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Sample records for dried blood specimens

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

    PubMed Central

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

    2014-01-01

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

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

    PubMed Central

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

    2013-01-01

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

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

    PubMed

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

    2014-07-01

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

  4. Comparison of Ahlstrom grade 226, Munktell TFN, and Whatman 903 filter papers for dried blood spot specimen collection and subsequent HIV-1 load and drug resistance genotyping analysis.

    PubMed

    Rottinghaus, Erin; Bile, Ebi; Modukanele, Mosetsanagape; Maruping, Maruping; Mine, Madisa; Nkengasong, John; Yang, Chunfu

    2013-01-01

    Dried blood spots (DBS) collected onto filter paper have eased the difficulty of blood collection in resource-limited settings. Currently, Whatman 903 (W-903) filter paper is the only filter paper that has been used for HIV load and HIV drug resistance (HIVDR) testing. We therefore evaluated two additional commercially available filter papers, Ahlstrom grade 226 (A-226) and Munktell TFN (M-TFN), for viral load (VL) testing and HIVDR genotyping using W-903 filter paper as a comparison group. DBS specimens were generated from 344 adult patients on antiretroviral therapy (ART) in Botswana. The VL was measured with NucliSENS EasyQ HIV-1 v2.0, and genotyping was performed for those specimens with a detectable VL (≥ 2.90 log(10) copies/ml) using an in-house method. Bland-Altman analysis revealed a strong concordance in quantitative VL analysis between W-903 and A-226 (bias = -0.034 ± 0.246 log(10) copies/ml [mean difference ± standard deviation]) and W-903 and M-TFN (bias = -0.028 ± 0.186 log(10) copies/ml) filter papers, while qualitative VL analysis for virological failure determination, defined as a VL of ≥ 3.00 log(10) copies/ml, showed low sensitivities for A-266 (71.54%) and M-TFN (65.71%) filter papers compared to W-903 filter paper. DBS collected on M-TFN filter paper had the highest genotyping efficiency (100%) compared to W-903 and A-226 filter papers (91.7%) and appeared more sensitive in detecting major HIVDR mutations. DBS collected on A-226 and M-TFN filter papers performed similarly to DBS collected on W-903 filter paper for quantitative VL analysis and HIVDR detection. Together, the encouraging genotyping results and the variability observed in determining virological failure from this small pilot study warrant further investigation of A-226 and M-TFN filter papers as specimen collection devices for HIVDR monitoring surveys.

  5. Drying drops of blood

    NASA Astrophysics Data System (ADS)

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

    2010-11-01

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

  6. Measures of Viral Load using Abbott Real-Time HIV-1 Assay on Venous and Fingerstick Dried Blood Spots from Provider-Collected Specimens in Malawian District Hospitals

    PubMed Central

    Rutstein, Sarah E.; Kamwendo, Deborah; Lugali, Lebah; Thengolose, Isaac; Tegha, Gerald; Fiscus, Susan A.; Nelson, Julie A. E.; Hosseinipour, Mina C.; Sarr, Abdoulaye; Gupta, Sundeep; Chimbwandira, Frank; Mwenda, Reuben; Mataya, Ronald

    2014-01-01

    Background Viral suppression is a key indicator of antiretroviral therapy (ART) response among HIV-infected patients. Dried blood spots (DBS) are an appealing alternative to conventional plasma-based virologic testing, improving access to monitoring in resource-limited settings. However, validity of DBS obtained from fingerstick in field settings remains unknown. Objectives Investigate feasibility and accuracy of DBS vs plasma collected by healthcare workers in real-world settings of remote hospitals in Malawi. Compare venous DBS to fingerstick DBS for identifying treatment failure. Study design We recruited patients from ART clinics at two district hospitals in Malawi, collecting plasma, venous DBS (vDBS), and fingerstick DBS (fsDBS) cards for the first 149 patients, and vDBS and fsDBS only for the subsequent 398 patients. Specimens were tested using Abbott RealTime HIV-1 Assay (lower detection limit 40 copies/ml (plasma) and 550 copies/ml (DBS)). Results 21/149 (14.1%) had detectable viremia (>1.6 log copies/ml), 13 of which were detectable for plasma, vDBS, and fsDBS. Linear regression demonstrated high correlation for plasma vs. DBS (vDBS: β=1.19, R2 0.93 (p<0.0001); fsDBS β=1.20, R2 0.90 (p<0.0001)) and vDBS vs. fsDBS (β=0.88, R2 0.73, (p<0.0001)). Mean difference between plasma and vDBS was 0.51 log copies/ml [SD: 0.33] and plasma and fsDBS 0.46 log copies/ml [SD: 0.30]. At 5000 copies/ml, sensitivity was 100%, and specificity was 98.6% and 97.8% for vDBS and fsDBS, respectively, compared to plasma. Conclusions DBS from venipuncture and fingerstick perform well at the failure threshold of 5000 copies/ml. Fingerstick specimen source may improve access to virologic treatment monitoring in resource-limited settings given task-shifting in high-volume, low-resource facilities. PMID:24906641

  7. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

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

  8. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

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

  9. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

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

  10. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

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

  11. 21 CFR 862.1675 - Blood specimen collection device.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

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

  12. Microwave applications to rock specimen drying in laboratory

    NASA Astrophysics Data System (ADS)

    Park, Jihwan; Park, Hyeong-Dong

    2014-05-01

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

  13. Morphology of drying blood pools

    NASA Astrophysics Data System (ADS)

    Laan, Nick; Smith, Fiona; Nicloux, Celine; Brutin, David; D-Blood project Collaboration

    2016-11-01

    Often blood pools are found on crime scenes providing information concerning the events and sequence of events that took place on the scene. However, there is a lack of knowledge concerning the drying dynamics of blood pools. This study focuses on the drying process of blood pools to determine what relevant information can be obtained for the forensic application. We recorded the drying process of blood pools with a camera and measured the weight. We found that the drying process can be separated into five different: coagulation, gelation, rim desiccation, centre desiccation, and final desiccation. Moreover, we found that the weight of the blood pool diminishes similarly and in a reproducible way for blood pools created in various conditions. In addition, we verify that the size of the blood pools is directly related to its volume and the wettability of the surface. Our study clearly shows that blood pools dry in a reproducible fashion. This preliminary work highlights the difficult task that represents blood pool analysis in forensic investigations, and how internal and external parameters influence its dynamics. We conclude that understanding the drying process dynamics would be advancement in timeline reconstitution of events. ANR funded project: D-Blood Project.

  14. Drone Transport of Microbes in Blood and Sputum Laboratory Specimens.

    PubMed

    Amukele, Timothy K; Street, Jeff; Carroll, Karen; Miller, Heather; Zhang, Sean X

    2016-10-01

    Unmanned aerial vehicles (UAVs) could potentially be used to transport microbiological specimens. To examine the impact of UAVs on microbiological specimens, blood and sputum culture specimens were seeded with usual pathogens and flown in a UAV for 30 ± 2 min. Times to recovery, colony counts, morphologies, and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS)-based identifications of the flown and stationary specimens were similar for all microbes studied.

  15. Drone Transport of Microbes in Blood and Sputum Laboratory Specimens

    PubMed Central

    Street, Jeff; Carroll, Karen; Miller, Heather; Zhang, Sean X.

    2016-01-01

    Unmanned aerial vehicles (UAVs) could potentially be used to transport microbiological specimens. To examine the impact of UAVs on microbiological specimens, blood and sputum culture specimens were seeded with usual pathogens and flown in a UAV for 30 ± 2 min. Times to recovery, colony counts, morphologies, and matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS)-based identifications of the flown and stationary specimens were similar for all microbes studied. PMID:27535683

  16. Spectral transmission by the specimens of blood-glucose mixture

    NASA Astrophysics Data System (ADS)

    Naumenko, Elena K.; Korolevich, Alexander N.

    2006-08-01

    Influence of glucose addition to specimens of diluted blood on the transmission spectrums have been investigated by experimentally and by the simulation. The coefficients of extinction, scattering and absorption of erythrocytes were calculated with using Mie theory for spherical particles of equal volume. Transmission spectrums were measured with the spectrophotometer Cary500 in the wavelength range 460-860 nm. Specimens of liquid for imbedding of erythrocytes were preparing by mixing blood plasma and 50-% glucose solution with the different concentrations. The volume concentrations (hematocrit) of red blood cells (RBC) were maintained to have the same values in all specimens by adding equal volume of whole blood to immersion liquid of equal volumes. It has been shown that transmission is decreasing for all wavelengths with the addition of glucose solution on condition that the volume concentrations less than 0.35-0.4. The subsequent increase of the glucose concentration results in increasing of spectral transmission.

  17. Molecular sex identification of dry human teeth specimens from Sokoto, Northwestern Nigeria

    PubMed Central

    Zagga, AD; Ahmed, H. OON; Ismail, SM; Tadros, AA

    2014-01-01

    Background: The advent of molecular techniques has revolutionized the ability of scientists to estimate the sex of individuals. Forensic odontology plays an important role in establishing the sex of victims with bodies mutilated beyond recognition due to major disaster. The genetic difference between males and females is defined by the presence or absence of the Y-chromosome. The use of alphoid-repeat primers in sex estimation was first applied on dried blood. Generally, the X, Y alphoid repeats blind test attest to the accuracy of genetic testing, and also point the potential for occasional error in morphometric sexing. Aim: To estimate genetic sex of dry human teeth specimens from Sokoto, Northwestern Nigeria, using polymerase chain reaction (PCR). Materials and Methods: A single-blind study of DNA analysis for sex estimation of nine dry human teeth specimens from Sokoto, Northwestern Nigeria, through PCR, using alphoid repeats primers, was undertaken. Results: The genetic sex of each group of the teeth samples were accurately (100%) identified. For each group of teeth, PCR Sensitivity = 100%, Specificity = 0%, Predictive value of positive test = 100%, Predictive value of negative test = 0%, False positive rate = 0%, False negative rate = 0%, Efficiency of test = 100%. Fisher's exact probability test P = 1. Z-test: z- and P values were invalid. Conclusion: This study has demonstrated the successful use of alphoid-repeat primers in genetic sex identification of human dry teeth samples from Sokoto, Northwestern Nigeria. This is the first known study estimating the sex of human dry teeth specimens by means of PCR in Nigeria. There is need for further studies in Nigeria to complement the findings of this study. PMID:25125922

  18. Assessment of viable bacteria and bacterial DNA in blood and bloodstain specimens stored under various conditions.

    PubMed

    Hosokawa-Muto, Junji; Fujinami, Yoshihito; Mizuno, Natsuko

    2013-11-01

    Microbial forensic specimens that are collected at biocrime and bioterrorism scenes include blood, tissue, cloths containing biological fluids, swabs, water, soil, and aerosols. It is preferable that pathogens in such specimens are alive and kept in a steady state. Specimens may be stored for a prolonged period before analysis; therefore, it is important to understand the effect of the storage conditions on the pathogens contained within the specimens. In this study, we prepared blood and bloodstain specimens containing Gram-negative or -positive bacteria, stored the samples for 482 days under various conditions, and measured viable bacterial counts and total bacterial contents in the samples. Viable bacteria were preserved well in the samples stored at -30 and -80 °C, but were diminished or undetectable in the samples stored at 4 °C and room temperature. The total bacterial content was maintained in the blood samples stored at -30 and -80 °C and in the bloodstain samples stored under all temperature conditions, but decreased in the blood samples stored at 4 °C and room temperature. This study showed that the storage conditions affected viable bacteria and bacterial DNA and that freezing and drying were significant for their long-term storage. We provide important information for the storage of microbial forensic specimens.

  19. Peptidomic analysis of human blood specimens: comparison between plasma specimens and serum by differential peptide display.

    PubMed

    Tammen, Harald; Schulte, Imke; Hess, Rudiger; Menzel, Christoph; Kellmann, Markus; Mohring, Thomas; Schulz-Knappe, Peter

    2005-08-01

    The human Plasma Proteome Project pilot phase aims to analyze serum and plasma specimens to elucidate specimen characteristics by various proteomic techniques to ensure sufficient sample quality for the HUPO main phase. We used our proprietary peptidomics technologies to analyze the samples distributed by HUPO. Peptidomics summarizes technologies for visualization, quantitation, and identification of the low-molecular-weight proteome (<15 kDa), the "peptidome." We analyzed all four HUPO specimens (EDTA plasma, citrate plasma, heparin plasma, and serum) from African- and Asian-American donors and compared them to in-house collected Caucasian specimens. One main finding focuses on the most suitable method of plasma specimen collection. Gentle platelet removal from plasma samples is beneficial for improved specificity. Platelet contamination or activation of platelets by low temperature prior to their removal leads to distinct and multiple peptide signals in plasma samples. Two different specimen collection protocols for platelet-poor plasma are recommended. Further emphasis is placed on the differences between plasma and serum on a peptidomic level. A large number of peptides, many of them in rather high abundance, are only present in serum and not detectable in plasma. This ex vivo generation of multiple peptides hampers discovery efforts and is caused by a variety of factors: the release of platelet-derived peptides, other peptides derived from cellular components or the clot, enzymatic activities of coagulation cascades, and other proteases. We conclude that specimen collection is a crucial step for successful peptide biomarker discovery in human blood samples. For analysis of the low-molecular-weight proteome, we recommend the use of platelet-depleted EDTA or citrate plasma.

  20. Freeze-Dried Human Red Blood Cells

    DTIC Science & Technology

    1992-04-15

    freeze-dried and rehydrated blood cells will be made radioactive with " chromium and infused into my other arm through a hypodermic needle . No more than...directed at: (1) development of buffer formulations based on the glass transition and water replacement theory : (2) establishing standard...survival of transfused red blood cells. The labelled RBC were infused through a 20 gauge needle into the volunteer via a scalp vein in the right arm

  1. Critical point drying for scanning electron microscopy: a semi-automatic method of preparing biological specimens.

    PubMed

    Hall, D J; Skerrett, E J; Thomas, W D

    1978-08-01

    Slow, controlled, rates of critical point bomb heating and of gas venting have been shown to improve the preservation of biological specimens in critical point drying. A procedure that represents a balance between avoidance of specimen damage and speed of operation has been developed for use with CO2 as the transitional fluid. Bomb heating is automated and controlled electronically, and manual venting of the gaseous CO2 is monitored using a gas flow meter.

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

    SciTech Connect

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

    2006-10-15

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

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

    PubMed

    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.

  4. Cross-reactivity of tapentadol specimens with DRI methadone enzyme immunoassay.

    PubMed

    Collins, Ayodele A; Merritt, A Paola; Bourland, James A

    2012-10-01

    A substantial incidence of positive methadone screens for pain management urine specimens using a commercial enzyme immunoassay (EIA) was observed in the absence of a methadone prescription, with negative methadone confirmation by ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS-MS). Tapentadol was the only common prescription among the investigated specimens. Tapentadol or one of its three major metabolites was tested at various concentrations (100-200,000 ng/mL) against the DRI EIAs for methadone and methadone metabolite, to evaluate cross-reactivity. Ninety-seven authentic tapentadol urine specimens that produced false-positive methadone EIA results (cutoff = 130 ng/mL) were analyzed for methadone and tapentadol in compound-specific UPLC-MS-MS confirmation tests. Tapentadol, tapentadol glucuronide, tapentadol sulfate and N-desmethyltapentadol exhibited cross-reactivity with the methadone EIA at 6,500 (2.2%), 25,000 (0.6%), 3,000 (4.4%) and 20,000 ng/mL (0.9%), respectively. No cross-reactivity was observed with the methadone metabolite 2-ethylidine-1,5-dimethyl-3,3-diphenylpyrrolidine EIA. All authentic urine specimens were confirmed to be negative for methadone, but positive for tapentadol and all monitored metabolites. Individual concentrations indicated that separate or combined urinary concentrations of tapentadol and its conjugates may produce false-positive methadone screens through cross-reactivity with the methadone immunoassay. The potential for false-positive results for methadone EIA screening of urine specimens associated with tapentadol prescriptions should be considered when interpreting results.

  5. Freeze-Dried Human Red Blood Cells.

    DTIC Science & Technology

    1992-07-15

    above in vivo survival data and half life fulfill the requirements by the American Association of Blood Banks and the Food and Drug Administration to...190-191 65-109 192-193 67-111 5. No known drug hypersensitivity, atopy or known seasonal or other allergy. 6. Subject does not take any medication on a...I S- Bleeding/coagulation disorder or severe anemia. 8. Negative urine drug and blood alcohol screens. 9. Subject is HIV antibody negative. 10

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

    PubMed

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

    2016-07-01

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

  7. Surface fixation of dried blood by glutaraldehyde and peracetic acid.

    PubMed

    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 < 10% (glutaraldehyde-based preparations). Fixation of the remainder was between 76.9 +/- 8.4% and 102.5 +/- 1.1% with glutaraldehyde and between 19.2% +/- 3.3% and 78.1% +/- 2.4% with peracetic acid. No other preparations showed a potential for blood fixation (< 1.3%). Our findings underline the potential for blood fixation, not only by glutaraldehyde, but also by peracetic acid, and support the evidence that effective cleaning should precede the chemical disinfection.

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

    DOEpatents

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

    1994-01-01

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

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

    DOEpatents

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

    1994-05-03

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

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

    PubMed

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

    2008-04-01

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

  11. Next-Generation Molecular Testing of Newborn Dried Blood Spots for Cystic Fibrosis

    PubMed Central

    Lefterova, Martina I.; Shen, Peidong; Odegaard, Justin I.; Fung, Eula; Chiang, Tsoyu; Peng, Gang; Davis, Ronald W.; Wang, Wenyi; Kharrazi, Martin; Schrijver, Iris; Scharfe, Curt

    2017-01-01

    Newborn screening for cystic fibrosis enables early detection and management of this debilitating genetic disease. Implementing comprehensive CFTR analysis using Sanger sequencing as a component of confirmatory testing of all screen-positive newborns has remained impractical due to relatively lengthy turnaround times and high cost. Here, we describe CFseq, a highly sensitive, specific, rapid (<3 days), and cost-effective assay for comprehensive CFTR gene analysis from dried blood spots, the common newborn screening specimen. The unique design of CFseq integrates optimized dried blood spot sample processing, a novel multiplex amplification method from as little as 1 ng of genomic DNA, and multiplex next-generation sequencing of 96 samples in a single run to detect all relevant CFTR mutation types. Sequence data analysis utilizes publicly available software supplemented by an expert-curated compendium of >2000 CFTR variants. Validation studies across 190 dried blood spots demonstrated 100% sensitivity and a positive predictive value of 100% for single-nucleotide variants and insertions and deletions and complete concordance across the polymorphic poly-TG and consecutive poly-T tracts. Additionally, we accurately detected both a known exon 2,3 deletion and a previously undetected exon 22,23 deletion. CFseq is thus able to replace all existing CFTR molecular assays with a single robust, definitive assay at significant cost and time savings and could be adapted to high-throughput screening of other inherited conditions. PMID:26847993

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

    PubMed Central

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

    2016-01-01

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

  13. Negative immunomagnetic selection of T cells from peripheral blood of presentation AML specimens.

    PubMed

    Le Dieu, Rifca; Taussig, David; Lister, T Andrew; Gribben, John G

    2009-08-31

    To date, studies on T cells in acute myeloid leukemia (AML) have been limited to flow cytometric analysis of whole peripheral blood mononuclear cell (PBMC) specimens or functional work looking at the impact of AML myeloblasts on normal or remission T cells. This lack of information on T cells at the time of presentation with disease is due in part to the difficulty in isolating sufficiently pure T cells from these specimens for further study. Negative immunomagnetic selection has been the method of choice for isolating immune cells for functional studies due to concerns that binding antibodies to the cell surface may induce cellular activation, block ligand-receptor interactions or result in immune clearance. In order specifically to study T cells in presentation AML specimens, we set out to develop a method of isolating highly pure CD4 and CD8 T cells by negative selection from the peripheral blood (PB) of newly diagnosed AML patients. This technique, unlike T cell selection from PB from normal individuals or from patients with chronic lymphocytic leukaemia, was extremely problematic due to properties of the leukaemic myeloblasts. A successful method was eventually optimized requiring the use of a custom antibody cocktail consisting of CD33, CD34, CD123, CD11c and CD36, to deplete myeloblasts.

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

    PubMed

    Hirtz, Christophe; Lehmann, Sylvain

    2015-01-01

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

  15. Comparison of Proteins in Whole Blood and Dried Blood Spot Samples by LC/MS/MS

    NASA Astrophysics Data System (ADS)

    Chambers, Andrew G.; Percy, Andrew J.; Hardie, Darryl B.; Borchers, Christoph H.

    2013-09-01

    Dried blood spot (DBS) sampling methods are desirable for population-wide biomarker screening programs because of their ease of collection, transportation, and storage. Immunoassays are traditionally used to quantify endogenous proteins in these samples but require a separate assay for each protein. Recently, targeted mass spectrometry (MS) has been proposed for generating highly-multiplexed assays for biomarker proteins in DBS samples. In this work, we report the first comparison of proteins in whole blood and DBS samples using an untargeted MS approach. The average number of proteins identified in undepleted whole blood and DBS samples by liquid chromatography (LC)/MS/MS was 223 and 253, respectively. Protein identification repeatability was between 77 %-92 % within replicates and the majority of these repeated proteins (70 %) were observed in both sample formats. Proteins exclusively identified in the liquid or dried fluid spot format were unbiased based on their molecular weight, isoelectric point, aliphatic index, and grand average hydrophobicity. In addition, we extended this comparison to include proteins in matching plasma and serum samples with their dried fluid spot equivalents, dried plasma spot (DPS), and dried serum spot (DSS). This work begins to define the accessibility of endogenous proteins in dried fluid spot samples for analysis by MS and is useful in evaluating the scope of this new approach.

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

    SciTech Connect

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

    1986-03-01

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

  17. Routine preparation of air-dried negatively stained and unstained specimens on holey carbon support films: a review of applications.

    PubMed

    Harris, J Robin; Scheffler, Dirk

    2002-01-01

    Several representative examples are given of the successful application of negative staining across the holes of holey carbon support films using 5% (w/v) ammonium molybdate solution containing trehalose. The inclusion of 0.1% (w/v) trehalose is considered to be most satisfactory, although good data have also been obtained in the presence of 0.01 and 1.0% (w/v) trehalose. The examples given fall into the following groups: protein molecules in the absence of polyethylene glycol (PEG), protein molecules in the presence of PEG (Mr 1000), lipoproteins, lipids and membranes, filaments and tubules, viruses in the absence of PEG, viruses in the presence of PEG, aqueous polymer solutions, and finally for comparison purposes, four unstained samples studied in the presence of trehalose alone. In all these cases, and many others not documented here, successful spreading of the sample across holes has been achieved, with the sample embedded within a thin film of air-dried ammonium molybdate+trehalose. These specimens can be rapidly produced and provide an alternative to negatively stained specimens on carbon support films. Specimen stability in the electron bean is good and such specimens can usually generate superior negatively stained TEM images without flattening and adsorption artefacts. The formation of 2-D arrays/crystals of protein molecules and viruses, suspended across holes in the presence of ammonium molbybdate+trehalose, and trehalose alone, is also demonstrated.

  18. Detection of Immunoglobulin Isotypes from Dried Blood Spots

    PubMed Central

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

    2015-01-01

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

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

    NASA Astrophysics Data System (ADS)

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

    2015-07-01

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

  20. Validation of Blood-Based Assays Using Dried Blood Spots for Use in Large Population Studies

    PubMed Central

    CRIMMINS, EILEEN; KIM, JUNG KI; McCREATH, HEATHER; FAUL, JESSICA; WEIR, DAVID; SEEMAN, TERESA

    2014-01-01

    Assessment of health in large population studies has increasingly incorporated measures of blood-based biomarkers based on the use of dried blood spots (DBS). The validity of DBS assessments made by labs used by large studies is addressed by comparing assay values from DBS collected using conditions similar to those used in the field with values from whole blood samples. The DBS approach generates values that are strongly related to whole blood levels of HbA1c, cystatin C, and C-reactive protein. Assessing lipid levels reliably with DBS appears to be a greater challenge. However, even when DBS values and values from venous blood are highly correlated, they are often on a different scale, and using conventional cutoffs may be misleading. PMID:24784986

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

    PubMed Central

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

    2016-01-01

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

  2. Endophytic fungi from Dragon's blood specimens: isolation, identification, phylogenetic diversity and bioactivity.

    PubMed

    Cui, Jin-long; Guo, Shun-xing; Dong, Hailing; Xiao, Peigen

    2011-08-01

    Endophytic fungi from Dragon's blood specimens of different locations of China were characterized taxonomically and investigated concerning their antimicrobial and antitumor activity against six pathogenic microbes and five tumor cells. A total of 49 endophytic fungi were obtained from Dragon's blood materials of Dracaena spp., 18 taxa were represented by 43 (87.8%) isolates and only six (12.2%) isolates were unknown. Twenty (40.8%) of the isolates displayed antimicrobial activity against at least one pathogenic microorganism. Three isolates YNDC07, BJDC06 and BJDC09 displayed significant antimicrobial activities against Staphylococcus aureus, Cryptococcus neoformans and Aspergillus fumigates, respectively. The results of antitumor activity by the MTT assay revealed that 26.5%, 69.4%, 48.9%, 6.1% and 42.9% of isolate fermentation broths displayed growth inhibition on HepG2 cells, SKVO3 cells, MCF7 cells, HL-60 cells and 293-T cells, respectively. HNDC09 and HNDC10 showed very strong antitumor activity against MCF7 and 293-T, respectively. The results showed that endophytic fungi in Dragon's blood samples were valuable in screening antitumor and antimicrobial bioactivity agents.

  3. Clinical feasibility of dried blood spots: Analytics, validation, and applications.

    PubMed

    Enderle, Yeliz; Foerster, Kathrin; Burhenne, Jürgen

    2016-10-25

    Dried blood spots (DBS) sampling and their specific advantages are becoming common in analytical and clinical routine. Being first established for metabolic disorder screening in neonates, its use emerged to a broad spectrum of clinical applications. Although DBS are easily generated, the conduction of specific analytical and clinical validation procedures should be obligatory when implementing DBS for clinical purposes, e.g. therapeutic drug monitoring or clinical drug trials. A respective recommendation has already been published by the European Bioanalysis Forum. Since no official guidelines are present, investigators are currently free in DBS procedure development and validation. This review summarizes and discusses published clinical validation procedures in relation to their applications to highlight the clinical feasibility of DBS.

  4. RNA sequencing of archived neonatal dried blood spots.

    PubMed

    Bybjerg-Grauholm, Jonas; Hagen, Christian Munch; Khoo, Sok Kean; Johannesen, Maria Louise; Hansen, Christine Søholm; Bækvad-Hansen, Marie; Christiansen, Michael; Hougaard, David Michael; Hollegaard, Mads V

    2017-03-01

    Neonatal dried blood spots (DBS) are routinely collected on standard Guthrie cards for all-comprising national newborn screening programs for inborn errors of metabolism, hypothyroidism and other diseases. In Denmark, the Guthrie cards are stored at - 20 °C in the Danish Neonatal Screening Biobank and each sample is linked to elaborate social and medical registries. This provides a unique biospecimen repository to enable large population research at a perinatal level. Here, we demonstrate the feasibility to obtain gene expression data from DBS using next-generation RNA sequencing (RNA-seq). RNA-seq was performed on five males and five females. Sequencing results have an average of > 30 million reads per sample. 26,799 annotated features can be identified with 64% features detectable without fragments per kilobase of transcript per million mapped reads (FPKM) cutoff; number of detectable features dropped to 18% when FPKM ≥ 1. Sex can be discriminated using blood-based sex-specific gene set identified by the Genotype-Tissue Expression consortium. Here, we demonstrate the feasibility to acquire biologically-relevant gene expression from DBS using RNA-seq which provide a new avenue to investigate perinatal diseases in a high throughput manner.

