyping was performed on DBS with detectable viral load with Versant® HCV Genotype 2.0 Assay (LiPA) and Abbott RealTime HCV Genotype II assay®. Among the 506 participants, 120 were tested as positive for anti-HCV antibodies and their samples were analyzed for HCV RNA viral load and genotype. Out of the 120 DBS tested, HCV RNA was detected on 25 (20.8%). The median viral load was 15,058 IU/ml (ranging from 710 to 766,740 IU/ml). All positive DBS were suitable for the genotyping assay, that showed a predominance of genotype 1 (21/25) including 16 genotypes 1a and 5 genotypes 1b. HCV genotype 1 prevails in a DU population in Dakar. DBS could be useful for HCV RNA genotyping, but optimal storage conditions should required avoiding RNA impairment. Acknowledging this limitation, DBS could be a great interest for detecting and genotyping HCV viremic patients. J. Med. Virol. 89:484-488, 2017. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

Assessment of Osmotic Pre-Drying Treatment on Drying Rates of Fresh Tomato Fruits

Directory of Open Access Journals (Sweden)

P. A. Idah

2014-06-01

Full Text Available The aim of this work is to investigate the influence of osmotic pre-drying treatments on drying rates of tomato (Lycopersiconesculentum at various drying temperatures. Fresh Roma tomato fruit samples were sliced to a thickness of 5 mm and the seeds were removed. Weight of 300 g was measured for each of the three replicates and immersed in a hypertonic solution of sucrose of different concentrations 40 and 60 oBrix each held for osmotic duration of 1 and 2 hours, drained for 10 min and then dried at 50, 60, and 70 oC in a mechanical dryer. Control samples were also weighed 300 g per replicate and dried at 50, 60, and 70 oC without pre-drying treatment. The initial moisture content of fresh tomato used was 94.5% (wb. Moisture loss of each sample was monitored and recorded hourly until the product has reached the desired final moisture content (≤ 7%.The data collected were subjected to statistical analysis of variance (ANOVA and Duncan New Multiple range tests (DNMRT to ascertain the level of significance differences between the individual treatments and their interaction at p ≤ 0.05.The results show that at all the drying temperatures used, the control tomato samples exhibited the fastest drying rate with an average of 35.2 g/hr, samples pre-treated at 40 oBrix has an average drying rate of 26.6 g/hr, while samples pre-treated at 60 oBrix has the slowest drying rate of 25.2 g/hr. It was also revealed that samples subjected to 1 hour osmotic time have faster drying rates than those treated for 2 hours osmotic time.

The Effect of Pneumatic Tube Systems on the Hemolysis of Biochemistry Blood Samples.

Science.gov (United States)

Cakirca, Gokhan; Erdal, Huseyin

2017-05-01

Pneumatic tube systems (PTSs) are widely used in many hospitals because they lead to reduced turnaround times and cost efficiency. However, PTSs may affect the quality of the blood samples transported to the laboratory. The aim of this study was to investigate the effect of the PTS used in our hospital on the hemolysis of the biochemical blood samples transported to the laboratory. A total of 148 samples were manually transported to the laboratory by hospital staff, 148 samples were transported with the PTS, and 113 were transported with the PTS without use of sponge-rubber inserts (PTSws). Hemolysis rates and the levels of biochemical analytes for the different transportation methods were compared. No significant difference was found between the samples transported manually and with the PTS with regard to hemolysis rate and the levels of biochemical analytes. However, the samples transported with the PTSws showed a significant difference compared with the samples transported manually and with the PTS with regard to hemolysis rate and potassium and lactate dehydrogenase levels. The percentages of the samples that exceeded the permissible threshold for the hemolysis among the samples transported manually, with the PTS, and with the PTSws were 10%, 8%, and 47%, respectively. A PTS can be used safely for transporting biochemistry blood samples to the laboratory. However, a sponge-rubber insert that holds sample tubes must be used with the PTS to prevent the hemolysis of blood samples. Copyright © 2016 Emergency Nurses Association. Published by Elsevier Inc. All rights reserved.

Design and implementation of an external quality assessment program for HIV viral load measurements using dried blood spots.

Science.gov (United States)

Prach, Lisa M; Puren, Adrian; Lippman, Sheri A; Carmona, Sergio; Stephenson, Sophie; Cutler, Ewalde; Barnhart, Scott; Liegler, Teri

2015-03-01

An external quality assurance program was developed for HIV-1 RNA viral load measurements taken from dried blood spots using a reference panel and field-collected specimens. The program demonstrated that accurate and reproducible quantitation can be obtained from field-collected specimens. Residual proviral DNA may confound interpretation in virologically suppressed subjects. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Blood Density Is Nearly Equal to Water Density: A Validation Study of the Gravimetric Method of Measuring Intraoperative Blood Loss.

Science.gov (United States)

Vitello, Dominic J; Ripper, Richard M; Fettiplace, Michael R; Weinberg, Guy L; Vitello, Joseph M

2015-01-01

Purpose. The gravimetric method of weighing surgical sponges is used to quantify intraoperative blood loss. The dry mass minus the wet mass of the gauze equals the volume of blood lost. This method assumes that the density of blood is equivalent to water (1 gm/mL). This study's purpose was to validate the assumption that the density of blood is equivalent to water and to correlate density with hematocrit. Methods. 50 µL of whole blood was weighed from eighteen rats. A distilled water control was weighed for each blood sample. The averages of the blood and water were compared utilizing a Student's unpaired, one-tailed t-test. The masses of the blood samples and the hematocrits were compared using a linear regression. Results. The average mass of the eighteen blood samples was 0.0489 g and that of the distilled water controls was 0.0492 g. The t-test showed P = 0.2269 and R (2) = 0.03154. The hematocrit values ranged from 24% to 48%. The linear regression R (2) value was 0.1767. Conclusions. The R (2) value comparing the blood and distilled water masses suggests high correlation between the two populations. Linear regression showed the hematocrit was not proportional to the mass of the blood. The study confirmed that the measured density of blood is similar to water.