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

    PubMed

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

    2016-01-01

    The preferred sampling medium for measuring human exposures of persistent organic compounds (POPs) is blood, and relevant sample types include whole blood, plasma, and dried blood spots (DBS). Because information regarding the performance and comparability of measurements across these sample types is limited, it is difficult to compare across studies. This study evaluates the performance of POP measurements in plasma, whole blood and DBS, and presents the distribution coefficients needed to convert concentrations among the three sample types. Blood samples were collected from adult volunteers, along with demographic and smoking information, and analyzed by GC/MS for organochlorine pesticides (OCPs), chlorinated hydrocarbons (CHCs), polychlorinated biphenyls (PCBs), and brominated diphenyl ethers (PBDEs). Regression models were used to evaluate the relationships between the sample types and possible effects of personal covariates. Distribution coefficients also were calculated using physically-based models. Across all compounds, concentrations in plasma were consistently the highest; concentrations in whole blood and DBS samples were comparable. Distribution coefficients for plasma to whole blood concentrations ranged from 1.74 to 2.26 for pesticides/CHCs, averaged 1.69 ± 0.06 for the PCBs, and averaged 1.65 ± 0.03 for the PBDEs. Regression models closely fit most chemicals (R (2) > 0.80), and whole blood and DBS samples generally showed very good agreement. Distribution coefficients estimated using biologically-based models were near one and did not explain the observed distribution. Among the study population, median concentrations of several pesticides/CHCs and PBDEs exceeded levels reported in the 2007-2008 National Health and Nutrition Examination Survey, while levels of other OCPs and PBDEs were comparable or lower. Race and smoking status appeared to slightly affect plasma/blood concentration ratios for several POPs. The experimentally

  6. Degradation of zopiclone during storage of spiked and authentic whole blood and matching dried blood spots.

    PubMed

    Jantos, Ricarda; Vermeeren, Annemiek; Sabljic, Danica; Ramaekers, Johannes G; Skopp, Gisela

    2013-01-01

    Z-drugs such as zopiclone are increasingly involved in forensic cases. Its degradation occurs in solvents and biological fluids. It is assumed that hydrolysis largely accounts for the breakdown of zopiclone in aqueous media. Therefore, a stability study in blood at different storage conditions (-20, 4, 20, and 40°C) was performed to establish changes of the drug's concentration with time, also including its degradation product 2-amino-5-chloropyridine (ACP). As removal of the aqueous phase may stabilize molecules that are prone to hydrolysis, it was assessed whether the use of dried blood spots (DBS) may be an alternative for storing and analyzing zopiclone and ACP. Spiked and authentic blood samples and corresponding DBS were analyzed using fully validated LC-MS/MS assays. There was agreement between the measurement of zopiclone from either blood or matching DBS in freshly prepared samples. Results showed that zopiclone was unstable in blood at all storage temperatures except at -20°C. Stability of zopiclone in spiked and authentic blood was increased in DBS compared to matching blood samples stored at the same condition. About 85 % of the initial concentration of zopiclone was still intact in DBS on day 8 at 20°C. ACP was formed from zopiclone in equimolar amounts in both media. Therefore, determination of both zopiclone and ACP may be helpful to estimate the initial concentration in both media. Pre-analytical conditions have a major impact on the recovery of zopiclone from blood. With respect to its known advantages, DBS can be recommended as a valuable alternative for the determination of zopiclone from blood.

  7. Malaria PCR detection in Cambodian low-transmission settings: dried blood spots versus venous blood samples.

    PubMed

    Canier, Lydie; Khim, Nimol; Kim, Saorin; Eam, Rotha; Khean, Chanra; Loch, Kaknika; Ken, Malen; Pannus, Pieter; Bosman, Philippe; Stassijns, Jorgen; Nackers, Fabienne; Alipon, SweetC; Char, Meng Chuor; Chea, Nguon; Etienne, William; De Smet, Martin; Kindermans, Jean-Marie; Ménard, Didier

    2015-03-01

    In the context of malaria elimination, novel strategies for detecting very low malaria parasite densities in asymptomatic individuals are needed. One of the major limitations of the malaria parasite detection methods is the volume of blood samples being analyzed. The objective of the study was to compare the diagnostic accuracy of a malaria polymerase chain reaction assay, from dried blood spots (DBS, 5 μL) and different volumes of venous blood (50 μL, 200 μL, and 1 mL). The limit of detection of the polymerase chain reaction assay, using calibrated Plasmodium falciparum blood dilutions, showed that venous blood samples (50 μL, 200 μL, 1 mL) combined with Qiagen extraction methods gave a similar threshold of 100 parasites/mL, ∼100-fold lower than 5 μL DBS/Instagene method. On a set of 521 field samples, collected in two different transmission areas in northern Cambodia, no significant difference in the proportion of parasite carriers, regardless of the methods used was found. The 5 μL DBS method missed 27% of the samples detected by the 1 mL venous blood method, but most of the missed parasites carriers were infected by Plasmodium vivax (84%). The remaining missed P. falciparum parasite carriers (N = 3) were only detected in high-transmission areas.

  8. Use of Dried Blood Spots: An Ideal Tool for Medical Anthropology “in the Field”

    PubMed Central

    Benyshek, Daniel C.

    2010-01-01

    The use of dried whole blood spot samples provides medical anthropological researchers—especially those working in remote, isolated communities—with several advantages over traditional methods. Anthropological research utilizing venous-drawn blood samples can create challenges in terms of phlebotomy training, personnel needs, storage and transportation requirements, and participant discomfort. Alternatively, research utilizing dried blood spot samples, via finger stick collection techniques, eliminates or reduces these problems greatly. While the use of dried blood spots is often the best sampling option for anthropologists or other population-level researchers, the method does have some limitations. Nevertheless, as the number of dried blood spot analyte protocols continues to increase, the logistical and participant advantages of dried blood spot methods assure their increased utility in biomedical anthropological research in the future. PMID:20307385

  9. High quality genome-wide genotyping from archived dried blood spots without DNA amplification.

    PubMed

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

    2013-01-01

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

  10. Comparison of S. stercoralis Serology Performed on Dried Blood Spots and on Conventional Serum Samples

    PubMed Central

    Formenti, Fabio; Buonfrate, Dora; Prandi, Rosanna; Marquez, Monica; Caicedo, Cintia; Rizzi, Eleonora; Guevara, Angel G.; Vicuña, Yosselin; Huerlo, Francisco R.; Perandin, Francesca; Bisoffi, Zeno; Anselmi, Mariella

    2016-01-01

    Background: Dried blood spots (DBS) are used for epidemiological surveys on infectious diseases in settings where limited resources are available. In fact, DBS can help to overcome logistic difficulties for the collection, transport and storage of biological specimens. Objective: To evaluate the accuracy of Strongyloides stercoralis serology performed on DBS. Methods: A survey was proposed to children attending a school in the village of Borbon, Ecuador, and to their parents/guardians. Each participant gave consent to the collection of both serum and DBS specimens. DBS absorbed on filter papers were analyzed with a commercially available ELISA test for S. stercoralis antibodies, as well as with standard serology. The agreement between the two methods was assessed through the Cohen’s kappa coefficient. Results: The study sample was composed of 174 children and 61 adults, for a total of 235 serum and 235 DBS samples. The serology was positive in 31/235 (13%) serum samples, and in 27/235 (11%) DBS: 4 samples resulted discordant (positive at standard serology). Cohen’s kappa coefficient was 0.921 (95% CI 0.845 – 0.998), indicating a high rate of concordance. Conclusion: DBS are suitable for in field-surveys requiring serological testing for S. stercoralis. PMID:27877170

  11. Dried blood spot analysis for therapeutic drug monitoring of pazopanib.

    PubMed

    de Wit, Djoeke; den Hartigh, Jan; Gelderblom, Hans; Qian, Yanwen; den Hollander, Margret; Verheul, Henk; Guchelaar, Henk-Jan; van Erp, Nielka P

    2015-12-01

    Dried blood spot (DBS) sampling is potentially a more patient-friendly and flexible alternative to venous sampling of pazopanib. This study determined the agreement between pazopanib DBS and plasma concentrations to facilitate implementation of pazopanib DBS sampling into clinical practice. Paired DBS and plasma samples were collected in 12 patients. Pazopanib plasma concentrations were calculated from DBS concentrations using the formula: plasma concentration = DBSconcentration /(1 - hematocrit). Passing-Bablok and Bland-Altman analyses were used to determine the agreement between calculated and measured plasma concentrations. We predefined a clinical acceptance limit of 25% for the Bland-Altman analysis. Passing-Bablok analysis showed a small constant (intercept estimate, -8.53 [95%CI, -12.22 to -4.41]) and slightly proportional (slope estimate, 1.15 [95%CI, 1.04-1.24]) bias between calculated and measured concentrations. This bias was clinically nonrelevant, as shown by Bland-Altman analysis; the mean ratio of calculated to measured concentrations was 0.94 (95%CI, 0.65-1.23). The clinical acceptance limits were well within these 95% limits of agreement. More specifically, 92.6% of the data points were within the predefined acceptance limits. Pazopanib plasma concentrations can be accurately calculated from DBS concentrations. Although validation of DBS cards prepared by patients themselves is required, these results show that DBS sampling can be used to monitor pazopanib therapy in clinical practice.

  12. Towards reproducible MRM based biomarker discovery using dried blood spots.

    PubMed

    Ozcan, Sureyya; Cooper, Jason D; Lago, Santiago G; Kenny, Diarmuid; Rustogi, Nitin; Stocki, Pawel; Bahn, Sabine

    2017-03-27

    There is an increasing interest in the use of dried blood spot (DBS) sampling and multiple reaction monitoring in proteomics. Although several groups have explored the utility of DBS by focusing on protein detection, the reproducibility of the approach and whether it can be used for biomarker discovery in high throughput studies is yet to be determined. We assessed the reproducibility of multiplexed targeted protein measurements in DBS compared to serum. Eighty-two medium to high abundance proteins were monitored in a number of technical and biological replicates. Importantly, as part of the data analysis, several statistical quality control approaches were evaluated to detect inaccurate transitions. After implementing statistical quality control measures, the median CV on the original scale for all detected peptides in DBS was 13.2% and in Serum 8.8%. We also found a strong correlation (r = 0.72) between relative peptide abundance measured in DBS and serum. The combination of minimally invasive sample collection with a highly specific and sensitive mass spectrometry (MS) technique allows for targeted quantification of multiple proteins in a single MS run. This approach has the potential to fundamentally change clinical proteomics and personalized medicine by facilitating large-scale studies.

  13. Towards reproducible MRM based biomarker discovery using dried blood spots

    PubMed Central

    Ozcan, Sureyya; Cooper, Jason D.; Lago, Santiago G.; Kenny, Diarmuid; Rustogi, Nitin; Stocki, Pawel; Bahn, Sabine

    2017-01-01

    There is an increasing interest in the use of dried blood spot (DBS) sampling and multiple reaction monitoring in proteomics. Although several groups have explored the utility of DBS by focusing on protein detection, the reproducibility of the approach and whether it can be used for biomarker discovery in high throughput studies is yet to be determined. We assessed the reproducibility of multiplexed targeted protein measurements in DBS compared to serum. Eighty-two medium to high abundance proteins were monitored in a number of technical and biological replicates. Importantly, as part of the data analysis, several statistical quality control approaches were evaluated to detect inaccurate transitions. After implementing statistical quality control measures, the median CV on the original scale for all detected peptides in DBS was 13.2% and in Serum 8.8%. We also found a strong correlation (r = 0.72) between relative peptide abundance measured in DBS and serum. The combination of minimally invasive sample collection with a highly specific and sensitive mass spectrometry (MS) technique allows for targeted quantification of multiple proteins in a single MS run. This approach has the potential to fundamentally change clinical proteomics and personalized medicine by facilitating large-scale studies. PMID:28345601

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

    PubMed

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

    2013-12-15

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

  15. Anatomical Analysis on the Lateral Bone Window of the Sella Turcica: A Study on 530 Adult Dry Skull Base Specimens

    PubMed Central

    WANG, Jianxin; WANG, Renzhi; LU, Yuntao; YAO, Yong; QI, Songtao

    2014-01-01

    Objective: To investigate the morphometric characteristics of the lateral bone window (LBW) of the sella turica. Methods: A descriptive anatomical study of LBW was performed in 530 cases of dry skull base specimens with relatively complete sella turcica. Detailed morphometric characteristics such as dimensions and calssification of the LBW was studied. All data analysis was performed using SPSS 17.0 statistical software. Results: LBW is located in the lateral bony structures of sella turcica. The mean area sizes of the LBW were 75.99 ± 25.81 mm2 (left) and 76.00 ± 25.53 mm2 (right). There was no significant difference and there was a ​​good positive correlation between bilateral areas of the LBWs. The area size of the LBWs is graded as follows: Grade A (< 60 mm2), B (60~90 mm2) and C (> 90 mm2). LBW morphology was typed as follows: Type I, II, III, IV. And Tpye III includes Type IIIa and IIIb; Type IV includes Type IVa, IVb, IVc, IVd. Conclusions: The lateral bone window of sella turcica is an important structure located between pituiary fossa and parasellar region. The morphological measurements and variations of LBW in this study will provide preliminary data for further anatomical study of sella turcica. Moreover, knowing detailed anatomy of this region is essential for neurosurgeons who make surgery on cranial base or for teaching about the sella turcica in the neuroanatomy lab. PMID:24465158

  16. Development of a PCR Assay to Detect Low Level Trypanosoma cruzi in Blood Specimens Collected with PAXgene Blood DNA Tubes for Clinical Trials Treating Chagas Disease

    PubMed Central

    Wei, Bo; Chen, Lei; Kibukawa, Miho; Kang, John; Waskin, Hetty; Marton, Matthew

    2016-01-01

    Chagas disease is caused by the parasitic infection of Trypanosoma cruzi (T. cruzi). The STOP CHAGAS clinical trial was initiated in 2011 to evaluate posaconazole in treating Chagas disease, with treatment success defined as negative qualitative PCR results of detecting the parasites in blood specimens collected post-treatment. PAXgene Blood DNA tubes were utilized as a simple procedure to collect and process blood specimens. However, the PAXgene blood specimens challenged published T. cruzi PCR methods, resulting in poor sensitivity and reproducibility. To accurately evaluate the treatment efficacy of the clinical study, we developed and validated a robust PCR assay for detecting low level T. cruzi in PAXgene blood specimens. The assay combines a new DNA extraction method with a custom designed qPCR assay, resulting in limit of detection of 0.005 and 0.01 fg/μl for K98 and CL Brener, two representative strains of two of T. cruzi’s discrete typing units. Reliable qPCR standard curves were established for both strains to measure parasite loads, with amplification efficiency ≥ 90% and the lower limit of linearity ≥ 0.05 fg/μl. The assay successfully analyzed the samples collected from the STOP CHAGAS study and may prove useful for future global clinical trials evaluating new therapies for asymptomatic chronic Chagas disease. PMID:27906977

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

    PubMed Central

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

    2014-01-01

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

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

    PubMed Central

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

    2015-01-01

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

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

    PubMed Central

    Kopp, Markus; Rychlik, Michael

    2015-01-01

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

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

    PubMed

    Kopp, Markus; Rychlik, Michael

    2015-01-01

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

  1. Zika Virus Infection and Prolonged Viremia in Whole-Blood Specimens.

    PubMed

    Mansuy, Jean Michel; Mengelle, Catherine; Pasquier, Christophe; Chapuy-Regaud, Sabine; Delobel, Pierre; Martin-Blondel, Guillaume; Izopet, Jacques

    2017-05-15

    We tested whole-blood and plasma samples from immunocompetent patients who had had benign Zika virus infections and found that Zika virus RNA persisted in whole blood substantially longer than in plasma. This finding may have implications for diagnosis of acute symptomatic and asymptomatic infections and for testing of blood donations.

  2. Dried tube specimens: a simple and cost-effective method for preparation of HIV proficiency testing panels and quality control materials for use in resource-limited settings.

    PubMed

    Parekh, Bharat S; Anyanwu, Juliana; Patel, Hetal; Downer, Marie; Kalou, Mireille; Gichimu, Catherine; Keipkerich, Bera Steven; Clement, Nelly; Omondi, Michael; Mayer, Oren; Ou, Chin-Yih; Nkengasong, John N

    2010-02-01

    HIV testing has rapidly expanded worldwide, but proficiency testing (PT) programs to monitor and improve the quality of testing are often lacking in resource-limited settings (RLS). Traditional PT programs and quality control reagents use serum or plasma specimens requiring stringent conditions for storage and transportation. A novel, simple and easy to use approach, based on dried tube specimens (DTS), was developed that can help monitor the quality of HIV antibody testing in RLS. DTS were prepared by drying 20 microl of specimen overnight at room temperature. The addition of a green dye (0.1%) made the DTS pellets visible without affecting the test results. Before testing, the DTS were rehydrated with 200 microl of PBS-Tween buffer. A panel of 303 DTS samples (135 HIV positive and 168 HIV negative) was evaluated with two rapid tests. Sensitivity and specificity with the Determine HIV-1/2 test were 99.3% and 99.4%, respectively, and with OraQuick were 98.5% and 100%, respectively. Stability studies showed that HIV-specific antibodies in the DTS specimens were stable at 4 degrees C and 25 degrees C for 4 weeks, with only marginal decline at 37 degrees C and 45 degrees C over 4 weeks. The DTS-based PT program was piloted successfully in 24 testing sites in Kenya. Results demonstrate that the DTS is a simple to use, practical method to prepare and distribute PT panels and quality control specimens to monitor HIV testing practices in RLS.

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

    PubMed

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

    2016-06-21

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

  4. Evaluation of Dried Blood Spots with a Multiplex Assay for Measuring Recent HIV-1 Infection

    PubMed Central

    Curtis, Kelly A.; Ambrose, Krystin M.; Kennedy, M. Susan; Owen, S. Michele

    2014-01-01

    Laboratory-based HIV tests for recent infection (TRIs), which primarily measure a specific serological biomarker(s) that distinguishes recent from long-term HIV infection, have facilitated the estimation of population-based incidence. Dried blood spots (DBS) on filter paper are an attractive sample source for HIV surveillance, given the simplified and cost-effective methods of specimen collection, storage, and shipment. Here, we evaluated the use of DBS in conjunction with an in-house multiplex TRI, the HIV-1-specific Bio-Plex assay, which measures direct antibody binding and avidity to multiple HIV-1 analytes. The assay performance was comparable between matched plasma and DBS samples from HIV-1 infected individuals obtained from diverse sources. The coefficients of variation, comparing the median antibody reactivity for each analyte between plasma and DBS, ranged from 2.78% to 9.40% and the correlation coefficients between the two sample types ranged from 0.89 to 0.97, depending on the analyte. The correlation in antibody reactivity between laboratory and site-prepared DBS for each analyte ranged from 0.87 to 0.98 and from 0.90 to 0.97 between site-prepared DBS and plasma. The correlation in assay measures between plasma and DBS indicate that the sample types can be used interchangeably with the Bio-Plex format, without negatively impacting the misclassification rate of the assay. PMID:25232736

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

    PubMed

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

    2016-07-01

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

  6. DNA methylome profiling using neonatal dried blood spot samples: a proof-of-principle study.

    PubMed

    Hollegaard, Mads Vilhelm; Grauholm, Jonas; Nørgaard-Pedersen, Bent; Hougaard, David Michael

    2013-04-01

    DNA methylation is the most common DNA modification and perhaps the best described epigenetic modification. It is believed to be important for genomic imprinting and gene regulation and has been associated with the development of diseases such as schizophrenia and some types of cancer. Neonatal dried blood spot samples, commonly known as Guthrie cards, are routinely collected worldwide to screen newborns for diseases. Some countries, including Denmark, have been storing the excess neonatal dried blood spot samples in biobanks for decades. Representing a high percentage of the population under a certain age, the neonatal dried blood spot samples are a potential alternative to collecting new samples to study diseases. As such, neonatal dried blood spot samples have previously been used for DNA genotyping studies with excellent results. However, the amount of material available for research is often limited, challenging researchers to generate the most data from a limited quantity of material. In this proof-of-principle study, we address whether two 3.2mm disks punched from a neonatal dried blood spot sample contain enough DNA for genome-wide methylome profiling, measuring 27,578 loci at the same time. We selected two subjects and carried out the following with each: 1) collected an adult whole-blood sample as reference, 2) spotted a fraction of the whole-blood sample onto a similar type of filter paper as used in the newborn screening and stored it for 3years to serve as a dried blood spot reference, and 3) identified the archived neonatal dried blood spot samples, stored for 26-28years, in the Danish Newborn Screening Biobank as a representative of the archived samples. For comparison, we used two different kits for DNA extraction. The DNA, extracted using the Extract-N-Amp Blood PCR kit, was analyzed, and no statistically significant differences were observed (P<0.001) when we compared the methylation profile of the reference whole-blood samples to the dried blood

  7. Comparison of a PCR assay in whole blood and serum specimens for canine brucellosis diagnosis.

    PubMed

    Keid, L B; Soares, R M; Vasconcellos, S A; Salgado, V R; Megid, J; Richtzenhain, L J

    2010-07-17

    The performance of a serum PCR assay was compared with that of a blood PCR assay for the diagnosis of canine brucellosis caused by Brucella canis in 72 dogs. The dogs were classified into three groups (infected, non-infected and suspected brucellosis) according to the results of blood culture and serological tests. The sensitivities of blood PCR and serum PCR were, respectively, 97.14 per cent and 25.71 per cent. The specificities of both were 100 per cent. In the group of dogs with suspected brucellosis, three were positive by blood PCR and none was positive by serum PCR. Serum PCR showed little value for the direct diagnosis of canine brucellosis as the assay had low diagnostic sensitivity and fewer positive dogs were detected by this test than by blood culture, blood PCR, rapid slide agglutination test (RSAT) and RSAT with 2-mercaptoethanol.

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

    PubMed Central

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

    2016-01-01

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

  9. Collecting in collections: a PCR strategy and primer set for DNA barcoding of decades-old dried museum specimens.

    PubMed

    Mitchell, Andrew

    2015-09-01

    Natural history museums are vastly underutilized as a source of material for DNA analysis because of perceptions about the limitations of DNA degradation in older specimens. Despite very few exceptions, most DNA barcoding projects, which aim to obtain sequence data from all species, generally use specimens collected specifically for that purpose, instead of the wealth of identified material in museums, constrained by the lack of suitable PCR methods. Any techniques that extend the utility of museum specimens for DNA analysis therefore are highly valuable. This study first tested the effects of specimen age and PCR amplicon size on PCR success rates in pinned insect specimens, then developed a PCR primer set and amplification strategy allowing greatly increased utilization of older museum specimens for DNA barcoding. PCR success rates compare favourably with the few published studies utilizing similar aged specimens, and this new strategy has the advantage of being easily automated for high-throughput laboratory workflows. The strategy uses hemi-nested, degenerate, M13-tailed PCR primers to amplify two overlapping amplicons, using two PCRs per amplicon (i.e. four PCRs per DNA sample). Initial PCR products are reamplified using an internal primer and a M13 primer. Together the two PCR amplicons yield 559 bp of the COI gene from Coleoptera, Lepidoptera, Diptera, Hemiptera, Odonata and presumably also other insects. BARCODE standard-compliant data were recovered from 67% (56 of 84) of specimens up to 25 years old, and 51% (102 of 197) of specimens up to 55 years old. Given the time, cost and specialist expertise required for fieldwork and identification, 'collecting in collections' is a viable alternative allowing researchers to capitalize on the knowledge captured by curation work in decades past.

  10. State-of-the-art dried blood spot analysis: an overview of recent advances and future trends.

    PubMed

    Meesters, Roland Jw; Hooff, Gero P

    2013-09-01

    Dried blood spots have become a popular method in a variety of micro blood-sampling techniques in the life sciences sector, consequently competing with the field of conventional, invasive blood sampling by venepuncture. Dried blood spots are widely applied in numerous bioanalytical assays and have gained a significant role in the screening of inherited metabolic diseases, in PK and PD modeling; in the treatment and diagnosis of infectious diseases; and in therapeutic drug monitoring. Recent technological developments such as automation, online extraction, mass spectrometric direct analysis and also conventional dried blood spot bioanalysis, as well as future developments in dried blood spot bioanalysis are highlighted and presented in this article.

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

    PubMed Central

    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

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

    EPA Science Inventory

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

  13. Dried Blood Spots for Viral Load Monitoring in Malawi: Feasible and Effective

    PubMed Central

    Rutstein, Sarah E.; Hosseinipour, Mina C.; Kamwendo, Deborah; Soko, Alice; Mkandawire, Memory; Biddle, Andrea K.; Miller, William C.; Weinberger, Morris; Wheeler, Stephanie B.; Sarr, Abdoulaye; Gupta, Sundeep; Chimbwandira, Frank; Mwenda, Reuben; Kamiza, Steve; Hoffman, Irving; Mataya, Ronald

    2015-01-01

    Objectives To evaluate the feasibility and effectiveness of dried blood spots (DBS) use for viral load (VL) monitoring, describing patient outcomes and programmatic challenges that are relevant for DBS implementation in sub-Saharan Africa. Methods We recruited adult antiretroviral therapy (ART) patients from five district hospitals in Malawi. Eligibility reflected anticipated Ministry of Health VL monitoring criteria. Testing was conducted at a central laboratory. Virological failure was defined as >5000 copies/ml. Primary outcomes were program feasibility (timely result availability and patient receipt) and effectiveness (second-line therapy initiation). Results We enrolled 1,498 participants; 5.9% were failing at baseline. Median time from enrollment to receipt of results was 42 days; 79.6% of participants received results within 3 months. Among participants with confirmed elevated VL, 92.6% initiated second-line therapy; 90.7% were switched within 365 days of VL testing. Nearly one-third (30.8%) of participants with elevated baseline VL had suppressed (<5,000 copies/ml) on confirmatory testing. Median period between enrollment and specimen testing was 23 days. Adjusting for relevant covariates, participants on ART >4 years were more likely to be failing than participants on therapy 1–4 years (RR 1.7, 95% CI 1.0-2.8); older participants were less likely to be failing (RR 0.95, 95% CI 0.92-0.98). There was no difference in likelihood of failure based on clinical symptoms (RR 1.17, 95% CI 0.65-2.11). Conclusions DBS for VL monitoring is feasible and effective in real-world clinical settings. Centralized DBS testing may increase access to VL monitoring in remote settings. Programmatic outcomes are encouraging, especially proportion of eligible participants switched to second-line therapy. PMID:25898365

  14. Use of dried blood spots and inductively coupled plasma mass spectrometry for multi-element determination in blood.

    PubMed

    Vacchina, Véronique; Huin, Vincent; Hulo, Sébastien; Cuny, Damien; Broly, Franck; Renom, Gilles; Perini, Jean-Marc

    2014-07-01

    The paper describes the development of an inductively coupled plasma mass spectrometry (ICP MS) method for multitrace element determination in dried blood spots (DBSs). The analytical conditions were optimized using Seronorm™ L-3 and L-1 Certified Reference Materials. The best results were obtained by sampling blood drops on a decontaminated PVDF filter membrane. After drying under metal-free conditions, the DBSs underwent acidic digestion and were analyzed with ICP MS. The method was then validated for As, Cd, Cu, Pb, Mo, Se and Zn. Using a matrix-matched calibration curve, the recovery levels ranged from 96% to 117%. The repeatability and reproducibility were generally below 15%. Limits of quantification ranging from 0.5 to 50 μg/L. In order to investigate the analytical procedure under real sampling conditions, the results obtained from DBSs and liquid blood aliquots (less subject to contamination) from two adult subjects were compared.

  15. Hemoglobin precipitation greatly improves 4-methylumbelliferone-based diagnostic assays for lysosomal storage diseases in dried blood spots.

    PubMed

    Oemardien, L F; Boer, A M; Ruijter, G J G; van der Ploeg, A T; de Klerk, J B C; Reuser, A J J; Verheijen, F W

    2011-01-01

    Derivatives of 4-methylumbelliferone (4MU) are favorite substrates for the measurement of lysosomal enzyme activities in a wide variety of cell and tissue specimens. Hydrolysis of these artificial substrates at acidic pH leads to the formation of 4-methylumbelliferone, which is highly fluorescent at a pH above 10. When used for the assay of enzyme activities in dried blood spots the light emission signal can be very low due to the small sample size so that the patient and control ranges are not widely separated. We have investigated the hypothesis that quenching of the fluorescence by hemoglobin leads to appreciable loss of signal and we show that the precipitation of hemoglobin with trichloroacetic acid prior to the measurement of 4-methylumbelliferone increases the height of the output signal up to eight fold. The modified method provides a clear separation of patients' and controls' ranges for ten different lysosomal enzyme assays in dried blood spots, and approaches the conventional leukocyte assays in outcome quality.

  16. [Determination of blood constituents with dry reagent carriers. Comparison with wet chemical tests (author's transl)].

    PubMed

    Thomas, L; Appel, W; Storz, G; Plischke, W

    1981-08-28

    Quantitative measurement of blood constituents (glucose, uric acid, urea-N) on dry reagent carriers was compared with corresponding wet-chemical methods. There was a good correlation (r greater than 0.95) and excellent linear regression with small scatter between the two methods for the three constituents. The dry test has proved to be a precise, simple, rapid and economic method, especially suitable for the practitioner and emergency laboratory.

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

    PubMed

    Bielanski, A

    2005-04-15

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

  18. Evaluation of the One-Step ELISA kit for the detection of buprenorphine in urine, blood, and hair specimens.

    PubMed

    Cirimele, V; Etienne, S; Villain, M; Ludes, B; Kintz, P

    2004-07-16

    A solid-phase enzyme immunoassay involving microtiter plates was recently proposed by International Diagnostic Systems corporation (IDS) to screen for buprenorphine in human serum. The performance of the kit led us to investigate its applicability in other biological matrices such as urine or blood, and also hair specimens. Low concentrations of buprenorphine were detected with the ELISA test and confirmed by HPLC/MS (buprenorphine concentrations measured by HPLC/MS: 0.3 ng/mL in urine, 0.2 ng/mL in blood, and 40 pg/mg in hair). The intra-assay precision values were 8.7% at 1 ng/mL of urine (n = 8), 11.5% at 2 ng/mL in serum (n = 8), and 11.5% at 250 pg/mg of hair (n = 8), respectively. The immunoassay had no cross-reactivity with dihydrocodeine, ethylmorphine, 6-monoacetylmorphine, pholcodine, propoxyphene, dextromoramide, dextrometorphan at 1 and 10 mg/L, or codeine, morphine, methadone, and its metabolite EDDP. A 1% cross-reactivity was measured for a norbuprenorphine concentration of 50 ng/mL. Finally, the immunoassay was validated by comparing authentic specimens results with those of a validated HPLC/MS method. From the 136 urine samples tested, 93 were positive (68.4%) after the ELISA screening test (cutoff: 0.5 ng/mL) and confirmed by HPLC/MS (buprenorphine concentrations: 0.3-2036 ng/mL). From the 108 blood or serum samples screened, 27 were positive (25%) after the ELISA test with a cutoff value of 0.5 ng/mL (buprenorphine concentrations: 0.2-13.3 ng/mL). Eighteen hair specimens were positive (72%) after the screening (cutoff: 10 pg/mg) and confirmed by LC/MS (buprenorphine concentrations: 40-360 pg/mg). The ELISA method produced false positive results in less than 21% of the cases, but no false negative results were observed with the immunological test. Four potential adulterants (hypochloride 50 mL/L, sodium nitrite 50 g/L, liquid soap 50 mL/L, and sodium chloride 50 g/L) that were added to 10 positive urine specimens (buprenorphine concentrations in

  19. Differential fluorescent staining method for detection of bacteria in blood cultures, cerebrospinal fluid and other clinical specimens.

    PubMed

    Fazii, P; Ciancaglini, E; Riario Sforza, G

    2002-05-01

    The aim of this study was to evaluate a differential staining method to distinguish gram-positive from gram-negative bacteria in fluorescence. The method is based on two fluorochromes, one acting in the wavelength of red, i.e. the acridine orange, and another acting in the wavelength of green, i.e. the fluorescein. With this method, gram-positive bacteria appear yellow and gram-negative bacteria appear green. In view of the importance of a rapid aetiological diagnosis in cases of septicaemia, the differential staining method in fluorescence was compared with Gram stain for the detection of bacteria in blood. Of 5,820 blood cultures entered into the study and identified by the Bactec 9120 fluorescent series instrument (Becton Dickinson Europe, France), 774 were positive. Of the 774 positive cultures, 689 yielded only a single organism. The differential staining method in fluorescence detected 626 of the 689 cultures, while Gram stain detected 468. On the basis of these results, the sensitivity of the differential staining method in fluorescence was 90.9%, while that of Gram stain was 67.9%. The difference between the two methods was statistically significant ( P<0.001). The differential fluorescent staining method was more sensitive than Gram stain in the detection of bacteria in blood cultures during the incubation period. This technique provides a rapid, simple and highly sensitive staining method that can be used in conjunction with subculture methods. Whereas subculture requires an incubation period of 18-24 h, the fluorescent staining technique can detect bacteria on the same day that smears are prepared and examined. The differential fluorescent staining method was also evaluated for its ability to detect microorganisms in cerebrospinal fluid and other clinical specimens. The microorganisms were easily detected, even when bacterial counts in the specimens were low.

  20. Stability Study of Cervical Specimens Collected by Swab and Stored Dry Followed by Human Papillomavirus DNA Detection Using the cobas 4800 Test.

    PubMed

    Lin, Chun-Qing; Zeng, Xi; Cui, Jian-Feng; Liao, Guang-Dong; Wu, Ze-Ni; Gao, Qian-Qian; Zhang, Xun; Yu, Xiu-Zhang; Chen, Wen; Xi, Ming-Rong; Qiao, You-Lin

    2017-02-01

    Safer, more convenient methods for cervical sample collection and storage are necessary to facilitate human papillomavirus (HPV) DNA testing in low-resource settings. Our study aimed to evaluate the stability of cervical specimens collected with dry swabs and stored dry, compared to liquid-based cytology (LBC) samples, as detected by HPV DNA testing. Women with abnormal cytological findings or HPV-positive results at colposcopy were recruited from the West China Second University Hospital, Sichuan University, between October 2013 and March 2014. From each woman, physicians collected cervical specimens with a swab placed into a Sarstedt tube and a CytoBrush placed into LBC medium. Samples were randomly assigned to be stored at uncontrolled ambient temperature for 2, 7, 14, or 28 days and then were tested for 14 high-risk HPV (HR-HPV) types using the cobas HPV test. The rates of agreement between dry swab and LBC samples for any HR-HPV type, HPV16, HPV18, and the 12 pooled HR-HPV types were 93.8%, 97.8%, 99.4%, and 93.2%, respectively, with kappa values of 0.87 (95% confidence interval [CI], 0.83 to 0.91), 0.94 (95% CI, 0.91 to 0.97), 0.94 (95% CI, 0.87 to 1.00), and 0.86 (95% CI, 0.82 to 0.90). The performance of swab samples for detection of cervical precancerous lesions by means of cobas HPV testing was equal to that of LBC samples, even with stratification by storage time. Dry storage of swab-collected cervical samples can last for 1 month without loss of test performance by cobas HPV testing, compared to LBC samples, which may offer a simple inexpensive approach for cervical cancer screening in low-resource settings.

  1. The stability of iso-α-acids and reduced iso-α-acids in stored blood specimens.

    PubMed

    Rodda, Luke N; Gerostamoulos, Dimitri; Drummer, Olaf H

    2014-06-01

    The long-term stability of the iso-α-acids, and three structurally similar but chemically altered iso-α-acids (known as 'reduced iso-α-acids' and consisting of the rho-, tetrahydro- and hexahydro-iso-α-acid groups) were investigated in whole blood. Pools of blank blood spiked with the four beer-specific ingredient congener groups at two different concentration levels were stored at 20°C, 4°C and -20°C; and extracted in duplicate in weeks 1, 3, 5 and 8, using a previously published method. A loss of 15% of the initial concentration was considered to indicate possible instability and losses greater than 30% demonstrated significant losses. The individual analytes within the four iso-α-acid groups were also measured to determine which iso-α-acids were subject to greater degradation and were responsible for the overall group instability. All four iso-α-acid groups showed significant losses after 8 weeks of storage under room temperature conditions in particularly the natural iso-α-acid group where major losses were observed (96% and 85% losses for low and high concentrations, respectively). Some degradation in all iso-α-acid groups were seen at 4°C samples predominantly due to the 'n' analogs of the groups showing an increased instability in blood. The -20°C storage conditions resulted in minimal changes in concentrations of all analytes. Higher than frozen storage temperatures can result in substantial changes on the stability of the iso-α-acid type groups in blood. The aim of this study was to highlight the stabilities of the IAA analytes in order to assist in the interpretation of IAA in stored blood specimens.

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

    PubMed Central

    Natan, Dity; Nagler, Arnon; Arav, Amir

    2009-01-01

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

  3. Dry film preparation from whole blood, plasma and serum for quantitative infrared diffuse reflectance spectroscopy

    NASA Astrophysics Data System (ADS)

    Bittner, A.; Heise, H. M.

    1998-06-01

    The potential of infrared spectroscopy in the analysis of biotic fluids for the determination of important clinical parameters such as glucose and other blood substrates has been investigated. For this purpose dried films from whole blood, blood plasma and serum were prepared on diffusely reflecting gold-coated substrates from sandpaper of different grades. This enabled measurements in the mid and near infrared spectral ranges by using special diffuse reflectance accessories. The removal of water leads to a considerable enrichment of the fluid constituents. Due to the reduced sample complexity a considerable gain in spectral information is obtained. This is especially valid for measurements in the near infrared where the problems associated with variability in the spectra of aqueous samples due to several parameters, i.e., temperature, electrolyte content etc., are well known. Additionally, mid infrared studies were carried out into the stability of dried samples.

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

    PubMed

    Jones, Alan W

    2006-01-01

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

  5. Comparison of cross-sectional HIV incidence assay results from dried blood spots and plasma

    PubMed Central

    Schlusser, Katherine E.; Pilcher, Christopher; Kallas, Esper G.; Santos, Breno R.; Deeks, Steven G.; Facente, Shelley; Keating, Sheila M.; Busch, Michael P.; Murphy, Gary; Welte, Alex; Quinn, Thomas; Eshleman, Susan H.

    2017-01-01

    Background Assays have been developed for cross-sectional HIV incidence estimation using plasma samples. Large scale surveillance programs are planned using dried blood spot (DBS) specimens for incidence assessment. However, limited information exists on the performance of HIV cross-sectional incidence assays using DBS. Methods The assays evaluated were: Maxim HIV-1 Limiting Antigen Avidity EIA (LAg-Avidity), Sedia HIV-1 BED-Capture EIA (BED-CEIA), and CDC modified BioRad HIV-1/2 Plus O Avidity-based Assay (CDC-BioRad Avidity) using pre-determined cutoff values. 100 matched HIV-1 positive plasma and DBS samples, with known duration of infection, from the Consortium for the Evaluation and Performance of HIV Incidence Assays repository were tested. All assays were run in duplicate. To examine the degree of variability within and between results for each sample type, both categorical and continuous results were analyzed. Associations were assessed with Bland Altman, R2 values and Cohen’s kappa coefficient (ĸ). Results Intra-assay variability using the same sample type was similar for all assays (R2 0.96 to 1.00). The R2 values comparing DBS and plasma results for LAg-Avidity, BED-CEIA, and CDC-BioRad Avidity were 0.96, 0.94, and 0.84, respectively. The concordance and ĸ values between DBS and plasma for all three assays were >87% and >0.64, respectively. The Bland-Altman analysis showed significant differences between plasma and DBS samples. For all three assays, a higher number of samples were classified as recent infections using DBS samples. Conclusions DBS and plasma sample results were highly correlated. However, when compared to plasma, each assay performed somewhat differently in DBS at the lower and higher ends of the dynamic range. DBS samples were more likely to be classified as recently infected by all three assays, which may lead to overestimation of incidence in surveys using performance criteria derived for plasma samples. PMID:28231277

  6. Use of dried blood samples for monitoring hepatitis B virus infection

    PubMed Central

    Lira, Rosalia; Maldonado-Rodriguez, Angelica; Rojas-Montes, Othon; Ruiz-Tachiquin, Martha; Torres-Ibarra, Rocio; Cano-Dominguez, Carlos; Valdez-Salazar, Hilda; Gomez-Delgado, Alejandro; Muñoz, Onofre; Alvarez-Muñoz, Ma-Teresa

    2009-01-01

    Background Hepatitis B virus (HBV) infection is a problem in several regions of the world with limited resources. Blood samples dried on filter paper (DBS) have been successfully used to diagnose and monitor several infectious diseases. In Mexico there is an urgent need for an affordable and easy sampling method for viral load (VL) testing and monitoring of chronic HBV infection. The purpose of this work was to validate the utility of DBS samples for monitoring HBV infection in patients from Mexico City. Methods Matched samples of plasma and DBS on filter paper from 47 HBV infected patients from the Instituto Mexicano del Seguro Social (IMSS), were included. To evaluate the DNA stability and purity from DBS stored at different temperature conditions, samples from ten patients were stored at 4 degree, 25 degree, and 37 degree C for 7 days. After DBS elution and DNA extraction, the purity of these samples was determined measuring the O.D. rate 260/280. The DBS utility for molecular studies was assessed with PCR assays to amplify a 322 bp fragment from the "a" determinant region of the HBV "S" gene. The VL from all samples was determined to evaluate the correlation between plasma and DBS matched samples. Results The quality of the DNA from DBS specimen is not adversely affected by storage at 4 degree, 25 degree and 37 degree C for up 7 days. Statistical ANOVA analyses did not show any significant difference. The same amplification efficiency was observed between DNA templates from samples stored at different temperatures. The Pearson correlation between the VL from DBS and plasma matched samples was 0.93 (p = 0.01). The SD was 1.48 for DBS vs.1.32 for Plasma, and an average of log10 copies/mL of 5.32 vs. 5.53. ANOVA analysis did not show any statistically significant difference between the analyzed groups (p = 0.92). Conclusion The results provide strong evidence that the isolation and quantification of DNA-HBV from DBS is a viable alternative for patient monitoring

  7. Liquid chromatography-mass spectrometric determination of losartan and its active metabolite on dried blood spots.

    PubMed

    Rao, R Nageswara; Raju, S Satyanarayana; Vali, R Mastan; Sankar, G Girija

    2012-08-01

    A simple and rapid quantitative bioanalytical liquid chromatography-tandem mass spectrometric (LC-MS/MS) method for simultaneous determination of losartan and its active metabolite, losartan carboxylic acid on rat dried blood spots was developed and validated as per regulatory guidelines. Losartan and its metabolite were extracted from dried blood spots using 50% aqueous methanol and separated on Waters XTerra(®) RP18 (250 mm × 4.6 mm, 5 μm) column using mobile phase composed of 40% acetonitrile and 60% aqueous ammonium acetate (10mM). The eluents were monitored using ESI tandem mass spectrometric detection with negative polarity in MRM mode using ion transitions m/z 421.2→179.0, m/z 435.3→157.0 and m/z 427.3→193.0 for losartan, losartan carboxylic acid and Irbesartan (internal standard), respectively. The method was validated over the linear range of 1-200 ng/mL and 5-1000 ng/mL with lower limits of quantification of 1.0 ng/mL and 5.0 ng/mL for losartan and losartan carboxylic acid, respectively. Inter and intra-day precision and accuracy (Bias) were below 5.96% and between -2.8 and 1.5%, respectively. The mean recoveries of the analytes from dried blood spots were between 89% and 97%. No significant carry over and matrix effects were observed. The stability of stock solution, whole blood, dried blood spot and processed samples were tested under different conditions and the results were found to be well within the acceptable limits. Additional validation parameters such as influence of hematocrit and spot volume were also evaluated and found to be well within the acceptable limits.

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

    PubMed Central

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

    2014-01-01

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

  9. Development of an UPLC-MS/MS method for the determination of antibiotic ertapenem on dried blood spots.

    PubMed

    la Marca, Giancarlo; Giocaliere, Elisa; Villanelli, Fabio; Malvagia, Sabrina; Funghini, Silvia; Ombrone, Daniela; Filippi, Luca; De Gaudio, Marina; De Martino, Maurizio; Galli, Luisa

    2012-03-05

    Ertapenem (Invanz) is a newly developed carbapenem β-lactam antimicrobial agent. The drug usage in pediatric age needs an accurate drug monitoring for effective patient management. The aim of this study was to evaluate the use of dried blood spot (DBS) specimens to measure ertapenem concentration during treatment. The analysis was performed by UPLC-MS/MS operating in multiple reaction monitoring (MRM) mode. The calibration curve in matrix was linear in the concentration range of 0.5-100 mg/L with correlation coefficient value higher than 0.997. Performance parameters of this method like lower limit of detection (LLOD, 0.2 mg/L), lower limit of quantification (LLOQ, 0.5 mg/L), matrix effect (20%), intra- and inter-day imprecision (CV within than 15%) and accuracy (between 94 and 155%) of drug concentrations have been evaluated. The drug stability at different temperatures was tested for one month, to evaluate the risks of sample delivery at different climatic conditions. The reported method allows now ertapenem analysis and offers many advantages for patients including the possibility of collecting samples at home. This new assay is both precise and accurate and is especially suitable for therapeutic drug monitoring and pharmacokinetic studies in neonates in whom obtaining larger blood samples is not convenient or possible.

  10. Rapid DNA extraction protocol for detection of alpha-1 antitrypsin deficiency from dried blood spots by real-time PCR.

    PubMed

    Struniawski, R; Szpechcinski, A; Poplawska, B; Skronski, M; Chorostowska-Wynimko, J

    2013-01-01

    The dried blood spot (DBS) specimens have been successfully employed for the large-scale diagnostics of α1-antitrypsin (AAT) deficiency as an easy to collect and transport alternative to plasma/serum. In the present study we propose a fast, efficient, and cost effective protocol of DNA extraction from dried blood spot (DBS) samples that provides sufficient quantity and quality of DNA and effectively eliminates any natural PCR inhibitors, allowing for successful AAT genotyping by real-time PCR and direct sequencing. DNA extracted from 84 DBS samples from chronic obstructive pulmonary disease patients was genotyped for AAT deficiency variants by real-time PCR. The results of DBS AAT genotyping were validated by serum IEF phenotyping and AAT concentration measurement. The proposed protocol allowed successful DNA extraction from all analyzed DBS samples. Both quantity and quality of DNA were sufficient for further real-time PCR and, if necessary, for genetic sequence analysis. A 100% concordance between AAT DBS genotypes and serum phenotypes in positive detection of two major deficiency S- and Z- alleles was achieved. Both assays, DBS AAT genotyping by real-time PCR and serum AAT phenotyping by IEF, positively identified PI*S and PI*Z allele in 8 out of the 84 (9.5%) and 16 out of 84 (19.0%) patients, respectively. In conclusion, the proposed protocol noticeably reduces the costs and the hand-on-time of DBS samples preparation providing genomic DNA of sufficient quantity and quality for further real-time PCR or genetic sequence analysis. Consequently, it is ideally suited for large-scale AAT deficiency screening programs and should be method of choice.

  11. Rapid and sensitive LC-MS/MS method for the analysis of antibiotic linezolid on dried blood spot.

    PubMed

    la Marca, Giancarlo; Villanelli, Fabio; Malvagia, Sabrina; Ombrone, Daniela; Funghini, Silvia; De Gaudio, Marina; Fallani, Stefania; Cassetta, Maria Iris; Novelli, Andrea; Chiappini, Elena; de Martino, Maurizio; Galli, Luisa

    2012-01-01

    Linezolid is a new drug from the oxazolidinone class of antibiotics used against mycobacteria and multi-drug resistant (MDR) Gram-positive bacterial infections, which may are also glycopeptide-resistant. The drug usage in pediatric age needs an accurate drug monitoring for effective patient management. The aim of this study was to evaluate the use of dried blood spot (DBS) specimens to determinate linezolid levels during treatment. Advantages of DBS include short collection time, low invasiveness, ease and low cost of sample collection, transport and storage. The analysis was performed in LC-MS/MS operating in positive ion mode and multiple reaction monitoring (MRM) mode. The calibration curve in matrix was linear in the concentration range of 1-100 mg/L with correlation coefficient value of 0.9987. Intraday and interday coefficients of variation were within 3.6% and 13.0%, respectively. We also tested the thermal and temporal drug stability in dried blood spots at four different temperatures to evaluate the risks of sample delivery in different conditions. The short term stability studies showed that linezolid concentration remained stable for at least one month under all the conditions tested. This new assay has favorable characteristics being highly precise and accurate and allows a fast linezolid analysis with a total run time 22 min long, in gradient analysis. Concentration data for plasma and DBS samples from patients after treatment were compared showing a good correlation. Correlation between DBS data and serum samples measured by HPLC-UV was satisfactory. The benefit for patients is the ability to monitor the treatment with a simple and convenient sample collection at home.

  12. EBF recommendation on the validation of bioanalytical methods for dried blood spots.

    PubMed

    Timmerman, Philip; White, Steve; Globig, Susanne; Lüdtke, Silke; Brunet, Leonarda; Smeraglia, John

    2011-07-01

    Over the last few years bioanalysts, pharmacokineticists and clinical investigators have rediscovered the technique of dried blood spots. The revival has provided pharmaceutical R&D a wealth of opportunities to optimize the drug-discovery and development process with respect to animal and patient ethics, new scientific insights and costs savings. On the bioanalytical front, multiple experiments have been performed and a lot of experience has been gained. Nevertheless, the technique still has a number of bioanalytical challenges. The European Bioanalysis Forum discussed the advantages and hurdles of the technique and summarized their current thinking in a recommendation on the validation of bioanalytical methods for dried blood spots, which can be used as a cornerstone for further discussions and experiments.

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

    PubMed

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

    2013-06-01

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

  14. Japanese reference panel of blood specimens for evaluation of hepatitis C virus RNA and core antigen quantitative assays.

    PubMed

    Murayama, Asako; Sugiyama, Nao; Watashi, Koichi; Masaki, Takahiro; Suzuki, Ryosuke; Aizaki, Hideki; Mizuochi, Toshiaki; Wakita, Takaji; Kato, Takanobu

    2012-06-01

    An accurate and reliable quantitative assay for hepatitis C virus (HCV) is essential for measuring viral propagation and the efficacy of antiviral therapy. There is a growing need for domestic reference panels for evaluation of clinical assay kits because the performance of these kits may vary with region-specific genotypes or polymorphisms. In this study, we established a reference panel by selecting 80 donated blood specimens in Japan that tested positive for HCV. Using this panel, we quantified HCV viral loads using two HCV RNA kits and five core antigen (Ag) kits currently available in Japan. The data from the two HCV RNA assay kits showed excellent correlation. All RNA titers were distributed evenly across a range from 3 to 7 log IU/ml. Although the data from the five core Ag kits also correlated with RNA titers, the sensitivities of individual kits were not sufficient to quantify viral load in all samples. As calculated by the correlation with RNA titers, the theoretical lower limits of detection by these core Ag assays were higher than those for the detection of RNA. Moreover, in several samples in our panel, core Ag levels were underestimated compared to RNA titers. Sequence analysis in the HCV core region suggested that polymorphisms at amino acids 47 to 49 of the core Ag were responsible for this underestimation. The panel established in this study will be useful for estimating the quality of currently available and upcoming HCV assay kits; such quality control is essential for clinical usage of these kits.

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

    PubMed Central

    Mössner, Belinda K; Staugaard, Benjamin; Jensen, Janne; Lillevang, Søren Thue; Christensen, Peer B; Holm, Dorte Kinggaard

    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 centers, a prison and outpatient clinics and included blood donors as negative controls. Five drops of finger capillary blood were spotted on filter paper, and a venous blood sample was obtained. The samples were analyzed for HBsAg, anti-HBc, anti-HBs, anti-HCV, and anti-HIV levels as well as subjected to a combined nucleic acid test (NAT) for HBV DNA, HCV RNA and HIV RNA. RESULTS Samples from 404 subjects were screened (85 CHB, 116 CHC, 114 HIV and 99 blood donors). DBS had a sensitivity of > 96% and a specificity of > 98% for the detection of all three infections. NAT testing did not improve sensitivity, 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 infections outside of the health care system. PMID:27672281

  16. Feline reference intervals for the Sysmex XT-2000iV and the ProCyte DX haematology analysers in EDTA and CTAD blood specimens.

    PubMed

    Granat, Fanny; Geffré, Anne; Bourgès-Abella, Nathalie; Mortier, Jeremy; Théron, Marie-Laure; Fauchon, Emilie; Braun, Jean-Pierre; Trumel, Catherine

    2014-06-01

    Laser-based haematology analysers are routinely used in veterinary clinical pathology laboratories, and are available to practitioners. However, feline haematological reference intervals (RIs) determined according to international recommendations are, to our knowledge, not available. Furthermore, platelet count RI is difficult to establish in cats because of the frequent occurrence of platelet aggregation in blood specimens. The purpose of this study was to establish feline haematological RIs with the Sysmex XT-2000iV and ProCyte DX analysers, in ethylenediamine tetra-acetic acid (EDTA) and in citrate, theophylline, adenosine and dipyridamole (CTAD), which is a combination of anticoagulants limiting platelet aggregation. Blood specimens from 120 healthy cats were analysed in duplicate, and the degree of platelet aggregation was assessed on blood smears. After exclusion of inadequate specimens, 81 sets of results (from 44 males and 37 females, aged from 6 to 116 months) were available for the determination of RIs by the non-parametric method. The effects of the anticoagulant, analyser and aggregation score were assessed. When the aggregation effect was significant, the RIs were determined using the subgroup of blood specimens with no or little aggregation. The effects of sex, age and weight were also investigated, but were moderate. The different RIs obtained with the Sysmex XT-2000iV and ProCyte DX analysers, and the two anticoagulants, were very similar to previous RIs established in EDTA with the ADVIA 120, another laser-based analyser, except for the platelet count in CTAD specimens. Its lower reference limit was higher in CTAD vs EDTA specimens, which confirms the interest in this anticoagulant in cats.

  17. The development and validation of a fast and robust dried blood spot based lipid profiling method to study infant metabolism.

    PubMed

    Koulman, Albert; Prentice, Philippa; Wong, Max C Y; Matthews, Lee; Bond, Nicholas J; Eiden, Michael; Griffin, Julian L; Dunger, David B

    2014-01-01

    Early life exposures and metabolic programming are associated with later disease risk. In particular lipid metabolism is thought to play a key role in the development of the metabolic syndrome and insulin resistance in later life. Investigative studies of metabolic programming are limited by the ethics and practicalities of sample collection in small infants. Dried blood spots on filter paper, derived from heel pricks are considered as the most suitable option for this age group. We validated a novel lipid profiling method, based on high resolution mass spectrometry to successfully determine the lipid composition of infants using dried blood spots. The spotting and air drying of blood on paper has noticeable effects on many of the lipids, leading to lipid oxidation and hydrolysis, which demand careful interpretation of the obtained data. We compared the lipid profiles from plasma or whole blood samples and the results from dried blood spots to determine if these revealed the same inter-subject differences. The results from dried blood spots were no less reproducible than other lipid profiling methods which required comparatively larger sample volumes. Therefore, lipid profiles obtained from dried blood spots can be successfully used to monitor infancy lipid metabolism and we show significant differences in the lipid metabolism of infants at age 3 versus 12 months.

  18. Effects of cold dry air nasal stimulation on airway mucosal blood flow in humans.

    PubMed

    Le Merre, C; Isber, J; Chediak, A D; Wanner, A

    2003-10-01

    Several studies have demonstrated that nasal challenges can induce reflex responses in the respiratory system. Some authors have described bronchoconstriction and modification of the pattern of breathing following nasal challenges by irritants and cold air. We propose to determine the effect of nasal stimulation with cold dry air on airway mucosal blood flow (Qaw) in the proximal tracheal bronchial tree of healthy humans. Nine healthy subjects participated in the study. Baseline measurement Qaw, nasal airway resistance (NAR) and airway caliber by specific airways conductance (SGaw) were followed by nasal challenge with cold dry air. Qaw, NAR and Sgaw were determined after the challenge. In those subjects in which a significant decline in Qaw was recorded the protocol was repeated after pretreatment with nasal anesthesia using topical lidocaine. Cold dry air challenge produced a significant decrease in mean Qaw for the nine subjects and this response was abolished by pretreatment with nasal anesthesia using topical lidocaine. There was no significant change in Sgaw and NAR after the challenge and topical lidocaine anesthesia. Our data indicates that nasal stimulation with cold dry air leads to a reduction in Qaw and that this effect may be mediated by a nasal reflex.

  19. Direct polymerase chain reaction (PCR) from human whole blood and filter-paper-dried blood by using a PCR buffer with a higher pH.

    PubMed

    Bu, Ying; Huang, Huan; Zhou, Guohua

    2008-04-15

    We described a novel approach to directly amplify genomic DNA from whole blood and dried blood spotted on filter paper without any DNA isolation by using the PCR buffer with a higher pH, which was optimized as pH 9.1-9.6. Direct PCR on blood treated with various anticoagulants showed that the buffer worked well with the blood treated by citrate, EDTA, or heparinate. DNA fragments with different lengths could be efficiently amplified directly from various forms of blood samples. By coupling the buffer with tetra-PCR, a "true" single-tube genotyping was realized by using whole blood or paper-dried blood as starting material.

  20. Molecular Epidemiology of Macrolide-Resistant Isolates of Streptococcus pneumoniae Collected from Blood and Respiratory Specimens in Norway

    PubMed Central

    Littauer, P.; Sangvik, M.; Caugant, D. A.; Høiby, E. A.; Simonsen, G. S.; Sundsfjord, A.

    2005-01-01

    Norway has a low prevalence of antimicrobial resistance, including macrolide-resistant Streptococcus pneumoniae (MRSP). In a nationwide surveillance program, a total of 2,200 S. pneumoniae isolates were collected from blood cultures and respiratory tract specimens. Macrolide resistance was detected in 2.7%. M-type macrolide resistance was found in 60% of resistant isolates, and these were mainly mef(A)-positive, serotype-14 invasive isolates. The erm(B)-encoded macrolide-lincosamide-streptogramin B (MLSB) type dominated among the noninvasive isolates. One strain had an A2058G mutation in the 23S rRNA gene. Coresistance to other antibiotics was seen in 96% of the MLSB-type isolates, whereas 92% of the M-type isolates were susceptible to other commonly used antimicrobial agents. Serotypes 14, 6B, and 19F accounted for 84% of the macrolide-resistant isolates, with serotype 14 alone accounting for 67% of the invasive isolates. A total of 29 different sequence types (STs) were detected by multilocus sequence typing. Twelve STs were previously reported international resistant clones, and 75% of the macrolide-resistant isolates had STs identical or closely related to these clones. Eleven isolates displayed 10 novel STs, and 7/11 of these “Norwegian strains” coexpressed MLSB and tetracycline resistance, indicating the presence of Tn1545. The invasive serotype-14 isolates were all classified as ST9 or single-locus variants of this clone. ST9 is a mef-positive M-type clone, commonly known as England14-9, reported from several European countries. These observations suggest that the import of major international MRSP clones and the local spread of Tn1545 are the major mechanisms involved in the evolution and dissemination of MRSP in Norway. PMID:15872231

  1. Silica coated paper substrate for paper-spray analysis of therapeutic drugs in dried blood spots.

    PubMed

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

    2012-01-17

    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 that 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 Whatman ET31 paper used for blood cards. Analysis using a hand-held 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.

  2. A Simple Method to Quantitate IP-10 in Dried Blood and Plasma Spots

    PubMed Central

    Aabye, Martine G.; Eugen-Olsen, Jesper; Werlinrud, Anne Marie; Holm, Line Lindebo; Tuuminen, Tamara; Ravn, Pernille; Ruhwald, Morten

    2012-01-01

    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 (r2>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. PMID:22761744

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

    NASA Astrophysics Data System (ADS)

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

    2013-08-01

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

  4. [Investigation into vegetative regulation of blood circulation during 7-day dry immersion].

    PubMed

    Eshmanova, A K; Luchinskaia, E S; Baevskiĭ, R M

    2008-01-01

    The paper presents the results of investigations into vegetative regulation of blood circulation and regulation modification by peroral amlodipine and myostimulation during 7-d dry immersion. It was shown that in immersion vegetative regulation readjusted towards predominance of the sympathetic mechanisms. Myostimulation and peroral amlodipine modified regulation substantially mobilizing high level suprasegmentary structures. It should be noted that amlodipine and myostimulation differ in mechanisms of their action on central regulation. Pharmaceutical intervention seems to have a more complex and varying effect on people including side-effects. Presumably, it was the cause of poor orthostatic tolerance of several test-subjects.

  5. Comparative evaluation of the role of single and multiple blood specimens in the outcome of blood cultures using BacT/ALERT 3D (automated) blood culture system in a tertiary care hospital

    PubMed Central

    Elantamilan, D.; Lyngdoh, Valarie Wihiwot; Khyriem, Annie B.; Rajbongshi, Jyotismita; Bora, Ishani; Devi, Surbala Thingujam; Bhattacharyya, Prithwis; Barman, Himesh

    2016-01-01

    Introduction: Bloodstream infection (BSI) is a leading cause of mortality in critically ill patients. The mortality directly attributable to BSI has been estimated to be around 16% and 40% in general hospital population and Intensive Care Unit (ICU) population, respectively. The detection rate of these infections increases with the number of blood samples obtained for culture. The newer continuous monitoring automated blood culture systems with enhanced culture media show increased yield and sensitivity. Hence, we aimed at studying the role of single and multiple blood specimens from different sites at the same time in the outcome of automated blood culture system. Materials and Methods and Results: A total of 1054 blood culture sets were analyzed over 1 year, the sensitivity of one, two, and three samples in a set was found to be 85.67%, 96.59%, and 100%, respectively, which showed a statistically significant difference (P < 0.0001). Similar findings were seen in few more studies, however, among individual organisms in contrast to other studies, the isolation rates of Gram-positive bacteria were less than that of Gram-negative Bacilli with one (or first) sample in a blood culture set. In our study, despite using BacT/ALERT three-dimensional continuous culture monitoring system with FAN plus culture bottles, 15% of positive cultures would have been missed if only a single sample was collected in a blood culture set. Conclusion: The variables like the volume of blood and number of samples collected from different sites still play a major role in the outcome of these automated blood culture systems. PMID:27688629

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

    PubMed Central

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

    2016-01-01

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

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

    PubMed Central

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

    2014-01-01

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

  8. Rapid screening assay of congenital adrenal hyperplasia by measuring 17 alpha-hydroxyprogesterone with high-performance liquid chromatography/electrospray ionization tandem mass spectrometry from dried blood spots.

    PubMed

    Lai, Chien-Chen; Tsai, Chang-Hai; Tsai, Fuu-Jen; Wu, Jer-Yuarn; Lin, Wei-De; Lee, Cheng-Chun

    2002-01-01

    A rapid, simple, and specific method was developed for the diagnosis of congenital adrenal hyperplasia (CAH) from dried blood spots on newborn screening cards based on high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS). The usefulness of 17 alpha-hydroxyprogesterone (17 OH-P) determination on dried filter-paper blood samples from patients with CAH caused by 21-hydroxylase deficiency was evaluated. The LC/MS/MS detection of 17 OH-P was rapid, <4 min. The intra- and interday accuracy and precision of the method were <7%. Our procedure maintained good linearities (R(2) > 0.992) and recovery rate (>83%). We used this new method to directly determine the 17 OH-P levels in dried blood specimens from abnormal children of various ages, with a detection limit of 20 ng/ml (approximately 240 pg), to avoid the time-consuming derivatization steps required by the gas-chromatography/mass spectrometry (GC/MS) method. Four dried filter-paper blood samples of CAH patients (three girls and one boy, 1-14 years old) were all quantified in an LC/MS/MS study and revealed high 17 OH-P levels (>90 ng/ml). After treatment, all of the elevated 17 OH-P levels either decreased or disappeared. Compared with CAH patients, 17 OH-P was nearly undetectable (<20 ng/ml) in the normal infants by LC/MS/MS. This LC/MS/MS assay is not only useful for both diagnosis and monitoring of treatment of CAH in all other age groups, it also can be used as a screening test for CAH infants. In this study, we provided the first data on 17 OH-P in dried blood specimens affected with CAH using HPLC/ESI-MS/MS.

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

    USGS Publications Warehouse

    Flannery, Jennifer A.; Poore, Richard Z.

    2013-01-01

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

  10. Rapid genotyping of cytomegalovirus in dried blood spots by multiplex real-time PCR assays targeting the envelope glycoprotein gB and gH genes.

    PubMed

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

    2012-02-01

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

  11. Bridging the Gap between Sample Collection and Laboratory Analysis: Using Dried Blood Spots to Identify Human Exposure to Chemical Agents.

    PubMed

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

    2016-05-13

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

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

    NASA Astrophysics Data System (ADS)

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

    2016-05-01

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

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

    PubMed

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

    2016-05-01

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

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

    PubMed Central

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

    2016-01-01

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

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

    PubMed

    Thilsing-Hansen, T; Jørgensen, R J; Enemark, J M; Zelvyte, R; Sederevicius, A

    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 the expected date of calving (exp. 1 and 2) and one week after calving (exp. 3 and 4). The zeolite supplementation significantly increased the mean serum Ca level on the day of calving. The efficiency of the zeolite supplementation to prevent hypocalcaemia (serum Ca < 2.00 mmol/l) on the day of calving was calculated. Efficiencies varied ranging from 3 to 100%, with a mean efficiency of 58%. The zeolite-calcium ratio (g of zeolite per cow per day/g of dietary calcium per cow per day) was calculated in each experiment. From the results it seemed, that zeolite-calcium ratios below 5 did not effectively prevent parturient hypocalcaemia, whereas ratios of 10 to 20 proved very efficient in preventing hypocalcaemia. There was apparently no additional effect from feeding zeolite for 4 instead of 2 weeks prepartum. Feeding zeolite in the dry period significantly decreased plasma phosphate before as well as after calving. The phosphate level was normalized within one week after calving. Plasma magnesium was significantly lower among the experimental cows on the day of calving

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

    PubMed Central

    Salazar-Gonzalez, Jesus F.; Salazar, Maria G.; Tully, Damien C.; Ogilvie, Colin B.; Learn, Gerald H.; Allen, Todd M.; Heath, Sonya L.; Goepfert, Paul; Bar, Katharine J.

    2016-01-01

    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 -20°C 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. PMID:27819061

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

    PubMed

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

    2016-02-05

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

  18. The Use of Dried Blood Spot Sampling in the National Social Life, Health, and Aging Project

    PubMed Central

    McDade, Thomas W.

    2009-01-01

    Objectives This paper describes the methods used for and issues associated with collection and analysis of dried blood spot (DBS) samples for the National Social Life, Health, and Aging Project and provides the basic distributions of the resulting analytes. Methods DBSs from capillary finger sticks were collected by nonmedically trained interviewers from 2,044 individuals, aged 57–85 years. The quality and quantity of DBS samples were evaluated to allow for analysis of interviewer performance. Levels of C-reactive protein, antibodies to the Epstein–Barr virus, hemoglobin, and glycosylated hemoglobin were assayed using various analytic methods. Results Cooperation rate for DBS collection was 84.5%, with 99% of the cards yielding enough sample for at least one analysis. The distribution, mean, and standard deviation of the analytes obtained from DBSs are also presented in this paper. Conclusions The high cooperation rate and quality of the spots collected suggest that the collection of DBSs in population-based research is a feasible and viable alternative to venous blood draws. The relative ease of sample collection, transport, and storage are significant benefits. Care should be taken, however, when comparing results from analysis of DBS samples with those obtained from serum or plasma samples. PMID:19244547

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

    PubMed

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

    2014-08-05

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

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

    PubMed

    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.

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

    PubMed Central

    Batterman, Stuart; Chernyak, Sergei

    2014-01-01

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

  2. Tenofovir, Emtricitabine, and Tenofovir Diphosphate in Dried Blood Spots for Determining Recent and Cumulative Drug Exposure

    PubMed Central

    Castillo-Mancilla, Jose R.; Zheng, Jia-Hua; Rower, Joseph E.; Meditz, Amie; Gardner, Edward M.; Predhomme, Julie; Fernandez, Caitlin; Langness, Jacob; Kiser, Jennifer J.; Bushman, Lane R.

    2013-01-01

    Abstract Tenofovir (TFV) disoproxil fumarate (TDF)±emtricitabine (FTC) are widely used for HIV treatment and chemoprophylaxis, but variable adherence may lead to suboptimal responses. Methods that quantify adherence would allow for interventions to improve treatment and prevention outcomes. Our objective was to characterize the pharmacokinetics of TFV-diphosphate (TFV-DP) and FTC-triphosphate (FTC-TP) in red blood cells (RBCs) and peripheral blood mononuclear cells (PBMCs); to extend the RBC analysis to dried blood spots (DBSs); and to model how RBC/DBS monitoring could inform recent and cumulative drug exposure/adherence. Blood samples were collected from 17 HIV-negative adults at 5 visits over a 30-day pharmacokinetics study of daily oral TDF/FTC. Dosing was discontinued on day 30 and blood was collected on days 35, 45, and 60 during the washout period. Plasma/RBCs/PBMCs/DBSs were all quantified by liquid chromatography/tandem mass spectrometry. DBSs were paired with RBCs and plasma for comparisons. The median (interquartile range) RBC TFV-DP half-life was 17.1 (15.7–20.2) versus 4.2 (3.7–5.2) days in PBMCs. At steady state, TFV-DP was 130 fmol/106 RBCs versus 98 fmol/106 PBMCs. FTC-TP was not quantifiable in most RBC samples. TFV-DP in RBCs versus DBSs yielded an r2=0.83. TFV-DP in DBSs was stable at −20°C. Simulations of TFV-DP in RBCs/DBSs, when dosed from one to seven times per week, demonstrated that each dose per week resulted in an average change of approximately 19 fmol/106 RBCs and 230 fmol/punch. TFV and FTC in plasma versus DBSs was defined by y=1.4x; r2=0.96 and y=0.8x; r2=0.99, respectively. We conclude that DBSs offer a convenient measure of recent (TFV/FTC) and cumulative (TFV-DP in RBCs) drug exposure with potential application to adherence monitoring. PMID:22935078

  3. Field Evaluation of the InBios Chagas Detect Plus Rapid Test in Serum and Whole-Blood Specimens in Bolivia

    PubMed Central

    Shah, Vishal; Ferrufino, Lisbeth; Gilman, Robert H.; Ramirez, Margot; Saenza, Eliana; Malaga, Edith; Sanchez, Gerardo; Okamoto, Emi E.; Sherbuck, Jacqueline E.; Clark, Eva H.; Galdos-Cardenas, Gerson; Bozo, Ricardo; Flores-Franco, Jorge Luis; Colanzi, Rony; Verastegui, Manuela

    2014-01-01

    Trypanosoma cruzi causes Chagas disease, which affects an estimated 7 million to 8 million people. Chagas disease is endemic throughout Latin America, with the highest prevalence in Bolivia. Conventional diagnosis requires a well-equipped laboratory with experienced personnel. We evaluated the Chagas Detect Plus (CDP) (InBios, Seattle, WA), a rapid immunochromatographic assay for IgG antibodies to T. cruzi. CDP performance was compared to infection status based on results obtained by indirect hemagglutination assay, immunofluorescent-antibody test, and enzyme-linked immunosorbent assay. Confirmed infection required positive results by at least 2 conventional assays. We used specimens from adults of both sexes in a general hospital in the city of Santa Cruz and from pregnant women in a hospital and children in villages in the Bolivian Chaco, an area of hyperendemicity. CDP was performed in paired whole-blood and serum specimens from 385 individuals in the two hospital studies and in 200 serum specimens from the community study. CDP showed sensitivities/specificities of 96.2% (95% confidence interval, 92.7 to 98.4)/98.8% (95.9 to 99.9) in whole blood and 99.3% (97.5 to 99.9)/96.9% (94.2 to 98.6) in serum, with no differences by sex, age group, or study site. CDP showed excellent sensitivity and specificity in our study population, comparable to those of conventional serology. The test is reliable for field surveys, requires no laboratory equipment, and performed well in serum and whole blood. The CDP could also be used for accurate maternal screening to identify neonates at risk of congenital transmission. CDP performance data in diverse geographic areas are needed to strengthen the evidence base for its use. PMID:25274804

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

    PubMed Central

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

    2015-01-01

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

  5. Blood Culture Collection through Peripheral Intravenous Catheters Increases the Risk of Specimen Contamination among Adult Emergency Department Patients

    PubMed Central

    Self, Wesley H.; Speroff, Theodore; McNaughton, Candace D.; Wright, Patty W.; Miller, Geraldine; Johnson, James G.; Daniels, Titus L.; Talbot, Thomas R.

    2017-01-01

    Five hundred five blood cultures collected through a peripheral intravenous catheter (PIV) in an emergency department were matched to cultures obtained by dedicated venipuncture from the same patient within 10 minutes. The relative risk of contamination for cultures collected through PIVs compared with dedicated venipuncture was 1.83 (95% confidence interval, 1.08–3.11). PMID:22476282

  6. Blood culture collection through peripheral intravenous catheters increases the risk of specimen contamination among adult emergency department patients.

    PubMed

    Self, Wesley H; Speroff, Theodore; McNaughton, Candace D; Wright, Patty W; Miller, Geraldine; Johnson, James G; Daniels, Titus L; Talbot, Thomas R

    2012-05-01

    Five hundred five blood cultures collected through a peripheral intravenous catheter (PIV) in an emergency department were matched to cultures obtained by dedicated venipuncture from the same patient within 10 minutes. The relative risk of contamination for cultures collected through PIVs compared with dedicated venipuncture was 1.83 (95% confidence interval, 1.08-3.11).

  7. Islet Autoantibody Measurements from Dried Blood Spots on Filter Paper Strongly Correlate to Serum Levels

    PubMed Central

    Simmons, Kimber M.; Alkanani, Aimon K.; McDaniel, Kristen A.; Goyne, Christopher; Miao, Dongmei; Zhao, Zhiyuan; Yu, Liping; Michels, Aaron W.

    2016-01-01

    Type 1 diabetes (T1D) is increasing in incidence and predictable with measurement of serum islet autoantibodies (iAb) years prior to clinical disease onset. Identifying iAb positive individuals reduces diabetic ketoacidosis and identifies individuals for T1D prevention trials. However, large scale screening for iAb remains challenging as assays have varying sensitivities and specificities, insulin autoantibodies remain difficult to measure and venipuncture is generally required to obtain serum. We developed an approach to reliably measure all four major iAb, including insulin autoantibodies, from dried blood spots (DBS) on filter-paper. By spiking iAb positive serum into iAb negative whole blood in a dose titration, we optimized the conditions for autoantibody elution from filter paper as measured by fluid phase radioimmunoassays. After assessing stability of measuring iAb from DBS over time, we then screened iAb from DBS and the corresponding serum in new-onset T1D (n = 52), and controls (n = 72) which included first-degree relatives of T1D patients. iAb measured from eluted DBS in new-onset T1D strongly correlated with serum measurements (R2 = 0.96 for mIAA, GADA = 0.94, IA-2A = 0.85, ZnT8A = 0.82, p<0.01 for each autoantibody). There were no false positives in control subjects, and 5/6 with previously unknown iAb positivity in sera were detected using DBS. With further validation, measuring iAb from DBS can be a reliable method to screen for T1D risk. PMID:27846247

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

    PubMed

    Capiau, Sara; Wilk, Leah S; Aalders, Maurice C G; Stove, Christophe P

    2016-06-21

    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 quantitation remains undoubtedly the most widely discussed one. Previously, we developed a method to derive the approximate hematocrit from a nonvolumetrically applied DBS based on its potassium content. Although this method yielded good results and was straightforward to perform, it was also destructive and required sample preparation. Therefore, we now developed a nondestructive method which allows to predict the hematocrit of a DBS based on its hemoglobin content, measured via noncontact diffuse reflectance spectroscopy. The developed method was thoroughly validated. A linear calibration curve was established after log/log transformation. The bias, intraday and interday imprecision of quality controls at three hematocrit levels and at the lower and upper limit of quantitation (0.20 and 0.67, respectively) were less than 11%. In addition, the influence of storage and the volume spotted was evaluated, as well as DBS homogeneity. Application of the method to venous DBSs prepared from whole blood patient samples (n = 233) revealed a good correlation between the actual and the predicted hematocrit. Limits of agreement obtained after Bland and Altman analysis were -0.076 and +0.018. Incurred sample reanalysis demonstrated good method reproducibility. In conclusion, mere scanning of a DBS suffices to derive its approximate hematocrit, one of the most important variables in DBS analysis.

  9. Exploring existing and deliberated community perspectives of newborn screening: informing the development of state and national policy standards in newborn screening and the use of dried blood spots

    PubMed Central

    Muchamore, Ian; Morphett, Luke; Barlow-Stewart, Kristine

    2006-01-01

    Objective Since the 1960s newborn screening (NBS) for several rare and serious disorders has been in place across Australia. Testing of a simple blood spot now enables the early detection of over 30 conditions. Policies across Australian states have diverged in some aspects of NBS, especially in the retention and further use of dried blood spots collected as part of the screening and attempts are underway to bring some further national consistency. Whilst this has initiated debate amongst health professionals and policy makers there is limited empirical evidence of wider community attitudes to such issues. Methods This research has explored the range and depth of views held by the wider community in New South Wales through moderated small group discussions. It has also assessed the range and depth of responses where the groups are reconvened after being given further information. Results The findings suggest that there is limited community awareness of the public health importance of NBS and especially that resulting biological samples are stored. Members of the wider community presented with opportunities to consider current procedures and policies appear reassured and to have high levels of trust. However there are clearly some groups who have concerns with the storage of dried blood spot specimens and perceive that these may be abused. Policy implications and conclusion The findings will inform health professionals and policy makers as to the perceived benefits and future challenges NBS raises for the wider community. The findings have implications for improving current communications about NBS, maintaining public confidence and the development of state and national initiatives in genetic health. PMID:17164009

  10. Dried blood spot analysis by digital microfluidics coupled to nanoelectrospray ionization mass spectrometry.

    PubMed

    Shih, Steve C C; Yang, Hao; Jebrail, Mais J; Fobel, Ryan; McIntosh, Nathan; Al-Dirbashi, Osama Y; Chakraborty, Pranesh; Wheeler, Aaron R

    2012-04-17

    Dried blood spot (DBS) samples on filter paper are surging in popularity as a sampling and storage vehicle for a wide range of clinical and pharmaceutical applications. For example, a DBS sample is collected from every baby born in the province of Ontario, Canada, for quantification of approximately one hundred analytes that are used to screen for 28 conditions, including succinylacetone (SA), a marker for hepatorenal tyrosinemia. Unfortunately, the conventional methods used to evaluate DBS samples for newborn screening and other applications are tedious and slow, with limited options for automated analysis. In response to this challenge, we have developed a method to couple digital microfluidics (DMF) to nanoelectrospray ionization mass spectrometry (nESI-MS) for SA quantification in DBS samples. The new system is formed by sandwiching a pulled glass capillary emitter between the two DMF substrates such that the capillary emitter is immobilized without external seals or gaskets. Moreover, we introduce a new feedback control system that enables high-fidelity droplet manipulation across DBS samples without manual intervention. The system was validated by application to on-chip extraction, derivatization, and analysis of SA and other analytes from DBS samples, with comparable performance to gold-standard methods. We propose that the new methods described here can potentially contribute to a new generation of analytical techniques for quantifying analytes in DBS samples for a wide range of applications.

  11. [Direct proteome profiling of human blood serum in the experiment with 5-day dry immersion].

    PubMed

    Pastushkova, L Kh; Pakharukova, N A; Trifonova, O P; Dobrokhotov, I V; Valeeva, O A; Larina, I M

    2011-01-01

    Purpose of the investigation was to determine changes in blood plasma proteome in healthy human subjects (n = 14, 19 to 26 y.o.) in an experiment with dry immersion (DI). Plasma samples were drawn 7 and 2 days before the exposure, on DI days 2, 3 and 5, and on days 1, 3, 7 and 15 after the experiment. Previous to direct MALDI-TOF mass-spectrometric profiling, serum samples were pre-fractionated and enriched with magnetic particles MB WCX (WCX--a weak cation exchanger) on ClinProt (Bruker Daltonics). In each spectrum, 175 MS-peaks were detected on average within the mass range from 1000 to 17,000 Da with the signal/noise ratio = 5. Student's criterion (p < 0.05) was used to define reliable differences between DI and baseline samples from 48 peaks (27.4 % of all the proteome profile peaks). On DI days 2 and 3, growth of peak areas was observed in fragments of complement system proteins C3 and C4, high-molecular kininogen and fibrinogen that can be attributed to organism adaptation to conditions of the experiment. Significant increases of the peak area of apolipoprotein CI (reduced form with segregated threonine and proline) and C4 enzymes of the complement system, and fibrinogen on the first day after the experiment can be related to changes in motor activities of the subjects.

  12. Dried Blood Spot sampling in psychiatry: Perspectives for improving therapeutic drug monitoring.

    PubMed

    Martial, Lisa C; Aarnoutse, Rob E; Mulder, Martina; Schellekens, Arnt; Brüggemann, Roger J M; Burger, David M; Schene, Aart H; Batalla, Albert

    2017-03-01

    Assessment of drug concentrations is indicated to guide dosing of a selected number of drugs used in psychiatry. Conventionally this is done by vena puncture. Novel sampling strategies such as dried blood spot (DBS) sampling have been developed for various drugs, including antipsychotics, antidepressants and mood-stabilizers. DBS sampling is typically performed by means of a finger prick. This method allows for remote sampling, which means that patients are not required to travel to a health care facility. The number of DBS assays for drugs used in psychiatry has increased over the last decade and includes antidepressants (tricyclic and serotonin and/or norepinephrine reuptake inhibitors), mood stabilizers and first- and second-generation antipsychotics. Available assays often comply with analytical validation criteria but are seldom used in routine clinical care. Little attention has been paid to the clinical validation and implementation processes of home sampling. Ideally, not only medicines but also clinical chemistry parameters should be measured within the same sample. This article reflects on the position of DBS remote sampling in psychiatry and provides insight in the requisites of making such a sampling tool successful.

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

    PubMed Central

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

    2015-01-01

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

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

    PubMed Central

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

    2016-01-01

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

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

    PubMed Central

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

    2016-01-01

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

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

    PubMed Central

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

    2015-01-01

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

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

    PubMed

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

    2014-11-01

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

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

    PubMed Central

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

    2014-01-01

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

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

    PubMed

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

    2016-05-01

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

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

    PubMed Central

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

    2016-01-01

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

  1. Determination of salmeterol in dried blood spot using an ionic liquid based dispersive liquid-liquid microextraction coupled with HPLC.

    PubMed

    Hatami, Mehdi; Karimnia, Elham; Farhadi, Khalil

    2013-11-01

    A novel analytical method based on the use of a modified dispersive liquid-liquid microextraction (DLLME) technique by ionic liquids has been developed for the first time. This method was applied for the determination of salmeterol in dried blood spot samples from asthmatic patients followed by HPLC system equipped with fluorescence detector. The main novelty of the present work deals with the analysis of salmeterol in dried blood spot with significant advantages with regard to invasive sampling, volume of blood used (<20 μL), storage and transport of biological materials and requirements for special biohazard arrangements. The effects of significant factors in microextraction procedure including extracting and disperser solvents volume, sample pH and salt contents were investigated. Under optimized conditions, the mean recoveries were higher than 90.0% with linear responses about 1.00-500 ng mL(-1) concentration range. The limit of detection (LOD) was 0.30 ng mL(-1) in S/N of 3. Intra-day (n=6) and inter-day (n=3) assay precision showed relative standard deviation lower than 6.0 and 8.5% for studied drug, respectively. The developed microextraction procedure was simpler and faster than conventional extraction techniques and was omitted hazard organic solvents. Finally, the proposed method is suitable for the analysis of salmeterol in dried blood spots from asthmatic patients for therapeutic drug monitoring purpose.

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

    PubMed

    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.

  3. Detection of IL28B SNP DNA from Buccal Epithelial Cells, Small Amounts of Serum, and Dried Blood Spots

    PubMed Central

    Halfon, Philippe; Ouzan, Denis; Khiri, Hacène; Pénaranda, Guillaume; Castellani, Paul; Oulès, Valerie; Kahloun, Asma; Amrani, Nolwenn; Fanteria, Lise; Martineau, Agnès; Naldi, Lou; Bourlière, Marc

    2012-01-01

    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. PMID:22412970

  4. Stability of dried blood spots for HIV-1 drug resistance analysis.

    PubMed

    Hearps, Anna C; Ryan, Claire E; Morris, Lisa M; Plate, Megan M; Greengrass, Vicki; Crowe, Suzanne M

    2010-03-01

    The wide scale application of dried blood spots (DBS) as a collection tool for low-cost HIV drug resistance testing requires a greater understanding of the accuracy of DBS for genotype analysis and the stability of DBS under various environmental conditions. Analysis of a 50microl DBS via a single amplicon, nested PCR-based in-house assay (the Burnet genotyping assay) showed an average nucleotide concordance of 98.9% with plasma samples, although only 65% of nucleotide mixtures detected in plasma were also detected within DBS. The analysis of three DBS resulted in the detection of a greater number of nucleotide mixtures (72 and 109 mixtures detected within one and three DBS, respectively, n=10). Two DBS extraction protocols (silica particle; NucliSENS, bioMerieux and spin column extraction; High Pure, Roche) were assessed and found to be equivalent (79% and 84% recovery success respectively, n=19). FTA Elute paper (Whatman) was an inferior DBS collection medium compared to Whatman 903 paper. DBS appeared relatively tolerant to multiple freeze/thaw cycles, with 79% of DBS subjected to ten freeze/thaw cycles successfully amplified compared to 93% of DBS defrosted once (n=14). High temperature (37 degrees C) and high humidity (>90%) substantially impaired DBS recovery within two weeks of storage (38%, n=8), whilst storage at -20 degrees C or 4 degrees C adequately preserved DBS for this period (100% recovery, n=8). Therefore, whilst DBS are suitable for HIV drug resistance surveillance, the use of multiple DBS may be required to ensure accurate detection of minor HIV quasispecies and short-term storage of samples at either 4 degrees C or -20 degrees C is recommended.

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

    PubMed Central

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

    2016-01-01

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

  6. A Rapid Screening Test on Dried Blood for the Neonatal Diagnosis of Tyrosinemia Type I

    PubMed Central

    Bodaghkhan, Farahnaz; Geramizadeh, Bita; Rajeh, Abbas Abdollah; Haghighat, Mahmoud; Dehghani, Mohsen; Honar, Naser; Zahmatkeshan, Mojgan; Imanieh, Mohammad-Hadi

    2016-01-01

    Background: Tyrosinemia is an inherited metabolic disorder characterized by elevated levels of tyrosine and its metabolites in plasma. Without treatment, the disease will progress to hepatic and renal failure, so that without liver transplantation will cause death in less than 10 years of age. So, early diagnosis and treatment can be life saving and crucial. It means that with early treatment starting in the neonatal period, the patient can have normal life with very few restrictions in diets containing tyrosine and phenylalanine. Objectives: In this study we wanted to evaluate an easy to perform, rapid and sensitive qualitative test with low cost, as a part of neonatal screening tests to help early diagnosis and treatment of hereditary tyrosinemia. Patients and Methods: In this cross sectional study, during the study period (2013 - 2014), 100 patients were selected. Fifty three (53) of these patients had proven tyrosinemia and the other 47 cases biliary atresia, paucity of intrahepatic bile ducts, cytomegalovirus (CMV) hepatitis, galactosemia and storage diseases. Results: There were 2 false negative and 14 false positive cases of hereditary tyrosinemia (HT-1) in the test. Six cases of biliary atresia, 7 cases of paucity of intrahepatic bile ducts and one patient with cytomegalovirus (CMV) hepatitis were falsely positive with the test. Sensitivity of the test was 96.23%, specificity 71.43%, positive predictive value (PPV) 78.46%, and negative predictive value (NPV) 94.59%. Conclusions: This rapid qualitative test on dried blood sample is an easy, cheap, and feasible method for the screening of hereditary tyrosinemia in neonatal period. PMID:28203327

  7. Virtual Specimens

    NASA Astrophysics Data System (ADS)

    de Paor, D. G.

    2009-12-01

    Virtual Field Trips have been around almost as long as the Worldwide Web itself yet virtual explorers do not generally return to their desktops with folders full of virtual hand specimens. Collection of real specimens on fields trips for later analysis in the lab (or at least in the pub) has been an important part of classical field geoscience education and research for generations but concern for the landscape and for preservation of key outcrops from wanton destruction has lead to many restrictions. One of the author’s favorite outcrops was recently vandalized presumably by a geologist who felt the need to bash some of the world’s most spectacular buckle folds with a rock sledge. It is not surprising, therefore, that geologists sometimes leave fragile localities out of field trip itineraries. Once analyzed, most specimens repose in drawers or bins, never to be seen again. Some end up in teaching collections but recent pedagogical research shows that undergraduate students have difficulty relating specimens both to their collection location and ultimate provenance in the lithosphere. Virtual specimens can be created using 3D modeling software and imported into virtual globes such as Google Earth (GE) where, they may be linked to virtual field trip stops or restored to their source localities on the paleo-globe. Sensitive localities may be protected by placemark approximation. The GE application program interface (API) has a distinct advantage over the stand-alone GE application when it comes to viewing and manipulating virtual specimens. When instances of the virtual globe are embedded in web pages using the GE plug-in, Collada models of specimens can be manipulated with javascript controls residing in the enclosing HTML, permitting specimens to be magnified, rotated in 3D, and sliced. Associated analytical data may be linked into javascript and localities for comparison at various points on the globe referenced by ‘fetching’ KML. Virtual specimens open up

  8. Hemoglobin Variant Analysis via Direct Surface Sampling of Dried Blood Spots Coupled with High-Resolution Mass Spectrometry

    PubMed Central

    2011-01-01

    Hemoglobinopathies are the most common inherited disorders. Newborn blood screening for clinically significant hemoglobin variants, including sickle (HbS), HbC, and HbD, has been adopted in many countries as it is widely acknowledged that early detection improves the outcome. We present a method for determination of Hb variants by direct surface sampling of dried blood spots by use of an Advion Triversa Nanomate automated electrospray system coupled to a high-resolution mass spectrometer. The method involves no sample preparation. It is possible to unambiguously identify homozygous and heterozygous HbS, HbC, and HbD variants in <10 min without the need for additional confirmation. The method allows for repeated analysis of a single blood spot over a prolonged time period and is tolerant of blood spot storage conditions. PMID:21341716

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

    PubMed

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

    2016-08-01

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

  10. Development of a fluorometric microtiter plate based enzyme assay for MPS IVA (Morquio type A) using dried blood spots.

    PubMed

    Ullal, Anirudh J; Millington, David S; Bali, Deeksha S

    2014-01-01

    Mucopolysaccharidosis type IVA or Morquio type-A disease is a hereditary lysosomal storage disorder caused by deficient activity of the lysosomal enzyme N-acetylgalactosamine-6-sulfate sulfatase (GALNS). The disease is caused by lysosomal accumulation of unprocessed glycosaminoglycans (GAGs) that manifests with severe to mild skeletal and cardiopulmonary abnormalities. We have developed a modified microtiter plate-based enzyme activity assay using dried blood spots and a fluorescent substrate for measuring specific GALNS activity to identify patients with MPS IVA.

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

    NASA Astrophysics Data System (ADS)

    Aarif, Ovais; Aggarwal, Anjali

    2016-03-01

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

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

    PubMed

    Aarif, Ovais; Aggarwal, Anjali

    2016-03-01

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

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

    PubMed Central

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

    2013-01-01

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

  14. A dried blood spots technique based LC-MS/MS method for the analysis of posaconazole in human whole blood samples.

    PubMed

    Reddy, Todime M; Tama, Cristina I; Hayes, Roger N

    2011-11-15

    A rugged and robust liquid chromatographic tandem mass spectrometric (LC-MS/MS) method utilizing dried blood spots (DBS) was developed and validated for the analysis of posaconazole in human whole blood. Posaconazole fortified blood samples were spotted (15 μL) onto Ahlstrom Alh-226 DBS cards and dried for at least 2h. Punched spots were then extracted by using a mixture of acetonitrile and water containing stable labeled internal standard (IS). Posaconazole and its IS were separated from endogenous matrix components on a Kinetex™ C18 column under gradient conditions with a mobile phase A consisting of 0.1% formic acid and a mobile phase B consisting of 0.1% formic acid in acetonitrile/methanol (70/30, v/v). The analyte and IS were detected using a Sciex API 4000 triple quadrupole LC-MS/MS system equipped with a TurboIonSpray™ source operated in the positive ion mode. The assay was linear over the concentration range of 5-5000 ng/mL. The inter-run accuracy and precision of the assay were -1.8% to 0.8% and 4.0% to 10.4%, respectively. Additional assessments unique to DBS were investigated including sample spot homogeneity, spot volume, and hematocrit. Blood spot homogeneity was maintained and accurate and precise quantitation results were obtained when using a blood spot volume of between 15 and 35 μL. Human blood samples with hematocrit values ranging between 25% and 41% gave acceptable quantitation results. The validation results indicate that the method is accurate, precise, sensitive, selective and reproducible.

  15. Blood cholesterol screening in several environments using a portable, dry-chemistry analyzer and fingerstick blood samples. Lipid Research Clinics Cholesterol Screening Study Group.

    PubMed

    Bradford, R H; Bachorik, P S; Roberts, K; Williams, O D; Gotto, A M

    1990-01-01

    A multicenter study of blood cholesterol screening was performed in several typical environments, such as community sites (shopping malls and a supermarket), health care sites, work sites, a blood bank and a school. Cholesterol was measured with a portable, dry-chemistry analyzer using capillary blood obtained by fingerstick. Data are reported from a total of 13,824 participants, spanning the entire age spectrum. Overall, 25% of screened subjects had blood cholesterol levels above the age-specific cutpoints used in the current study. Although in the aggregate this screening experience very closely approximates the expected level of referrals, the proportion of referred screened subjects differed significantly among the 5 types of screening environments and by gender. Follow-up telephone interviews indicated that 53% of referrals had initiated a physician contact. More than 75% of those who had seen a physician reported that the diagnosis of hypercholesterolemia had been confirmed, and almost 72% had been prescribed a diet. A large proportion of referred screened subjects reported having modified their diet, particularly when recommended to do so by a physician. This study has yielded encouraging evidence that physicians gave referred screened subjects appropriate initial advice for managing hypercholesterolemia. The new technology for blood cholesterol measurement evaluated in the current study has proven to be a feasible and reliable means for measuring blood cholesterol in typical screening settings.

  16. Emtricitabine-Triphosphate in Dried Blood Spots as a Marker of Recent Dosing.

    PubMed

    Castillo-Mancilla, Jose; Seifert, Sharon; Campbell, Kayla; Coleman, Stacey; McAllister, Kevin; Zheng, Jia-Hua; Gardner, Edward M; Liu, Albert; Glidden, David V; Grant, Robert; Hosek, Sybil; Wilson, Craig M; Bushman, Lane R; MaWhinney, Samantha; Anderson, Peter L

    2016-11-01

    New objective measures of antiretroviral adherence are needed. We determined if emtricitabine triphosphate (FTC-TP) in dried blood spots (DBS) can be used as a marker of recent dosing with tenofovir disoproxil fumarate-emtricitabine (TDF-FTC). The half-life of FTC-TP was estimated in DBS samples obtained from an intensive pharmacokinetic (PK) study of coformulated TDF-FTC in HIV-negative and HIV-infected participants. The concordance of quantifiable FTC-TP in DBS with tenofovir (TFV)/FTC in plasma was evaluated by utilizing paired plasma-DBS samples from participants enrolled in 2 large preexposure prophylaxis (PrEP) open-label trials. The time to FTC-TP nondetectability after TDF-FTC dosing was evaluated utilizing DBS from HIV-negative participants enrolled in a directly observed therapy study of variable adherence to TDF-FTC. The mean (95% confidence interval [CI]) terminal half-life of FTC-TP in the PK study was 35 (23 to 47) h. A total of 143/163 (88%) samples obtained 0 to 48 h post-TDF-FTC dose had quantifiable FTC-TP in DBS, compared with 2/93 (2%) and 0/87 (0%) obtained >48 and >96 h postdose. In 746 paired plasma-DBS samples from 445 participants enrolled in PrEP trials, when both TFV/FTC in plasma were below the limit of quantification, FTC-TP was as well in 98.9% of the samples, and when either TFV or FTC in plasma was quantifiable, FTC-TP was as well in 90.5% of the samples. The half-life of FTC-TP in DBS is short relative to that of TFV-diphosphate (TFV-DP), making it a surrogate for TFV-FTC detection in plasma. FTC-TP can be quantified in DBS simultaneously with TFV-DP, which quantifies cumulative adherence to TDF-FTC. (The clinical trials discussed in this article have been registered at ClinicalTrials.gov under identifiers NCT01040091, NCT02022657, NCT00458393, NCT01772823, and NCT02012621.).

  17. Development and Implementation Challenges of a Quality Assured HIV Infant Diagnosis Program in Nigeria Using Dried Blood Spots and DNA Polymerase Chain Reaction

    PubMed Central

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

    2015-01-01

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

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

    PubMed

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

    2015-04-01

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

  19. Evaluation of fibronectin 1 in one dried blood spot and in urine after rhGH treatment.

    PubMed

    Ferro, P; Ventura, R; Pérez-Mañá, C; Farré, M; Segura, J

    2016-10-07

    Since the appearance of recombinant human growth hormone (rhGH) in the 1980s, its expansion and acquisition through the black market has increased, so the detection of its abuse continues to be a challenge. New biomarkers that are more reliable and sensitive, allowing a larger detection window, are still needed. In this line, Fibronectin 1 (FN1) has been proposed as a potential genetic and protein biomarker of rhGH abuse in peripheral blood lymphocytes, serum, and plasma. However, logistic problems associated with current blood collection in sports drug testing point towards potential new alternative matrices that could be good candidates to be evaluated. Results obtained in this study showed high ELISA FN1 levels in one dried blood spot and in urine samples in ten healthy male volunteers treated with rhGH. Results showed that especially dried blood spots appear as a potential good matrix to detect rhGH abuse by means of FN1 biomarker. Copyright © 2016 John Wiley & Sons, Ltd.

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

    PubMed

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

    2016-07-01

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

  1. Cost Evaluation of Dried Blood Spot Home Sampling as Compared to Conventional Sampling for Therapeutic Drug Monitoring in Children

    PubMed Central

    Martial, Lisa C.; Aarnoutse, Rob E.; Schreuder, Michiel F.; Henriet, Stefanie S.; Brüggemann, Roger J. M.; Joore, Manuela A.

    2016-01-01

    Dried blood spot (DBS) sampling for the purpose of therapeutic drug monitoring can be an attractive alternative for conventional blood sampling, especially in children. This study aimed to compare all costs involved in conventional sampling versus DBS home sampling in two pediatric populations: renal transplant patients and hemato-oncology patients. Total costs were computed from a societal perspective by adding up healthcare cost, patient related costs and costs related to loss of productivity of the caregiver. Switching to DBS home sampling was associated with a cost reduction of 43% for hemato-oncology patients (€277 to €158) and 61% for nephrology patients (€259 to €102) from a societal perspective (total costs) per blood draw. From a healthcare perspective, costs reduced with 7% for hemato-oncology patients and with 21% for nephrology patients. Total savings depend on the number of hospital visits that can be avoided by using home sampling instead of conventional sampling. PMID:27941974

  2. Cost Evaluation of Dried Blood Spot Home Sampling as Compared to Conventional Sampling for Therapeutic Drug Monitoring in Children.

    PubMed

    Martial, Lisa C; Aarnoutse, Rob E; Schreuder, Michiel F; Henriet, Stefanie S; Brüggemann, Roger J M; Joore, Manuela A

    2016-01-01

    Dried blood spot (DBS) sampling for the purpose of therapeutic drug monitoring can be an attractive alternative for conventional blood sampling, especially in children. This study aimed to compare all costs involved in conventional sampling versus DBS home sampling in two pediatric populations: renal transplant patients and hemato-oncology patients. Total costs were computed from a societal perspective by adding up healthcare cost, patient related costs and costs related to loss of productivity of the caregiver. Switching to DBS home sampling was associated with a cost reduction of 43% for hemato-oncology patients (€277 to €158) and 61% for nephrology patients (€259 to €102) from a societal perspective (total costs) per blood draw. From a healthcare perspective, costs reduced with 7% for hemato-oncology patients and with 21% for nephrology patients. Total savings depend on the number of hospital visits that can be avoided by using home sampling instead of conventional sampling.

  3. Therapeutic drug monitoring of everolimus using the dried blood spot method in combination with liquid chromatography-mass spectrometry.

    PubMed

    van der Heijden, J; de Beer, Y; Hoogtanders, K; Christiaans, M; de Jong, G J; Neef, C; Stolk, L

    2009-11-01

    An assay of everolimus based on finger prick sampling and consecutive application as a blood spot on sampling paper has been developed. We explored several methods [K. Hoogtanders, J. van der Heijden, M. Christiaans, P. Edelbroek, J. van Hooff, L. Stolk, J. Pharm. Biomed. Anal. 44 (2006) 658-664; A. Allanson, M. Cotton, J. Tettey, et al., J. Pharm. Biomed. Anal. 44 (2007) 963-969] and developed a new method, namely the impregnation of sampling paper with a solution of plasma-protein, formic acid and ammonium acetate, in combination with the extraction of the blood spot by filter filtration. This kind of sample preparation provides new possibilities for blood spot sampling especially if analytes are adsorbed to the paper. The dried blood spot was analysed using the HPLC-electrospray-tandem mass spectrometry method, with 32-desmethoxyrapamycin as the internal standard. The working range of our study was 2-30 microg/l. Within this range, intra-and inter-assay variability for precision and accuracy was <15%. Everolimus blood spot samples proved stable for 3 days at 60 degrees C and for 32 days at 4 degrees C. Everolimus concentrations of one stable out-patient were compared after both blood spot sampling and conventional venous sampling on various occasions. Results indicate that this new method is promising for therapeutic drug monitoring in stable renal transplant patients.

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

    PubMed

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

    2011-10-01

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

  5. DRIED BLOOD SPOT REAL-TIME POLYMERASE CHAIN REACTION ASSAYS TO SCREEN NEWBORNS FOR CONGENITAL CYTOMEGALOVIRUS INFECTION

    PubMed Central

    Boppana, Suresh B.; Ross, Shannon A.; Novak, Zdenek; Shimamura, Masako; Tolan, Robert W.; Palmer, April L.; Ahmed, Amina; Michaels, Marian G.; Sánchez, Pablo J.; Bernstein, David I.; Britt, William J.; Fowler, Karen B.

    2010-01-01

    Context Reliable methods to screen newborns for congenital cytomegalovirus (CMV) infection are needed for identification of infants at increased risk for hearing loss. Since dried blood spots (DBS) are routinely collected for metabolic screening from all newborns in the United States, there has been interest in using DBS polymerase chain reaction (PCR)-based methods for newborn CMV screening. Objective To determine the diagnostic accuracy of DBS real-time PCR assays for newborn CMV screening Design, Setting, and Participants Between March 2007 and May 2008, infants born at seven medical centers in the U.S. were enrolled in the CMV and Hearing Multicenter Screening (CHIMES) study. Newborn saliva specimens were tested for the detection of early antigen fluorescent foci (DEAFF). Results of saliva DEAFF were compared with a single-primer (from 03/07 to 12/07) and a two-primer (from 01/08 to 05/08) DBS real-time PCR. Infants positive by screening DEAFF or PCR were enrolled in follow-up to confirm congenital infection by the reference standard method, DEAFF on saliva or urine. Main Outcome Measures Sensitivity, specificity and positive and negative likelihood ratios (LRs) of single-primer and two-primer DBS real-time PCR assays for identifying infants with confirmed congenital CMV infection. Results Congenital CMV infection was confirmed in 92 of 20,448 (0.45%; 95% CI, 0.36–0.55) infants. Ninety-one of 92 infants were saliva DEAFF positive on screening. Of the 11,422 infants screened using the single-primer DBS PCR, 17 of 60 (28%) infants were positive with this assay, whereas, among the 9,026 infants screened using the two-primer DBS PCR, 11 of 32 (34%) infants were positive. The single-primer DBS PCR identified congenital CMV infection with a sensitivity of 28.3% (95% CI, 17.4–41.4%), specificity, 99.9% (95% CI, 99.9–100%), positive LR, 803.7 (95% CI, 278.7–2317.9), and negative LR, 0.7 (95% CI, 0.6–0.8). The positive and negative predictive values of the

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

    PubMed

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

    2013-03-25

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

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

    PubMed Central

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

    2016-01-01

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

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

    PubMed

    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.

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

    PubMed Central

    Wickremsinhe, Enaksha R; Perkins, Everett J

    2015-01-01

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

  10. Determination of lead in blood by laser ablation ICP-TOF-MS analysis of blood spotted and dried on filter paper: a feasibility study.

    PubMed

    Cizdziel, James V

    2007-06-01

    Screening programs for pediatric lead (Pb) poisoning routinely collect blood by finger stick onto specially designed filter paper (FP) for subsequent laboratory testing. Here, laser ablation inductively coupled plasma time-of-flight mass spectrometry is evaluated as a novel method for the determination of Pb in blood spotted onto FP cards. The method requires no sample pretreatment and may be particularly suitable for rapidly screening a large number of samples. Signal intensity from the ICP-MS strongly correlated (r (2) = 0.996) with Pb target values for five blood cards from a laboratory proficiency testing program. Better precision was obtained by replicate ablation line scans (<15% rsd) compared to a grid of 25 individual ablation spots (>30% rsd). Because <2% of the sample is consumed in the laser ablation analysis, selected samples can also be analyzed by traditional techniques or archived. Lead isotope ratios determined during the same total-Pb analysis appear to be effective for discerning contamination extraneous to the blood sample. Determining that an elevated result was due to contamination may negate the need for retesting with invasive pediatric venipuncture. Some elements monitored along with Pb exhibited signal profiles conducive to potential quantitation (Ca, V, Fe, Cu Zn), whereas others did not (Cr, Mn, Co, Ni, Cd, Hg, Tl, U). Copper, Ca, and V were inversely correlated with Pb (r = -0.89, -0.90, and -0.92, respectively). Scanning electron microscope images revealed surface modality differences between FP spotted with fresh whole blood and reconstituted freeze-dried blood reference material.

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

    PubMed

    Verplaetse, Ruth; Henion, Jack

    2016-01-01

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

  12. Bosentan Pharmacokinetics in Pediatric Patients with Pulmonary Arterial Hypertension: Comparison of Dried Blood Spot and Plasma Analysis.

    PubMed

    Géhin, Martine; Sidharta, Patricia N; Dingemanse, Jasper

    2016-01-01

    FUTURE-3, a phase III pediatric pharmacokinetic (PK) trial conducted to compare 2 bosentan dosing regimens in 64 patients with pulmonary arterial hypertension, offered the opportunity to compare dried blood spot (DBS)-derived data to plasma data. Bosentan PK parameters obtained with both methods were compared by the geometric mean ratio (GMR; DBS/plasma) and its 90% CI after correction for the blood-to-plasma partition ratio (0.6). Bosentan GMRs were 1.10 (1.03, 1.16) and 1.12 (1.04, 1.20) for AUCτ and Cmax, respectively. Bosentan concentrations measured by DBS were therefore good estimations of bosentan plasma concentrations. DBS can be considered a valid alternative to bosentan assessed in plasma.

  13. Examination of Eurasian griffon vultures (Gyps fulvus fulvus) in Israel for exposure to environmental toxicants using dried blood spots.

    PubMed

    Shlosberg, Alan; Wu, Qian; Rumbeiha, Wilson K; Lehner, Andreas; Cuneah, Olga; King, Roni; Hatzofe, Ohad; Kannan, Kurunthachalam; Johnson, Margaret

    2012-04-01

    The griffon vulture (Gyps fulvus) is one of seven species of Old World Gyps vultures found over a wide range from the Iberian peninsula in the west through the Balkans, Turkey, and the Middle East to India in the east. The population of the griffon vultures in Israel has suffered a dramatic decrease, and in recent years productivity has been severely reduced. In this study, whole-blood samples taken from 25 apparently healthy griffon vultures at various stages of maturity were examined to investigate whether the vultures are being excessively exposed to environmental contaminants that might deleteriously affect their reproduction. Five groups of environmental contaminants, comprising toxic elements, organochlorine pesticides, polychlorinated biphenyls, polybrominated diphenyl ethers, and perfluorinated compounds, were monitored in dried blood spots. Results of the analyses showed low levels of exposure of griffon vultures to environmental contaminants compared with the sparse data available on griffon vultures and other diurnal raptors in other countries.

  14. Evaluation of Two Techniques for Viral Load Monitoring Using Dried Blood Spot in Routine Practice in Vietnam (French National Agency for AIDS and Hepatitis Research 12338)

    PubMed Central

    Taieb, Fabien; Tram, Tran Hong; Ho, Hien Thi; Pham, Van Anh; Nguyen, Lan; Pham, Ban Hien; Tong, Linh An; Tuaillon, Edouard; Delaporte, Eric; Nguyen, Anh Tuan; Bui, Duc Duong; Do, NhanThi; Madec, Yoann

    2016-01-01

    Background. Although it is the best method to detect early therapeutic failure, viral load (VL) monitoring is still not widely available in many resource-limited settings because of difficulties in specimen transfer, personnel shortage, and insufficient laboratory infrastructures. Dried blood spot (DBS) use, which was introduced in the latest World Health Organization recommendations, can overcome these difficulties. This evaluation aimed at validating VL measurement in DBS, in a laboratory without previous DBS experience and in routine testing conditions. Methods. Human immunodeficiency virus (HIV)-infected adults were observed in a HIV care site in Hanoi, and each patient provided 2 DBS cards with whole blood spots and 2 plasma samples. Viral load was measured in DBS and in plasma using the COBAS Ampliprep/TaqMan and the Abbott RealTime assays. To correctly identify those with VL ≥ 1000 copies/mL, sensitivity and specificity were estimated. Results. A total of 198 patients were enrolled. With the Roche technique, 51 plasma VL were ≥1000 copies/mL; among these, 28 presented a VL in DBS that was also ≥1000 copies/mL (sensitivity, 54.9; 95% confidence interval [CI], 40.3–68.9). On the other hand, all plasma VL < 1000 copies/mL were also <1000 copies/mL in DBS (specificity, 100; 95% CI, 97.5–100). With the Abbott technique, 45 plasma VL were ≥1000 copies/mL; among these, 42 VL in DBS were also ≥1000 copies/mL (sensitivity, 93.3%; 95% CI, 81.7–98.6); specificity was 94.8 (95% CI, 90.0–97.7). Conclusions. The Abbott RealTime polymerase chain reaction assay provided adequate VL results in DBS, thus allowing DBS use for VL monitoring. PMID:27704001

  15. Multicenter comparison of ESP Culture System II with BACTEC 460TB and with Lowenstein-Jensen medium for recovery of mycobacteria from different clinical specimens, including blood.

    PubMed

    Tortoli, E; Cichero, P; Chirillo, M G; Gismondo, M R; Bono, L; Gesu, G; Simonetti, M T; Volpe, G; Nardi, G; Marone, P

    1998-05-01

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

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

    PubMed

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

    2016-02-01

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

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

    NASA Astrophysics Data System (ADS)

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

    2012-11-01

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

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

    PubMed Central

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

    2015-01-01

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

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

    PubMed

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

    2015-01-01

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

  20. Low risk of contamination with automated and manual excision of dried blood spots for HIV DNA PCR testing in the routine laboratory.

    PubMed

    Driver, Glenn A; Patton, Janet C; Moloi, Jackie; Stevens, Wendy S; Sherman, Gayle G

    2007-12-01

    In low resource settings the inability to diagnose HIV in infants early presents a major obstacle to providing care for HIV-infected children. Dried blood spot samples offer a solution to the scarcity of skills available for venesecting young infants but pose challenges to laboratories because their processing requirements are distinct from that of the liquid blood samples widely used for viral detection assays. Different methods of excising 287 paired HIV-positive and HIV-negative dried blood spot samples (n=574) for testing by the Roche Amplicor HIV-1 DNA assay version 1.5 were assessed. A manual punch in conjunction with three different cleaning protocols (n=372) and an automated punch (BSD 1000 GenePunch) using a single cleaning protocol (n=202) was assessed for the risk of cross contamination between samples. A single false positive result obtained using the automated punch may have been attributable to contamination during the excision step of the assay. Excision of dried blood spot samples is associated with a very low risk of cross contamination regardless of the instrument or cleaning intervention used. The process of excising dried blood spot samples should not compromise the results of HIV viral detection assays performed on dried blood spots in routine laboratories which is encouraging for increasing access to an accurate diagnosis of HIV in infants.

  1. [Direct proteomic profiling of human urine and blood serum in an experiment with 5-day dry immersion].

    PubMed

    2012-01-01

    Changes in proteome of urine and blood serum obtained from 14 healthy humans (age 21-29 yrs) medically certified for an experiment with dry immersion were analyzed. Urine and serum samples were pre-fractionated and enriched with magnetic particles MB-WCX and MB-HIC, respectively, on robot ClinProt (Bruker Daltonics) for direct mass-spectrometry profiling by MALDI-TOF. As a result, 143 protein peaks on the average were identified in urine samples. It was shown that a high variation coefficient in 23.7% of protein peaks, i.e. double technical, points to the most plastic fraction of the urine proteome. In blood serum, 175 peaks were identified in a sample on the average. Comparison of baseline and immersion mass-spectra of the blood proteome revealed significant differences. Increased peak areas of several protein fragments--C3 and C4 fragments of complement system, high-molecular kininogen and fibrinogen--can be ascribed to human body adaptation to the experimental conditions.

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

    PubMed Central

    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

  3. Simultaneous quantitation of metformin and sitagliptin from mouse and human dried blood spots using laser diode thermal desorption tandem mass spectrometry.

    PubMed

    Swales, John G; Gallagher, Richard T; Denn, Mark; Peter, Raimund M

    2011-06-01

    A simple, rapid and robust high-throughput assay for the simultaneous analysis of metformin and sitagliptin from mouse and human dried blood spot samples using laser diode thermal desorption interfaced with atmospheric pressure chemical ionization tandem mass spectrometry (LDTD-APCI-MS/MS) was developed for use in a pharmaceutical discovery environment as an alternative to traditional plasma analysis. Analytes were extracted from dried blood spots using a simple punch disc and solvent extract procedure. Details of the method development and optimization of the instrumental parameters are presented. The method was successfully applied to spiked mouse and human dried blood spot samples. Analyte stability was determined in dried blood spots on FTA cards and as extracts of dried blood spots. The method was subsequently used to determine the oral pharmacokinetics of metformin and sitagliptin after dosing to male mice. Metformin and Sitagliptin results are compared to data generated by more traditional liquid chromatography-mass spectrometry methods. Intra-assay and inter-assay accuracy and precision across the analytes and species deviated by less than 30% at all calibration levels and less than 20% at all quality control levels.

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

    PubMed Central

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

    2015-01-01

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

  5. Methylmalonic acid quantified in dried blood spots provides a precise, valid, and stable measure of functional vitamin B-12 status in healthy women.

    PubMed

    Schroder, Theresa H; Quay, Teo A W; Lamers, Yvonne

    2014-10-01

    Methylmalonic acid (MMA) is a sensitive and specific functional biomarker of vitamin B-12 status, commonly assessed in plasma or serum. Dried blood spots (DBSs) allow simpler and more cost-efficient blood sampling than plasma. To facilitate convenient testing for vitamin B-12 deficiency in large-scale surveys and in population groups from remote areas, we developed a method for MMA quantification in DBSs and tested its applicability as well as the long-term stability of MMA in DBSs at various temperatures. MMA was extracted from an 8-mm DBS punch with water:methanol (95:5, v:v) and methyl-d3-malonic acid as the internal standard. After sample cleanup by ultrafiltration and hexane extraction, MMA was quantified by using reversed-phase LC-tandem mass spectrometry. Extraction conditions were optimized to maximize the detection signal and achieve DBS extract concentrations above the lowest limit of quantification (signal-to-noise ratio ≥ 10) of 10 nmol/L. Recovery was between 93% and 96%. Intra- and interassay variation (CV%) for DBS MMA was 0.49% and 2.3%, respectively. Calibrators showed linearity (R(2) = 0.998) between 10 and 10,000 nmol/L. In 94 healthy women, MMA concentrations in DBS extract (min-max: 10.2-80.5 nmol/L) and plasma (min-max: 68-950 nmol/L) were correlated (ρ = 0.90) (P < 0.001). MMA concentrations in DBSs were stable at room temperature for 1 wk, in the refrigerator for 8 wk, and at -80°C for at least 1 y. This simple and robust method allows quantification of MMA in DBSs of healthy individuals. The linear relation between plasma and DBS MMA suggests that DBS MMA could predict plasma MMA, the current reference indicator for functional vitamin B-12 deficiency. With the advantages of minimally invasive specimen collection and no need for laborious blood processing steps, this method has the potential to be a reliable, convenient, and field-applicable alternative for assessment of vitamin B-12 status.

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

    PubMed Central

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

    2014-01-01

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

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

    PubMed Central

    Turecek, Frantisek; Scott, C. Ron; Chamoles, Nestor A.

    2008-01-01

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

  8. A simple screening method for individuals at risk of developing type 1 diabetes: measurement of islet cell autoantibodies (GADA, IA-2A, and IAA) on dried capillary blood spots collected on filter paper.

    PubMed

    Siraj, E S; Rogers, D G; Gupta, M K; Reddy, S S K

    2012-10-01

    There is limited information regarding the use of dried capillary blood spots collected on a filter paper (FP) to test for islet cell antibodies. The aim of this study was to validate the use of dried capillary blood spots collected on a FP for the analysis of islet cell antibodies. FP eluates were tested using both single and combined assay for antibodies to glutamic acid decarboxylase (GADA) and/or to the protein tyrosine phosphatase like IA-2 (IA-2A), and a single assay for antibodies to insulin (IAA). The results were compared with those of serum assays. Ninety-one subjects were studied. Forty had Type 1 diabetes mellitus (T1DM) and 51 were first-degree relatives (FDR) of patients with T1DM. The GADA and IA-2A were measured by radio-binding assays, which utilize 35S-labeled GAD65 and IA-2. IAA was measured by a microtiter plate assay using 125I-labeled insulin. Twenty-six of those with T1DM (65%) and 5 of the FDRs (10%) had at least 1 positive test on the single serum assays. The FP combi-assay for GADA and IA-2A had 97.8% concordance rate when compared with serum single assays for GADA and IA-2A. The concordance rate for individual assays were 96.7% for GADA, and 100% for both IA-2A and IAA There was significant correlation of the antibody levels between FP and serum specimen for all 3 antibodies. We conclude that antibody screening performed using dried capillary blood spots collected on a FP correlates well with serum assays, and provides an easy alternative for population screening.

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

    PubMed

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

    2014-08-01

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

  10. Mass spectrometric studies on the in vivo metabolism and excretion of SIRT1 activating drugs in rat urine, dried blood spots, and plasma samples for doping control purposes.

    PubMed

    Höppner, Sebastian; Delahaut, Philippe; Schänzer, Wilhelm; Thevis, Mario

    2014-01-01

    The NAD(+) depending enzyme SIRT1 regulates the mitochondrial biogenesis, fat and glucose metabolism through catalyzing the deacetylation of several metabolism-related protein-substrates. Recently, synthetic activators of SIRT1 referred to as STACs (Sirtuin activating compounds, e.g. SRT2104) were identified and tested in clinical studies for the treatment of aging-related diseases such as type 2 diabetes, Alzheimer's and obesity. Although the mechanism of SIRT1 activation by small molecules has caused considerable controversy, STACs demonstrated a significant performance enhancement in mice experiments including an improvement of endurance, muscle strength, and locomotor behavior. Due to their potential to increase exercise tolerance in healthy individuals, SIRT1 activators are currently being monitored by anti-doping authorities. In the present study, the in vivo metabolic clearance of three SIRT1 activators was investigated in rats by the collection of urine, DBS (dried blood spots) and plasma samples following a single oral administration. The resulting metabolic products were studied by positive electrospray ionization - (tandem) mass spectrometry and confirmed by the comparison with in vitro generated metabolites using human and rat liver microsomal preparations. Subsequently, a screening procedure for five SIRT1 activators and the metabolite M1-SRT1720 in DBS specimens was developed. Liquid-liquid-extraction and liquid chromatography/tandem mass spectrometry was employed based on diagnostic ion transitions recorded in multiple reaction monitoring mode and two deuterated internal standards namely d8-SRT1720 and d8-M1-SRT1720 were utilized. The doping control assay was characterized with regard to specificity, limit of detection (10-50ng/ml), recovery (65-83%) and imprecision (7-20%) and ion suppression/enhancement effects (<10%), demonstrating its fitness-for-purpose for sports drug testing applications.

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

    PubMed Central

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

    2003-01-01

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

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

    NASA Astrophysics Data System (ADS)

    Shyamkumar, Prashanth

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

  13. Effects of the usage of dried brewing yeast in the diets on the performance, egg traits and blood parameters in quails.

    PubMed

    Yalçın, S; Erol, H; Ozsoy, B; Onbaşılar, I; Yalçın, S

    2008-12-01

    This experiment was carried out to determine the effects of the usage of dried brewing yeast in quail diets on laying performance, egg traits and blood parameters. A total of 240 Japanese quails (Coturnix coturnix japonica) aged 10 weeks were randomly allocated into one control group and three treatment groups. Each group was divided into five replicates as subgroups, comprising 12 quails each. Dried brewing yeast (Saccharomyces cerevisiae) was used at the levels of 1.5%, 3.0% and 4.5% in the diets of the first, second and third treatment groups, respectively. Soyabean meal was replaced with dried brewing yeast. The diets were formulated to be isocaloric and isonitrogenous. The experimental period lasted 18 weeks. Dietary treatments did not significantly affect body weight, daily feed intake, daily protein intake, egg production, egg weight, feed efficiency, mortality, egg shell thickness, egg albumen index, egg yolk index, egg Haugh unit, the percentages of egg shell, albumen and yolk, excreta moisture and small intestinal pH. Inclusion of 3% and 4.5% dried brewing yeast in diets reduced egg yolk cholesterol concentration as mg per yolk and mg per g yolk (P < 0.01). Blood serum cholesterol of groups fed diets with dried brewing yeast was significantly lower (P < 0.01) than that of the control group. Feeding diets containing 3.0% and 4.5% dried brewing yeast resulted in significant increases (P < 0.01) in blood serum levels of total protein, alanine aminotransferase at the end of the experiment. Blood serum levels of uric acid, triglyceride, aspartate aminotransferase and alkaline phosphatase were not affected by dietary dried brewing yeast. It is concluded that dried brewing yeast can be used up to 4.5% in the diets of laying quails without adverse effects on the measured parameters.

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

    PubMed Central

    Chung, So-Hyang

    2016-01-01

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

  15. Structure build-up and evolution in the drying of sessile blood droplets

    NASA Astrophysics Data System (ADS)

    Craster, Richard; Uppal, Aran; Matar, Omar

    2016-11-01

    Experimental observations have recorded blood undergoing a sol-gel transition during the evaporation process. Consequently, the rheology becomes non-uniform throughout the droplet and exhibits transitional complex phenomena that we must capture if we wish to accurately model the evaporative/cracking process. We propose a model where thixotropy is introduced to capture the evolving rheology as evaporation occurs. Thixotropy is often used to describe fluids which exhibit a decrease in viscosity due to flow and subsequent slow recovery of viscosity after the cessation of the flow. We introduce an additional parameter to describe the internal structure of the fluid at each point and consider a droplet in the limit of the lubrication approximation. We present a discussion of our results that demonstrates the dependence of structure build-up, which accompanies the spatio-temporal evolution of the drop, on system parameters. EPSRC UK Centre for Doctoral Training.

  16. Determination of venlafaxine and O-desmethylvenlafaxine in dried blood spots for TDM purposes, using LC-MS/MS.

    PubMed

    Berm, E J J; Brummel-Mulder, E; Paardekooper, J; Hak, E; Wilffert, B; Maring, J G

    2014-04-01

    Dried blood spot (DBS) sampling and quantitative analyses of many current therapeutic drug monitoring (TDM)-guided drugs are advantageous because of the minimal invasive sampling strategy. Here, a fast and robust LC-MS/MS method was developed and analytically validated for simultaneous determination of venlafaxine (VEN) and O-desmethylvenlafaxine (ODV) in DBS. Six-millimeter circles were punched out from DBS collected on Whatman DMPK-C paper, and the DBS was extracted with acetonitrile/methanol at 1:3. The total run time was 4.8 min. The assay was linear in the range of 20-1,000 μg/L for both VEN and ODV. Assay accuracy and precision was well within limits of acceptance (LLOQ = 20 μg/L). Normal hematocrit concentrations (0.30-0.50) did not influence the results neither did a normal spot volume (40-80 μL). Punch position at the perimeter instead of the center of the blood spot gave a bias ranging from 2.4 to 10.4%. Correlation between plasma and spiked DBS samples was high. The concentrations found in spiked DBS samples were higher than those in plasma, indicating that a conversion factor for translation of DBS to plasma values is needed. This analytically validated method is suitable for determination of VEN and ODV in DBS and applicable for TDM. The method will be used for TDM of VEN in the Dutch CYSCE multicenter trial (NCT01778907).

  17. Dried blood spots: liquid chromatography-mass spectrometry analysis of Δ9-tetrahydrocannabinol and its main metabolites.

    PubMed

    Mercolini, Laura; Mandrioli, Roberto; Sorella, Vittorio; Somaini, Lorenzo; Giocondi, Daniele; Serpelloni, Giovanni; Raggi, Maria Augusta

    2013-01-04

    A sensitive and selective HPLC-MS/MS method has been developed for the first time for the analysis of Δ(9)-tetrahydrocannabinol (the most important active cannabinoid) and its hydroxylated and carboxylated metabolites in human Dried Blood Spots (DBSs). The simultaneous determination of Δ(9)-tetrahydrocannabinol and its two main metabolites allows assessing the time elapsed after the drug intake and distinguishing between acute or former consumption. This is an important information in specific contexts such as "on street" controls by police forces. DBSs have been chosen as the optimal biological matrix for this kind of testing, since they provide information on the actual state of intoxication, without storage and transportation problems usually associated with classical blood testing. The analysis is carried out on a C8 reversed phase column with a mobile phase composed of 0.1% formic acid in a water/methanol mixture and an electrospray ionisation (ESI) source, coupled to a triple quadrupole mass spectrometer. The method was validated according to international guidelines, with satisfactory results in terms of extraction yields, precision, stability and accuracy. Application to real DBS samples from Cannabis abusers gave reliable results, thus confirming the methodology suitability for roadside testing.

  18. Development and validation of a dried blood spot LC-MS/MS assay to quantify ranitidine in paediatric samples.

    PubMed

    Yakkundi, Shirish; Millership, Jeff; Collier, Paul; Shields, Michael D; McElnay, James

    2011-12-15

    A novel approach has been developed to determine ranitidine in paediatric samples using dried blood spots (DBS) on Guthrie cards (Whatman 903). A selective and sensitive HPLC-MS/MS assay has been developed and validated using small volumes of blood (30 μl). A 6 mm disc was punched from each DBS and extracted with methanolic solution of the internal standard (IS) nizatidine. This was further subjected to solid phase extraction (SPE), followed by reversed phase HPLC separation, using a XBridge™ C18 column and mobile phase 10 mM ammonium acetate/methanol (98:2 v/v) with a flow rate of 0.3 mL/min. This was combined with multiple reaction monitoring (MRM) mass detection using electrospray ionisation (ESI). The calibration curve for ranitidine was found linear over the range 10-500 ng/mL (r=0.996). The limit of quantification (LOQ) of the method was validated at 10 ng/mL. Accuracy and precision values for within and between days were <20% at the LOQ and <15% at all other concentrations. The validated DBS method was successfully applied to a clinical study employing 81 samples from 36 paediatric patients.

  19. Validation of an optimized ELISA for quantitative assessment of Epstein-Barr virus antibodies from dried blood spots

    PubMed Central

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

    2016-01-01

    Our objective was to validate a commercially-available ELISA to measure antibody titers against Epstein-Barr virus (EBV) in dried blood spots (DBS) to replace a previously validated assay for DBS that is no longer available. We evaluated the precision, reliability, and stability of the assay for the measurement of EBV antibodies in matched plasma, fingerprick DBS, and venous blood DBS samples from 208 individuals. Effects of hematocrit and DBS sample matrix on EBV antibody determination were also investigated, and the cut-off for seropositivity in DBS was determined. A conversion equation was derived to enable comparison of results generated using this method with the former DBS method. There was a high correlation between plasma and DBS EBV antibody titers (R2 = 0.93) with very little bias (−0.07 based on Bland-Altman analysis). The assay showed good linearity, did not appear to be affected by the DBS matrix, and physiological hematocrit levels had no effect on assay performance. There was reasonable agreement between DBS EBV titer estimates obtained using this assay and the previously-validated assay (R2 = 0.72). The commercially-available ELISA assay for EBV antibody titers that we validated for use with DBS will facilitate continued investigation of EBV antibody titers in DBS. PMID:27337556

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

    PubMed

    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.

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

    PubMed

    Kip, A E; Rosing, H; Hillebrand, M J X; Blesson, S; Mengesha, B; Diro, E; Hailu, A; Schellens, J H M; Beijnen, J H; Dorlo, T P C

    2016-04-01

    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'sr= 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.

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

    PubMed Central

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

    2016-01-01

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

  3. Carbohydrate supplements and their effects on pasture dry matter intake, feeding behavior, and blood factors associated with intake regulation.

    PubMed

    Sheahan, A J; Kay, J K; Roche, J R

    2013-01-01

    Supplementary feeds are offered to grazing dairy cows to increase dry matter (DM) and metabolizable energy (ME) intakes; however, offering feed supplements reduces pasture DM intake, a phenomenon known as substitution. The objective of the study was to investigate changes in blood factors associated with intake regulation in monogastric species in pasture-fed dairy cows supplemented with either a starch- or nonforage fiber-based concentrate. Fifteen multiparous Friesian × Jersey cross cows were assigned to 1 of 3 treatments at calving. Measurements were undertaken in wk 8 of lactation. Treatments were pasture only, pasture plus a starch-based concentrate (3.5 kg of DM/cow per day; STA), and pasture plus a nonforage fiber-based concentrate (4.4 kg of DM/cow per day). Pelleted concentrates were fed at an isoenergetic rate in 2 equal portions at a.m. and p.m. milkings. Measurements were undertaken to investigate differences in pasture DM intake, feeding behavior, and profiles of blood factors for 4h after a.m. and p.m. milkings, the periods of intensive feeding in grazing cows. Supplementing cows with STA concentrate reduced pasture DM intake to a greater extent than the fiber concentrate, although time spent eating did not differ between treatments. The blood factor response to feeding differed between the a.m. and p.m. feeding events. Blood factors associated with a preprandial or fasted state were elevated prefeeding in the a.m. and declined following feeding, whereas satiety factors increased. In comparison, the blood factor response to feeding in the p.m. differed, with responses to feeding delayed for most factors. Plasma ghrelin concentration increased during the p.m. feeding event, despite the consumption of feed and the positive energy state remaining from the previous a.m. feeding, indicating that environmental factors (e.g., sunset) supersede physiological cues in regulating feeding behavior. The greater reduction in pasture DM intake for the STA treatment

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

    PubMed

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

    2016-02-01

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

  5. Investigation of endogenous blood lipids components that contribute to matrix effects in dried blood spot samples by liquid chromatography-tandem mass spectrometry.

    PubMed

    Ismaiel, Omnia A; Jenkins, Rand G; Karnes, H Thomas

    2013-08-01

    Dried blood spot (DBS) sampling coupled to liquid chromatography-tandem mass spectrometry (LC-MS/MS) is a rapidly developing approach in the field of biopharmaceutical analysis. DBS sampling enables analysis of small sample volumes with high sensitivity and selectivity while providing a convenient easy to store and ship format. Lipid components that may be extracted during biological sample processing may result in matrix ionization effects and can significantly affect the precision and accuracy of the results. Glycerophosphocholines (GPChos), cholesterols and triacylglycerols (TAG) are the main lipid components that contribute to matrix effects in LC-MS/MS. Various organic solvents such as methanol, acetonitrile, methyl tertiary butyl ether, ethyl ether, dichloromethane and n-hexane were investigated for elution of these lipid components from DBS samples. Methanol extracts demonstrated the highest levels of GPChos whereas ethyl ether and n-hexane extracts contained less than 1.0 % of the GPChos levels in the methanol extracts. Ethyl ether extracts contained the highest levels of cholesterols and TAG in comparison to other investigated organic solvents. Acetonitrile is recommended as an elution solvent due to low lipid recoveries. Matrix effects resulted from different extracted lipid components should be studied and assessed carefully in DBS samples.

  6. Genotyping of celiac disease-related-risk haplotypes using a closed-tube polymerase chain reaction analysis of dried blood and saliva disk samples.

    PubMed

    Ollikka, Pia; Raussi, Hanna-Mari; Laitala, Ville; Jaakkola, Lassi; Hovinen, Jari; Hemmilä, Ilkka; Ylikoski, Alice

    2009-03-01

    Expansion of molecular diagnostics more widely into clinical routines requires simplified methods allowing automation. We developed a homogeneous, multilabel polymerase chain reaction (PCR) method based on time-resolved fluorometry, and studied the use of dried disk samples in PCR. Celiac disease-related HLA-DQA1*05, HLA-DQB1*02, and HLA-DQB1*0302 genotyping was used to verify the method with blood and saliva samples dried on S&S 903 and IsoCode sample collection papers. Three sample preparation procedures, including manufacturer's manual elution, an automated elution, and direct use of disk samples, were compared using dried disk samples. The three procedures gave successful amplification and correct genotyping results. Owing to the simplicity of the direct use of disk samples in PCR, this method was chosen for the subsequent homogeneous analysis of blood (n=194) and saliva (n=30) disk samples on S&S 903 paper. The results revealed that, in addition to DNA samples (n=29), both blood and saliva disk samples were successfully amplified and genotyped using the homogeneous PCR assays for HLA-DQA1 and HLA-DQB1. The homogeneous PCR assays developed provide a useful tool to genotype celiac disease-related HLA-DQA1*05, HLA-DQB1*02, and HLA-DQB1*0302 alleles. Furthermore, the method provides a direct way to perform a closed-tube PCR analysis of dried blood and saliva disk samples enabling simple automation.

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

    PubMed

    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.

  8. Prediction of the hematocrit of dried blood spots via potassium measurement on a routine clinical chemistry analyzer.

    PubMed

    Capiau, Sara; Stove, Veronique V; Lambert, Willy E; Stove, Christophe P

    2013-01-02

    The potential of dried blood spot (DBS) sampling as an alternative for classical venous sampling is increasingly recognized, with multiple applications in, e.g., therapeutic drug monitoring and toxicology. Although DBS sampling has many advantages, it is associated with several issues, the hematocrit (Hct) issue being the most widely discussed challenge, given its possible strong impact on DBS-based quantitation. Hitherto, no approaches allow Hct prediction from nonvolumetrically applied DBS. Following a simple and rapid extraction protocol, K(+) levels from 3 mm DBS punches were measured via indirect potentiometry, using the Roche Cobas 8000 routine chemistry analyzer. The extracts' K(+) concentrations were used to calculate the approximate Hct of the blood used to generate DBS. A linear calibration line was established, with a Hct range of 0.19 to 0.63 (lower limit of quantification, LLOQ, to upper limit of quantification, ULOQ). The procedure was fully validated; the bias and imprecision of quality controls (QCs) at three Hct levels and at the LLOQ and ULOQ was less than 5 and 12%, respectively. In addition, the influence of storage (pre- and postextraction), volume spotted, and punch homogeneity was evaluated. Application on DBS from patient samples (n = 111), followed by Bland and Altman, Passing and Bablok, and Deming regression analysis, demonstrated a good correlation between the "predicted Hct" and the "actual Hct". After correcting for the observed bias, limits of agreement of ±0.049 were established. Incurred sample reanalysis demonstrated assay reproducibility. In conclusion, potassium levels in extracts from 3 mm DBS punches can be used to get a good prediction of the Hct, one of the most important "unknowns" in DBS analysis.

  9. Disparate detection outcomes for anti-HCV IgG and HCV RNA in dried blood spots.

    PubMed

    Tejada-Strop, Alexandra; Drobeniuc, Jan; Mixson-Hayden, Tonya; Forbi, Joseph C; Le, Ngoc-Thao; Li, Lixia; Mei, Joanne; Terrault, Norah; Kamili, Saleem

    2015-02-01

    Dried blood spots (DBS) expedite the collection, storage and shipping of blood samples, thereby facilitating large-scale serologic studies. We evaluated the sensitivity of anti-HCV IgG testing and HCV-RNA quantitation using freshly prepared and stored DBS derived from HCV-infected patients. Protocols for elution were optimized using DBS prepared from plasma of 52 HCV-infected persons and 51 uninfected persons (control DBS), then applied to DBS from 33 chronic hepatitis C patients that had been stored at -20°C for 5 years (stored DBS). Control and stored DBS, and their corresponding plasma, were processed for anti-HCV IgG testing using the VITROS chemiluminescence assay (CIA) and the HCV 3.0 enzyme immunoassay (EIA) (Ortho-Clinical Diagnostics), and for HCV RNA quantitation by quantitative (q) RT-PCR. HCV genotyping was conducted by nucleotide sequencing. The sensitivity of CIA and EIA in control DBS was 92% and 90%, respectively, compared to 100% and 97%, respectively, in stored DBS. The sensitivity of HCV RNA detection was 88% in control DBS, compared to 36% in stored DBS. Specificity was 100% for all the assays in both control and stored DBS. Genotypes 1, 2 and 3 were detected in 16 (62%), 6 (23.1%), and 4 (15.3%) samples, respectively. Sequences generated from DBS and their corresponding plasma samples were identical. Whereas the sensitivity of anti-HCV IgG detection in stored DBS was equivalent to that in recently prepared DBS, the sensitivity of HCV RNA detection was markedly lower in stored DBS compared to recently prepared DBS. Stored DBS may be reliably used for anti-HCV detection but for HCV-RNA-based testing freshly prepared DBS is preferable to stored DBS.

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

    PubMed

    Schwartz, Alanna; Baidjoe, Amrish; Rosenthal, Philip J; Dorsey, Grant; Bousema, Teun; Greenhouse, Bryan

    2015-05-01

    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 investigated the effects of storage conditions, length of storage, and DNA extraction methods on amplification via three PCR-based assays using field samples and laboratory controls. Samples stored as DBS for 2 or more years at ambient temperature showed a significant loss of sensitivity that increased with time; after 10 years only 10% samples with parasite densities > 1,000 parasites/μL were detectable by nested polymerase chain reaction (PCR). Conversely, DBS and extracted DNA stored at -20°C showed no loss of sensitivity with time. Samples with low parasite densities amplified more successfully with saponin/Chelex compared with spin-column-based extraction, though the latter method performed better on samples with higher parasite densities stored for 2 years at ambient temperature. DNA extracted via both methods was stable after 20 freeze-thaw cycles. Our results suggest that DBS should be stored at -20°C or extracted immediately, especially if anticipating 2 or more years of storage.

  11. Governing biological material at the intersection of care and research: the use of dried blood spots for biobanking

    PubMed Central

    Douglas, Conor M.W.; van El, Carla G.; Faulkner, Alex; Cornel, Martina C.

    2012-01-01

    A series of governance issues currently surrounds the multiple uses and multiple users of dried blood spots (DBS) for research purposes. Internationally there is a discussion on storing DBS resulting from newborn screening for public health and using them as the basis for large biobank-like collections to facilitate biomedical research. If such a transformation were to be formalized, then DBS would sit at the intersection of care (ie, public health) and research, with the mechanisms through which such a collection could be managed not totally self-evident. What is more, a DBS collection raises questions about the fuzzy boundaries between privacy and anonymity; how to control or define quality control uses of DBS; medical vs nonmedical uses; as well as benefit sharing and stakeholder involvement. Our goal here is to explore some of the key questions relating to DBS governance by way of the bio-objects and bio-objectification concepts. By embracing – rather than resisting to – the blurring of boundaries and problems in categorization that have come to characterize bio-objects and bio-objectification processes recently described in this journal, we attempt to highlight some issues that might not be currently considered, and to point to some possible directions to go (or avoid). Building from our knowledge of the current DBS situation in the Netherlands, we outline questions concerning the uses, management, collection, and storage of DBS. PMID:22911534

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

    PubMed

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

    2015-01-01

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

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

    SciTech Connect

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

    1980-05-01

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

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

    PubMed Central

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

    2015-01-01

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

  15. Quantification of the HIV-integrase inhibitor raltegravir and detection of its main metabolite in human plasma, dried blood spots and peripheral blood mononuclear cell lysate by means of high-performance liquid chromatography tandem mass spectrometry.

    PubMed

    Ter Heine, R; Hillebrand, M J X; Rosing, H; van Gorp, E C M; Mulder, J W; Beijnen, J H; Huitema, A D R

    2009-02-20

    For the quantification of the HIV-integrase inhibitor raltegravir in human plasma, dried blood spots and peripheral blood mononuclear cell (PBMC) lysate, an assay was developed and validated, using liquid chromatography coupled with tandem mass spectrometry. The assay also allowed detection, but no quantification due to absence of reference substance, of the main metabolite, raltegravir-glucuronide. Raltegravir was extracted from plasma by means of protein precipitation with a mixture of methanol and acetonitrile using only 50microL plasma. Extraction from dried blood spots was performed with a simple one-step extraction with a mixture of methanol, acetonitrile and 0.2M zincsulphate in water (1:1:2, v/v/v) and extraction from cell lysate was performed in 50% methanol in water. Chromatographic separation was performed on a reversed phase C18 column (150mmx2.0mm, particle size 5microm) with a quick stepwise gradient using an acetate buffer (pH 5) and methanol, at a flow rate of 0.25mL/min. The analytical run time was 10min. The triple quadrupole mass spectrometer was operated in the positive ion-mode and multiple reaction monitoring was used for drug quantification. The method was validated over a range of 50-10,000ng/mL in plasma and dried blood spots and a range of 1-500ng/mL in PBMC lysate. Dibenzepine was used as the internal standard. The method was proven to be specific, accurate, precise and robust. Accuracies ranged from 104% to 105% in plasma, from 93% to 105% in dried blood spots and from 82% to 113% in PBMC lysate. Precision over the complete concentration range was less than 6%, 11% and 13% in plasma, dried blood spots and PBMC lysate, respectively. The method is now applied for therapeutic drug monitoring and pharmacological research in HIV-infected patients treated with raltegravir.

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

    PubMed

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

    2015-07-01

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

  17. Blood

    MedlinePlus

    ... The liquid part, called plasma, is made of water, salts, and protein. Over half of your blood is plasma. The solid part of your blood contains red blood cells, white blood cells, and platelets. Red ...

  18. Prediction of Vitamin A, Vitamin E, Selenium and Zinc Status of Periparturient Dairy Cows Using Blood Sampling During the Mid Dry Period

    PubMed Central

    Meglia, GE; Holtenius, K; Petersson, L; Öhagen, P; Waller, K Persson

    2004-01-01

    Vitamins A and E, and the trace elements selenium (Se) and zinc (Zn) are essential for the health and performance of dairy cows. Their concentrations often decrease around calving and extra supplementation is sometimes recommended at that time. However, the need for this varies, for example depending on quantity and quality of feedstuffs in the diet. The aim of this study was to measure the concentrations of serum vitamin A (S-vit A) and vitamin E (S-vit E), plasma Se (P-Se) and serum Zn (S-Zn) in blood samples taken at several time points from one month before to one month after calving, and to evaluate if a blood sample taken during the mid dry period can accurately predict the blood concentration at calving and early lactation. Dairy cows on 3 different feeding regimens during the dry period were included in the study. A significant decrease in the concentrations of S-vit A and S-vit E, and S-Zn, was observed at calving, and P-Se was significantly lower during the dry period and at calving than in early lactation. The blood concentrations of S-vit E and P-Se in the mid dry period significantly predicted the occurrence of values considered marginal or deficient at the time of calving. The data indicate that a mid dry period concentration of ≥5.4 mg/l of S-vit E and ≥0.09 mg/l of P-Se will result in a 90% chance that the cow stays above marginal levels at calving given that a feed of the same quality is offered. PMID:15535092

  19. Cinemicrographic specimen housing

    NASA Technical Reports Server (NTRS)

    Wilkins, J. R.

    1979-01-01

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

  20. DNA extraction from herbarium specimens.

    PubMed

    Drábková, Lenka Záveská

    2014-01-01

    With the expansion of molecular techniques, the historical collections have become widely used. Studying plant DNA using modern molecular techniques such as DNA sequencing plays an important role in understanding evolutionary relationships, identification through DNA barcoding, conservation status, and many other aspects of plant biology. Enormous herbarium collections are an important source of material especially for specimens from areas difficult to access or from taxa that are now extinct. The ability to utilize these specimens greatly enhances the research. However, the process of extracting DNA from herbarium specimens is often fraught with difficulty related to such variables as plant chemistry, drying method of the specimen, and chemical treatment of the specimen. Although many methods have been developed for extraction of DNA from herbarium specimens, the most frequently used are modified CTAB and DNeasy Plant Mini Kit protocols. Nine selected protocols in this chapter have been successfully used for high-quality DNA extraction from different kinds of plant herbarium tissues. These methods differ primarily with respect to their requirements for input material (from algae to vascular plants), type of the plant tissue (leaves with incrustations, sclerenchyma strands, mucilaginous tissues, needles, seeds), and further possible applications (PCR-based methods or microsatellites, AFLP).

  1. Measurement of succinyl-carnitine and methylmalonyl-carnitine on dried blood spot by liquid chromatography-tandem mass spectrometry.

    PubMed

    Rizzo, Cristiano; Boenzi, Sara; Inglese, Rita; la Marca, Giancarlo; Muraca, Maurizio; Martinez, Tegra Barreiro; Johnson, David W; Zelli, Eleonora; Dionisi-Vici, Carlo

    2014-02-15

    Methylmalonic aciduria (MMA) is one of the most frequent organic acidurias, a class of diseases caused by enzymatic defects mainly involved in the catabolism of branched-chain amino acids. Recently, mild MMA and C4-dicarboxylyl-carnitine (C4DC-C) accumulation have been reported in patients carrying mutation in genes encoding the α-subunit (SUCLG1) and the β-subunit (SUCLA2) of the ADP-forming succinyl-CoA synthetase (SCS). We developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify in dried blood spot the two isobaric compounds of C4DC-C, succinyl-carnitine and methylmalonyl-carnitine, to allow the differential diagnosis between classical MMA and SCS-related defects. This method, with an easy liquid-phase extraction and derivatization procedure, has been validated to demonstrate the specificity, linearity, recovery, lowest limit of quantification (LLOQ), accuracy and precision for quantitative determination of blood succinyl-carnitine and methylmalonyl-carnitine. The assay was linear over a concentration range of 0.025-10 μmol/L and achieved the LLOQ of 0.025 μmol/L for both metabolites. The average slope, intercept, and coefficient of linear regression (r(2)) were respectively: 0.3389 (95% confidence interval 0.2888-0.3889), 0.0113 (95% confidence interval -0.0157 to 0.0384), 0.9995 (95% confidence interval 0.9990-1.0000) for succinyl-carnitine and 0.5699 (95% confidence interval 0.5263-0.6134), 0.0319 (95% confidence interval -0.0038 to 0.0677), 0.9997 (95% confidence interval 0.9995-1.0000) for methylmalonyl-carnitine. Within-day and between-day coefficients of variation (CV) were 1.94% and 3.19% for succinyl-carnitine and 3.21%, and 2.56 for methylmalonyl-carnitine. This method is accurate and provides a new tool to differentiate patients with classical methylmalonic acidemia from those with SCS-related defects.

  2. Dry Blood Spots a Reliable Method for Measurement of Hepatitis B Viral Load in Resource-Limited Settings

    PubMed Central

    Stene-Johansen, Kathrine; Yaqoob, Nadeem; Overbo, Joakim; Aberra, Hanna; Desalegn, Hailemichael; Berhe, Nega; Johannessen, Asgeir

    2016-01-01

    Background & Aims Hepatitis B virus (HBV) quantification is essential in the management of chronic hepatitis B, both to determine treatment eligibility and in the monitoring of treatment effect. This test, however, is rarely available in resource-limited settings due to high costs and stringent requirements for shipment and storage of plasma. Dried Blood Spots (DBS) can be a convenient alternative to plasma, but its use for HBV monitoring has not been investigated under real-life conditions in Africa. Methods The performance of DBS in HBV quantification was investigated using a modified commercial test (Abbott RealTime HBV assay). Paired DBS and plasma samples were collected from an HBV positive cohort in Addis Ababa, Ethiopia. DBS were stored at ambient temperature for 4–39 days before shipment to the laboratory. Results Twenty-six paired samples were selected covering the total range of quantification, from 2.14 log IU/ml to >7 log IU/ml. HBV was detected in 21 of 21 (100%) DBS from patients with a corresponding plasma viral load above 2.70 log IU/ml. The mean difference between plasma and DBS was 0.59 log IU/ml, and the correlation was strong (R2 = 0.92). In stability studies there was no significant change in DBS viral load after storage at room temperature for up to 12 weeks. Conclusions This study suggests that DBS can be a feasible and reliable alternative to plasma for quantification of HBV in resource-limited settings. DBS can expand access to antiviral treatment for patients in low- and middle-income countries. PMID:27820845

  3. A cyclic-olefin-copolymer microfluidic immobilized-enzyme reactor for rapid digestion of proteins from dried blood spots.

    PubMed

    Wouters, Bert; Dapic, Irena; Valkenburg, Thalassa S E; Wouters, Sam; Niezen, Leon; Eeltink, Sebastiaan; Corthals, Garry L; Schoenmakers, Peter J

    2017-03-31

    A critical step in the bottom-up characterization of proteomes is the conversion of proteins to peptides, by means of endoprotease digestion. Nowadays this method typically uses overnight digestion and as such represents a considerable bottleneck for high-throughput analysis. This report describes protein digestion using an immobilized-enzyme reactor (IMER), which enables accelerated digestion times that are completed within seconds to minutes. For rapid digestion to occur, a cyclic-olefin-copolymer microfluidic reactor was constructed containing trypsin immobilized on a polymer monolithic material through a 2-vinyl-4,4-dimethylazlactone linker. The IMER was applied for the rapid offline digestion of both singular protein standards and a complex protein mixture prior to liquid chromatography-electrospray ionisation-tandem mass spectrometry (LC-ESI-MS/MS) analysis. The effects of protein concentration and residence time in the IMER were assessed for protein standards of varying molecular weight between 11 and 240kDa. Compared to traditional in-solution digestion, IMER-facilitated protein digestion at room temperature for 5min yielded similar results in terms of sequence coverage and number of identified peptides. Good repeatability was demonstrated with a relative standard deviation of 6% for protein-sequence coverage. The potential of the IMER was also demonstrated for a complex protein mixture in the analysis of dried blood spots. Compared to a traditional workflow a similar number of proteins could be identified, while reducing the total analysis time from 22.5h to 4h and importantly omitting the sample-pre-treatment steps (denaturation, reduction, and alkylation). The identified proteins from two workflows showed similar distributions in terms of molecular weight and hydrophobic character.

  4. Simple, Sensitive, and Specific Detection of Human Immunodeficiency Virus Type 1 Subtype B DNA in Dried Blood Samples for Diagnosis in Infants in the Field

    PubMed Central

    Beck, Ingrid A.; Drennan, Kathryn D.; Melvin, Ann J.; Mohan, Kathey M.; Herz, Arnd M.; Alarcón, Jorge; Piscoya, Julia; Velázquez, Carlos; Frenkel, Lisa M.

    2001-01-01

    The detection of virus is used to diagnose human immunodeficiency virus type 1 (HIV-1) infection in infants due to the persistence of maternal antibodies for a year or more. An HIV-1 DNA PCR assay with simple specimen collection and processing was developed and evaluated. Whole blood was collected on filter paper that lysed cells and bound the DNA, eliminating specimen centrifugation and extraction procedures. The DNA remained bound to the filter paper during PCR amplification. Assays of copy number standards showed reproducible detection of 5 to 10 copies of HIV-1 in 5 μl of whole blood. The sensitivity of the assay did not decrease after storage of the standards on filter paper for 3 months at room temperature or after incubation at 37 or 45°C for 20 h. The primers used for nested PCR of the HIV-1 pol gene amplified templates from a reference panel of multiple HIV-1 subtypes but did not amplify a subtype A or a subtype C virus from children living in Seattle. The assay had a sensitivity of 98.4% and a specificity of 98.3% for testing of 122 specimens from 35 HIV-1-infected and 16 uninfected children and 43 seronegative adults living in Washington. The assay had a sensitivity of 99% and a specificity of 100% for testing of 102 HIV-1-positive (as determined by enzyme immunoassay) Peruvian women and 6 seropositive and 34 seronegative infants. This assay, with adsorption of whole blood to filter paper and no specimen processing, provides a practical, economical, sensitive, and specific method for the diagnosis of HIV-1 subtype B infection in infants. PMID:11136743

  5. Urine culture - catheterized specimen

    MedlinePlus

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

  6. NASA Biological Specimen Repository

    NASA Technical Reports Server (NTRS)

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

    2010-01-01

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

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

    PubMed

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

    2016-05-01

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

  8. Blood

    MedlinePlus

    ... that die or are lost from the body. White Blood Cells White blood cells (WBCs, and also ... of severe pain. previous continue Diseases of the White Blood Cells Neutropenia (pronounced: new-truh-PEE-nee- ...

  9. The ratio of alpha-galactosidase to beta-glucuronidase activities in dried blood for the identification of female Fabry disease patients.

    PubMed

    Lukacs, Z; Keil, A; Kohlschütter, A; Beck, M; Mengel, E

    2005-01-01

    Female heterozygous patients with Fabry disease are difficult to identify because of the relatively high residual activity of alpha-galactosidase. We systematically evaluated the activities of various lysosomal enzymes in dried blood samples from Fabry patients and found that the beta-glucuronidase activity was frequently elevated. The ratio of alpha-galactosidase to beta-glucuronidase proved to be a helpful tool for the diagnosis of female Fabry disease patients.

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

    PubMed Central

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

    2016-01-01

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

  11. Effects of rumen-protected choline and dry propylene glycol on feed intake and blood parameters for Holstein dairy cows in early lactation.

    PubMed

    Chung, Y-H; Brown, N E; Martinez, C M; Cassidy, T W; Varga, G A

    2009-06-01

    A 6 x 6 Latin square design was used to test 3 sets of comparisons simultaneously to study response in dry matter intake, milk yield, and blood parameters to propylene glycol (PG) supplementation delivered by 2 methods [incorporating PG into the total mixed ration (TMR) vs. top dressing; comparison I]; individual or combined dietary choline and PG supplementation as a 2 x 2 factorial (comparison II); or increasing amounts of dietary choline (comparison III). Six multiparous (lactation number = 1.5 +/- 0.8 SD) Holstein dairy cows were at 41 d in milk (+/- 9 SD) at the start of the experiment. Propylene glycol used was a dry product containing 65% PG, and choline was a rumen-protected choline product (RPC; estimated to be 50% rumen-protected) containing 50% choline chloride. In comparison I, treatments compared were 1) control: no PG; 2) PG-TMR: 250 g/d of dry PG (corresponding to 162.5 g/d of PG) incorporated into the TMR; and 3) PG-top dress: 250 g/d of dry PG top-dressed onto the TMR. In comparison II, treatments compared were 1) control: no PG and no RPC; 2) PG: 250 g/d of dry PG incorporated into the TMR; 3) RPC: 50 g/d of RPC top-dressed onto the TMR; and 4) PG+RPC: combination of treatments 2 and 3. In comparison III, treatments compared were 0, 25, and 50 g/d of RPC top-dressed onto the TMR. Each experimental period lasted 10 d with 9 d of adaptation followed by 1 d of serial blood sampling. Dry matter intake and milk yield were recorded daily. During the serial blood sampling, jugular blood was sampled every 20 min for the first 4 h and at 8 and 12 h after treatment administration. Results obtained from comparison I showed that feeding 250 g/d of PG as a dry product decreased plasma beta-hydroxybutyrate (BHBA) concentration (mean +/- SEM) from 701 +/- 81 (control) to 564 +/- 76 micromol/L without affecting serum insulin, plasma glucose, or plasma nonesterified fatty acid concentrations. Top-dressing PG decreased plasma BHBA concentrations more than by

  12. Detection of binding of a synthetic granzyme B-like peptide fluorescent conjugate within platelet-like structures in cancer-related peripheral blood specimens and tissue sections.

    PubMed

    Lo, Wai Chun Jennifer; Luther, Donald Gene

    2014-09-01

    Platelets and cytotoxic T lymphocytes (CTL) are important whole blood components in peripheral blood. Studies have shown that platelets, from precursor megakaryocytes, are significant factors in cancer prognosis, cancer progression, and metastasis; but a direct platelet-cancer relationship remains unclear. CTL play an essential role in cancer surveillance by inducing cancer cell death with granzyme B. A recent report has shown the presence of binding targets with binding affinity to a synthetic granzyme B-like peptide fluorescent conjugate (GP1R) in different types of cancer cells grown in vitro. It suggests that these binding targets may serve as a "universal-pathologic-biomarker". It is not known if similar biomarkers may be present in platelets of cancer patients. We show with fluoroscopic images that GP1R can bind to binding targets: 1) within platelets in methanol-fixed whole blood smears of patients with breast cancer and lung cancer, and 2) within platelet-like structures in formalin-fixed-paraffin-embedded (FFPE) nude mouse xenogeneic breast tumor tissues. Samples without cancer-association displayed no discernible GP1R-binding in platelet-like structures. Our data demonstrate for the first time that a similar "universal-pathologic-biomarker" detectable by GP1R-binding is present in circulating platelets of cancer patients. The data depict a co-existence of animal-platelets and human-breast cancer cells, both have a common pathologic biomarker detectable by GP1R, in the tumor growth. The fluoroscopic images indicate a visual direct connection between pathologic platelets and cancer. These preliminary results may lead to developments of novel platelet-based cancer diagnostics and therapeutics and a better understanding of the potential multifunction of GP1R and its relationship to megakaryocytes and PD1.

  13. NASA Biological Specimen Repository

    NASA Technical Reports Server (NTRS)

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

    2009-01-01

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

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

    PubMed

    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

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

    PubMed

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

    2014-07-01

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

  16. Short communication: Effect of heat stress during the dry period on gene expression in mammary tissue and peripheral blood mononuclear cells.

    PubMed

    Tao, S; Connor, E E; Bubolz, J W; Thompson, I M; do Amaral, B C; Hayen, M J; Dahl, G E

    2013-01-01

    Heat stress (HT) during the dry period compromises mammary gland development, decreases future milk production, and impairs the immune status of dairy cows. Our objective was to evaluate the effect of cooling HT cows during the dry period on gene expression of the mammary gland and peripheral blood mononuclear cells (PBMC). Cows were dried off 46 d before their expected calving and assigned to 2 treatments, HT or cooling (CL). Cows in the CL group were cooled with sprinklers and fans whereas HT cows were not. After parturition, all cows were housed in a freestall barn with cooling. The PBMC were isolated at dry-off and at -20, 2, and 20 d relative to calving from a subset of cows (HT, n=9; CL, n=10), and mammary biopsies were taken at the same intervals (HT, n=7; CL, n=6) for RNA extraction. Gene expression was assessed using a custom multiplex gene expression assay based on traditional reverse transcription-PCR. Genes involved in prolactin (PRL) signaling [PRL receptor long form, PRL receptor short form, suppressor of cytokine signaling (SOCS)2, SOCS3, IGF2, IGF binding protein 5, and cyclin D1], fatty acid metabolism (acetyl-CoA carboxylase α (ACACA) and lipoprotein lipase (LPL)], and IGF1 were evaluated in mammary tissue, and genes related to fatty acid metabolism [ACACA, fatty acid synthase (FASN), and LPL], cytokine production [IL6, IL8, and tumor necrosis factor (TNF)], and IGF1 were evaluated in PBMC. No differences were observed in PRL signaling or fatty acid metabolism gene expression in the mammary gland. In PBMC, HT cows had greater mRNA expression of IGF1 and TNF during the transition period relative to CL and upregulated IL8 and downregulated FASN mRNA expression at 2 d relative to calving. We conclude that cooling HT cows during the dry period alters expression of genes involved in cytokine production and lipid metabolism in PBMC.

  17. Tandem Mass Spectrometry Assays of Palmitoyl Protein Thioesterase 1 and Tripeptidyl Peptidase Activity in Dried Blood Spots for the Detection of Neuronal Ceroid Lipofuscinoses in Newborns

    PubMed Central

    2015-01-01

    We report new substrates for quantitative enzyme activity measurements of human palmitoyl protein thioesterase (PPT1) and tripeptidyl peptidase (TPP1) in dried blood spots from newborns using tandem mass spectrometry. Deficiencies in these enzyme activities due to inborn errors of metabolism cause neuronal ceroid lipofuscinoses. The assays use synthetic compounds that were designed to mimic the natural substrates. Incubation produces nanomole quantities of enzymatic products per a blood spot that are quantified by tandem mass spectrometry using synthetic internal standards and selected reaction monitoring. The assays utilize a minimum steps for sample workup and can be run in a duplex format for the detection of neuronal ceroid lipofuscinoses or potentially multiplexed with other mass spectrometry-based assays for newborn screening of lysosomal storage disorders. PMID:25019629

  18. High blood sugar

    MedlinePlus

    ... High blood glucose - self-care; Diabetes - high blood sugar ... Symptoms of high blood sugar can include: Being very thirsty or having a dry mouth Having blurry vision Having dry skin Feeling weak or tired ...

  19. Dry Mouth

    MedlinePlus

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

  20. The Effect of the Dried-Bonito Broth on Blood Pressure, 8-Hydroxydeoxyguanosine (8-OHdG), an Oxidative Stress Marker, and Emotional States in Elderly Subjects.

    PubMed

    Umeki, Youko; Hayabuchi, Hitomi; Hisano, Manami; Kuroda, Motonaka; Honda, Masashi; Ando, Bunei; Ohta, Masanori; Ikeda, Masaharu

    2008-11-01

    Dried-bonito broth (DBB, katsuo-bushi dashi) is commonly used in Japanese cuisine, and is also used as a traditional remedy for recovery from fatigue and improvement of blood circulation. To clarify the effect of DBB on blood pressure, oxidative stress and emotional states, a randomized crossover human trial was performed. Twenty-seven elderly Japanese subjects ingested DBB or water for one month. Measurement of blood pressure and urinary 8-hydroxydeoxyguanosine (8-OHdG) and evaluation of emotional states were performed before and after the ingestion periods. The changes in systolic blood pressure (SBP) during DBB ingestion was significantly lower than that during water ingestion (p = 0.037). Urinary 8-OHdG significantly decreased during DBB ingestion (p = 0.0002). Evaluation of emotional states indicated that composure significantly improved during DBB ingestion (p = 0.034). These results suggest that the daily ingestion of DBB lower SBP, reduce urinary 8-OHdG and might improve emotional states in elderly subjects.

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

    PubMed

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

    2015-02-01

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

  2. Tandem Mass Spectrometry for the Direct Assay of Lysosomal Enzymes in Dried Blood Spots: Application to Screening Newborns for Mucopolysaccharidosis I

    PubMed Central

    Blanchard, Sophie; Sadilek, Martin; Scott, C. Ronald; Turecek, Frantisek; Gelb, Michael H.

    2008-01-01

    Background Treatments now available for Mucopolysaccharidosis I require early detection for optimum therapy. Therefore, we have developed an assay appropriate for newborn screening of the activity of the relevant enzyme, α-l-iduronidase. Methods We synthesized a new α-l-iduronidase substrate that can be used to assay the enzyme by use of tandem mass spectrometry together with an internal standard. The assay uses a dried blood spot on a newborn screening card as the enzyme source. The assay protocol uses a simple liquid-liquid extraction step prior to mass spectrometry. We optimized enzyme reaction conditions and procedures for the assay, including the concentration of substrate, the reaction pH, the incubation time, and mass spectrometer operation. We also assessed inter- and intra-assay imprecision. Results When the assay was tested on dried blood spots, the α-l-iduronidase activity measured for 5 patients with Mucopolysaccharidosis I was well below the interval found for 10 randomly chosen newborns. Inter- and intra-assay imprecision were less than 10 %. The synthesis of the α-l-iduronidase substrate is practical on a scale needed to support newborn screening demands. Conclusions This newly developed tandem mass spectrometry assay has the potential to be adopted for newborn screening of Mucopolysaccharidosis I. It presents advantages compared to the one previously published from this laboratory and has the potential to be performed in a multiplex fashion with several lysosomal enzymes relevant to treatable lysosomal storage diseases. PMID:19042989

  3. Comparative evaluation of Amplicor HIV-1 DNA test, version 1.5, by manual and automated DNA extraction methods using venous blood and dried blood spots for HIV-1 DNA PCR testing.

    PubMed

    Nsojo, Anthony; Aboud, Said; Lyamuya, Eligius

    2010-10-01

    Human immunodeficiency virus (HIV) DNA polymerase chain reaction (PCR) test using venous blood sample has been used for many years in low resource settings for early infant diagnosis of HIV infection in children less than 18 months. The aim of this study was to evaluate and compare the performance characteristics of Amplicor HIV-1 DNA assay version 1.5 following processing of venous blood and dried blood spot (DBS) samples by Roche manual DNA extraction and automated Roche MagNA Pure LC instrument (MP) for HIV-1 DNA PCR testing in Dar es Salaam, Tanzania, in order to scale up early infant diagnosis of HIV infection in routine practice. Venous blood samples from children under 18 months born to HIV-infected mothers between January and April 2008 were collected. Venous blood was used to prepare cell pellet and DBS samples. DNA extractions by manual procedure and MP were performed each on cell pellet, venous blood and DBS samples and tested by Amplicor HIV-1 DNA assay. Of 325 samples included, 60 (18.5%) were confirmed HIV-infected by manual extraction performed on cell pellets. Sensitivity of the assay following MP processing of venous blood was 95% (95% CI; 86.1-99.0%) and 98.3% (95% CI; 91.1 to 99.9%) for the manual extraction and processing by MP performed on DBS samples. Specificity of the assay with all DNA extraction methods was 99.6% (95% CI; 97.9 to 100%). Performance of the assay with Roche manual extraction and processing by MP on DBS samples compared well with Roche manual extraction performed on cell pellet samples. The choice of DNA extraction method needs to be individualized based on the level of laboratory facility, volume of testing and cost benefit analysis before it is adopted for use.

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

    PubMed

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

    2015-11-01

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

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

    EPA Science Inventory

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

  6. Blood gases and oximetry: calibration-free new dry-chemistry and optical technology for near-patient testing.

    PubMed

    Boalth, N; Wandrup, J; Larsson, L; Frischauf, P A; Lundsgaard, F C; Andersen, W L; Jensen, N; Singer, R; Troldborg, C P; Lunding, G

    2001-05-01

    The first calibration-free Near-Patient-Testing instrument (NPT7) for blood gases, pH and oximetry has been developed. With cartridges of 30 single-use cuvettes, the NPT7 needs no preparation prior to sample aspiration, no manual calibration, and no maintenance apart from paper and cartridge changes and regulatory quality control. Each cuvette measures pCO2, pO2, pH, total hemoglobin (ctHb), oxygen saturation (sO2), fractions of carboxyhemoglobin (FCOHb) and methemoglobin (FMetHb) on 95 microl whole blood with a 110-s measuring cycle. The measurement principles are as follows: pCO2-three-wavelength infrared spectroscopy of dissolved CO2; pO2-measurement of O2-induced changes in the decay time of phosphorescence; pH-the absorbance spectra change of an azo-dye color indicator; and oximetry is performed with a 128-wavelength spectrophotometer. We determined the within and between instrument variations with tonometered whole blood on seven prototype instruments, using between one and five control levels per analyte. The 95% analytical performance limits: +/-(/Bias/ +2 xS(T)) in the NPT7 instrument matched the analytical performance criteria for the measured quantities as defined by AACC guidelines. The application of these optical measuring methods for blood gases, pH and oximetry in single-use devices introduces a new concept into point-of-care testing (POCT), where preanalytical activities otherwise associated with instrument preparation are eliminated.

  7. 37 CFR 2.59 - Filing substitute specimen(s).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

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

  8. Development of an LC-MS/MS method for determination of bicalutamide on dried blood spots: application to pharmacokinetic study in mice.

    PubMed

    P S, Suresh; Vijay Kumar, S; Kumar, Avinash; Mullangi, Ramesh

    2015-02-01

    A rapid and highly sensitive assay method has been developed and validated for the estimation of bicalutamide (BCL) on mouse dried blood spots (DBS) using liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the negative-ion mode. The assay procedure involves a simple liquid extraction of BCL and tolbutamide (internal standard, IS) from mouse blood DBS cards using tert-butyl methyl ether. Chromatographic separation was achieved with 5 mm ammonium acetate (pH 6.5)-acetonitrile (35:65, v/v) at a flow rate of 0.60 mL/min on an Atlantis dC18 column with a total run time 3.0 min. The MS/MS ion transitions monitored were 428.80 → 254.70 for BCL and 269.00 → 169.60 for IS. Method validation was performed as per regulatory guidelines. A linear response function was observed from 0.92 to 1911 ng/mL for BCL in mouse blood. The intra- and inter-day precisions were in the ranges of 1.86-12.5 and 3.19-10.8%, respectively. This novel DBS method has been applied to a pharmacokinetic study in mice.

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

    PubMed

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

    2010-06-01

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

  10. Comparative evaluation of newborn bloodspot specimen cards by experienced laboratory personnel and by an optical scanning instrument.

    PubMed

    Dantonio, Paul D; Stevens, Germaine; Hagar, Arthur; Ludvigson, David; Green, Daron; Hannon, Harry; Vogt, Robert F

    2014-01-01

    A major factor in determining the suitability of a dried blood spot (DBS) specimen is the subjective nature of evaluation by laboratory personnel. Using newborn screening DBS specimen cards as they were submitted to a public health NBS program, we conducted a systematic pilot study of DBS evaluation by multiple experienced laboratory personnel (ELP) and by an automated optical scanning instrument (OSI) (CardScan (tm), BSD Robotics). OSI confirmed the satisfactory status of all newborn DBS specimen cards that passed initial review by the first ELP. Among the questionable cards selected for further review, 58% passed multiple ELP consensus assessment, and 62% passed OSI evaluation. The overall agreement between ELP and OSI was 86%. Among questionable specimen cards, ELP and OSI were more strongly correlated when multiple ELP assessment was unanimous. We conclude that subjective assessment by ELP is essential and that OSI evaluation is a useful adjunct when ELP assessment does not reach consensus. OSI further allows the selection of optimal locations for punching DBS from unsatisfactory or questionable specimens, optimizing the quality of interim analyses that may be conducted while repeat specimens are being collected. Instrument evaluation of specimen cards would also be valuable as an independent reference method for training laboratory and specimen collection personnel. OSI technology merits further studies to confirm and extend our findings.

  11. Blood

    MedlinePlus

    ... increased red blood cell destruction can affect teens: G6PD deficiency. G6PD is an enzyme that helps to protect ... can cause red cells to hemolyze, or burst. G6PD deficiency is a common hereditary disease among people of ...

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

    PubMed Central

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

    2014-01-01

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

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

    PubMed

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

    2015-03-01

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

  14. Short communication: HIV type 1 tropism determination in a novel dried blood spot membrane and the use of a mixture of outer nested polymerase chain reaction primers.

    PubMed

    Lavigne, Shirlene; Santos, Carlos; Arif, Muhammad Shoaib; Reis, Alexanda; Samer, Sadia; Dos Santos, Cintia Vilhena; Diaz, Ricardo Sobhie

    2014-02-01

    Genotropism was determined in 608 Brazilian samples collected in dried blood spots using Polyethersulfone collection cards. Patients were infected by subtype B (88.8%), F (5.6%), C (3.3%), A (1.8%), and G (0.5%). All patients were exposed to three classes of antiretrovirals, and 59.8% of the samples harbored R5 viruses, 35% non-R5-tropic viruses, and 5.1% harbored mixtures of R5 and non-R5-tropic viruses, with non-R5 more prevalent among clade B-infected patients as compared to non-B (42.8% versus 19.1%; p<0.0003). A strategy using a mixture of outer nested polymerase chain reaction (PCR) primers reduced the number of negative PCR results from 39% to 19%.

  15. Single step detection of HIV-1 proviral DNA and housekeeping β-actin gene from dried blood spots by a monoplex polymerase chain reaction.

    PubMed

    Choudhary, Ipsita; Chimanpure, Vaishali; Patil, Ajit; Mukhopadhyaya, Robin; Paranjape, Ramesh; Bhattacharya, Jayanta

    2013-01-01

    There is a growing need for developing a simple, rapid, reliable and cost effective method for detection of HIV-1 for early diagnosis of the infection especially in developing countries and in resource limited settings. A method for simultaneous detection of the HIV-1 p17 gene and the house keeping human β-actin gene from dried blood spots (DBS) by a monoplex polymerase chain reaction (PCR) is described. Genomic DNA was extracted from 40 HIV-1 positive and 40 HIV-1 negative DBS and used as templates to amplify both the HIV-1 p17 and β-actin genes simultaneously under the same cycling condition by a single round PCR. This method of detection of HIV-1 may provide a simple, rapid and cost effective alternative in resource limited settings; however, it would require testing a larger number of samples before widespread use.

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

    PubMed Central

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

    2015-01-01

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

  17. Impact of salt reduction on biogenic amines, fatty acids, microbiota, texture and sensory profile in traditional blood dry-cured sausages.

    PubMed

    Laranjo, Marta; Gomes, Ana; Agulheiro-Santos, Ana Cristina; Potes, Maria Eduarda; Cabrita, Maria João; Garcia, Raquel; Rocha, João Miguel; Roseiro, Luísa Cristina; Fernandes, Maria José; Fraqueza, Maria João; Elias, Miguel

    2017-03-01

    Meat industry needs to reduce salt in their products due to health issues. The present study evaluated the effect of salt reduction from 6% to 3% in two Portuguese traditional blood dry-cured sausages. Physicochemical and microbiological parameters, biogenic amines, fatty acids and texture profiles and sensory panel evaluations were considered. Differences due to salt reduction were perceptible in a faint decline of water activity, which slightly favoured microbial growth. Total biogenic amines content ranged from 88.86 to 796.68mgkg(-1) fresh matter, with higher amounts, particularly of cadaverine, histamine and tyramine, in low-salt products. Still, histamine and other vasoactive amines remained at low levels, thus not affecting consumers' health. Regarding fatty acids, no significant differences were observed due to salt. However, texture profile analysis revealed lower resilience and cohesiveness in low-salt products, although no textural changes were observed by the sensory panel. Nevertheless, low-salt sausages were clearly preferred by panellists.

  18. Fast method for simultaneous quantification of tamoxifen and metabolites in dried blood spots using an entry level LC-MS/MS system.

    PubMed

    Tré-Hardy, Marie; Capron, Arnaud; Antunes, Marina Venzon; Linden, Rafael; Wallemacq, Pierre

    2016-11-01

    The purpose of this study was to develop and validate a new liquid chromatography-tandem mass spectrometric (LC-MSMS) assay for the simultaneous quantification of tamoxifen (TAM) and its main therapeutically active metabolites, N-desmethyltamoxifen (NDT), 4-hydroxytamoxifen (4HT) and endoxifen (END) in dried blood spots. Ultrasound assisted methanolic extraction was used for TAM and metabolites extraction from dried blood spot. After evaporation and methanol reconstitution, the extract was injected into a LC-MSMS system. Reversed phase chromatography was performed on a C18 grafted column in gradient mode. TAM, metabolites, and internal standard (diazepam-d5; IS) were identified in positive electrospray ionization mode using m/z transition of 372.5>72.1 (TAM); 374.23>58.10 (END); 358.27>58.10 (NDT); 388.23>44.80 (4HT) and 290.00>198.00 (IS). Total analytical run time was 6.5min. Assay was linear from 1 to 500ng/mL for all substances and presented intra and inter-assay precision and accuracy <15%. TAM, NDT, 4HT and END limits of quantification and detection were of 1 and 0.5ng/mL; 1 and 3ng/mL; 1.7 and 3ng/mL; 0.6 and 2ng/mL, respectively. Recovery ranged from 83.8 to 96.3% with matrix effect ranged from 4.3 to 29.8% for TAM and its metabolites. Hematocrit value ≤40% appeared to negatively influence accuracy of the method. In conclusion, the method described here is somewhat accessible, relatively fast, sensitive and selective with no interference. This assay might be used to investigate the level of TAM and its metabolites in DBS for therapeutic drug monitoring purposes.

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

    PubMed

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

    2016-03-01

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

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

    PubMed Central

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

    2016-01-01

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

  1. A simple dried blood spot method for therapeutic drug monitoring of the tricyclic antidepressants amitriptyline, nortriptyline, imipramine, clomipramine, and their active metabolites using LC-MS/MS.

    PubMed

    Berm, E J J; Paardekooper, J; Brummel-Mulder, E; Hak, E; Wilffert, B; Maring, J G

    2015-03-01

    Therapeutic drug monitoring (TDM) of tricyclic antidepressants (TCAs) is considered useful in patients with major depressive disorder, since these drugs display large individual differences in clearance, and the therapeutic windows of these drugs are relatively small. We developed an assay for determination of amitriptyline (ATP), nortriptyline (NTP), imipramine (IMP), desipramine (DSP) clomipramine (CMP) and desmethyl-clomipramine (DCMP) in dried blood spots (DBS). A fast and robust LC-MS/MS method was developed and analytically validated for simultaneous determination of ATP, NTP, IMP, DSP, CMP, and DCMP in DBS. Six mm circles were punched out from DBS collected on Whatman DMPK-C paper and mixed with acetonitrile: methanol 1:3 containing the internal standard. The extract was analyzed by LC-MS/MS. Total LC-MS/MS runtime was 4.8 min. The assay was linear in the range 20-500 µg/L for all compounds. Overall-assay accuracy and precision were<20% for the lower limit of quantification (LLOQ), except for CMP (CV=22.3%), and <15% at other concentrations. The initial LLOQ was 20 µg/L however for CMP and DMCP it was increased to 40 µg/L. The blood volume per spot did not influence the results, but a low hematocrit (≤ 30%) was associated with a >15% negative bias for all compounds. Punching at the perimeter of the blood spot instead of the center was associated with a positive bias. A good correlation was found between patients plasma and DBS samples of ATP, NTP and DMCP, but not for CMP. In addition, proportional differences were found. This LC-MS/MS method was analytically validated for determination of TCAs in DBS. Future validation will focus on the clinical application of the method.

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

    PubMed

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

    2016-01-25

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

  3. Temporal Trends of Polybrominated Diphenyl Ethers (PBDEs) in the Blood of Newborns from New York State during 1997 through 2011: Analysis of Dried Blood Spots from the Newborn Screening Program

    PubMed Central

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

    2013-01-01

    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 1,224 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 BDEs −47, −99, and −100, respectively. A significant correlation was found among the concentrations of three major congeners (p < 0.001). PBDE concentrations were similar during 1997 through 2002 and, thereafter, decreased significantly, which was similar to the trends observed for perfluorinated compounds (PFCs) in DBS samples. Occurrence of PBDEs in the whole blood of newborns confirms that these compounds do cross the placental barrier. PMID:23755886

  4. Preparation of Articular Cartilage Specimens for Scanning Electron Microscopy.

    PubMed

    Stupina, T A

    2016-08-01

    We developed and adapted a technology for preparation of articular cartilage specimens for scanning electron microscopy. The method includes prefixation processing, fixation, washing, and dehydration of articular cartilage specimens with subsequent treatment in camphene and air-drying. The technological result consists in prevention of deformation of the articular cartilage structures. The method is simpler and cheaper than the known technologies.

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

    PubMed

    Hollegaard, Mads Vilhelm; Grauholm, Jonas; Nielsen, Ronni; Grove, Jakob; Mandrup, Susanne; Hougaard, David Michael

    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 neonatal DBSS (neoDBSS) stored at -20°C in the Danish Newborn Screening Biobank. The reference samples were genotyped using an Illumina Omni2.5M array, and all samples were sequenced on a HighSeq2000 Paired-End flow cell. First, we compared the array single nucleotide polymorphism (SNP) genotype data to the single nucleotide variation (SNV) calls from the WGS and WES SNV calls. We also compared the WGS and WES reference sample SNV calls to the DBSS SNV calls. The overall performance of the archived DBSS was similar to the whole blood reference sample. Plotting the error rates relative to coverage revealed that the error rates of DBSS were similar to that of their reference samples. SNVs called with a coverage<×8 had error rates between 1.5 and 35%, whereas the error rates of SNVs called with a coverage≥8 were <1.5%. In conclusion, the wgaDNA amplified from both new and old neonatal DBSS perform as well as their whole-blood reference samples with regards to error rates, strongly indicating that neonatal DBSS collected shortly after birth and stored for decades comprise an

  6. Chemokine receptor CCR5 Delta 32 genetic analysis using multiple specimen types and the NucliSens Basic Kit.

    PubMed

    Tetali, S; Lee, E M; Kaplan, M H; Romano, J W; Ginocchio, C C

    2001-09-01

    Resistance to HIV-1 infection and delayed disease progression have been associated with a 32-bp deletion (Delta32) in the gene encoding the CCR5 chemokine receptor. In the present study we describe the modification of a nucleic acid sequence-based amplification (NASBA)-based CCR5 genotyping assay for a NucliSens Basic Kit (Organon Teknika, Durham, N.C.) format using a new target-specific sandwich oligonucleotide detection methodology. The new method permitted the use of generic electrochemiluminescent probes supplied in the NucliSens Basic Kit, whereas the original NASBA method required expensive target-specific ruthenium detection probes. The Basic Kit CCR5 Delta32 genotypic analysis was in 100% concordance with both the original NASBA assay and DNA PCR results. This study also evaluated the use of multiple specimen types, including peripheral blood mononuclear cells (PBMC), whole blood, dried blood spots, buccal scrapings, and plasma, for CCR5 genotype analysis. The sensitivities of the three assays were comparable when PBMC or whole blood was the specimen source. In contrast, when dried blood spots, buccal scrapings, or plasma was used as the sample source, the sensitivity of DNA PCR was 80.95, 42.8, or 0%, respectively, compared to 100% sensitivity obtained with the original NASBA and Basic Kit NASBA assays. Our study indicates that the NucliSens Basic Kit NASBA assay is very sensitive and specific for CCR5 Delta32 genotyping using multiple sample types.

  7. Chemokine Receptor CCR5 Δ32 Genetic Analysis Using Multiple Specimen Types and the NucliSens Basic Kit†

    PubMed Central

    Tetali, Suryakumari; Lee, Eun Mi; Kaplan, Mark H.; Romano, Joseph W.; Ginocchio, Christine C.

    2001-01-01

    Resistance to HIV-1 infection and delayed disease progression have been associated with a 32-bp deletion (Δ32) in the gene encoding the CCR5 chemokine receptor. In the present study we describe the modification of a nucleic acid sequence-based amplification (NASBA)-based CCR5 genotyping assay for a NucliSens Basic Kit (Organon Teknika, Durham, N.C.) format using a new target-specific sandwich oligonucleotide detection methodology. The new method permitted the use of generic electrochemiluminescent probes supplied in the NucliSens Basic Kit, whereas the original NASBA method required expensive target-specific ruthenium detection probes. The Basic Kit CCR5 Δ32 genotypic analysis was in 100% concordance with both the original NASBA assay and DNA PCR results. This study also evaluated the use of multiple specimen types, including peripheral blood mononuclear cells (PBMC), whole blood, dried blood spots, buccal scrapings, and plasma, for CCR5 genotype analysis. The sensitivities of the three assays were comparable when PBMC or whole blood was the specimen source. In contrast, when dried blood spots, buccal scrapings, or plasma was used as the sample source, the sensitivity of DNA PCR was 80.95, 42.8, or 0%, respectively, compared to 100% sensitivity obtained with the original NASBA and Basic Kit NASBA assays. Our study indicates that the NucliSens Basic Kit NASBA assay is very sensitive and specific for CCR5 Δ32 genotyping using multiple sample types. PMID:11527812

  8. Review of the problems involved in using enzymes in blood group serology--provision of freeze-dried ICSH/ISBT protease enzyme and anti-D reference standards. International Council for Standardization in Haematology. International Society of Blood Transfusion.

    PubMed

    Scott, M L; Voak, D; Phillips, P K; Hoppe, P A; Kochman, S A

    1994-01-01

    Proteolytic enzyme preparations and techniques used routinely in blood group serology for the detection of atypical patient antibodies prior to transfusion vary widely and are often poorly standardised. Recent advances have been made in the use of biochemical methods to standardise and stabilise the potency of the enzyme preparations used. A joint working party of the International Council for Standardization in Haematology (ICSH) and the International Society of Blood Transfusion (ISBT) has investigated possibilities for the provision of standards for the protease preparations and techniques. The specification for these standards was that the performance of enzyme reference preparation in the reference technique should be of equivalent sensitivity to the ICSH/ISBT LISS spin indirect antiglobulin test using a titration series of a reference weak anti-D, and be free from false-positive reactions. The working party circulated materials for evaluation in inter-laboratory trials, followed by a laboratory workshop meeting to achieve agreement on the specification for reference materials and methods. Reference freeze-dried papain at 0.6 azoalbumin units and weak anti-D preparations (91/562) have been prepared and validated to meet these specifications. The performance of a test enzyme preparation in the technique for which it is recommended for use should be at least equal to that of the reference papain preparation, by the reference two-stage technique in terms of sensitivity, using a titration series of the reference anti-D, and freedom from false-positive reactions, using six fresh inert sera. The reference papain and weak anti-D can also be used to calibrate the level of proteolytic activity required in other procedures in blood group serology, such as new technology methods for antibody detection, and automated and microplate cell grouping procedures. These preparations and an agreed method for their use are now available from listed centres as ICSH/ISBT and Food

  9. Application of dried blood spots to determine vitamin D status in a large nutritional study with unsupervised sampling: the Food4Me project.

    PubMed

    Hoeller, Ulrich; Baur, Manuela; Roos, Franz F; Brennan, Lorraine; Daniel, Hannelore; Fallaize, Rosalind; Forster, Hannah; Gibney, Eileen R; Gibney, Mike; Godlewska, Magdalena; Hartwig, Kai; Kolossa, Silvia; Lambrinou, Christina P; Livingstone, Katherine M; Lovegrove, Julie A; Macready, Anna L; Manios, Yannis; Marsaux, Cyril F M; Martinez, J Alfredo; Celis-Morales, Carlos; Moschonis, George; Navas-Carretero, Santiago; O'Donovan, Clare B; San-Cristobal, Rodrigo; Saris, Wim H M; Surwiłło, Agnieszka; Traczyk, Iwona; Tsirigoti, Lydia; Walsh, Marianne C; Woolhead, Clara; Mathers, John C; Weber, Peter

    2016-01-28

    An efficient and robust method to measure vitamin D (25-hydroxy vitamin D3 (25(OH)D3) and 25-hydroxy vitamin D2 in dried blood spots (DBS) has been developed and applied in the pan-European multi-centre, internet-based, personalised nutrition intervention study Food4Me. The method includes calibration with blood containing endogenous 25(OH)D3, spotted as DBS and corrected for haematocrit content. The methodology was validated following international standards. The performance characteristics did not reach those of the current gold standard liquid chromatography-MS/MS in plasma for all parameters, but were found to be very suitable for status-level determination under field conditions. DBS sample quality was very high, and 3778 measurements of 25(OH)D3 were obtained from 1465 participants. The study centre and the season within the study centre were very good predictors of 25(OH)D3 levels (P<0·001 for each case). Seasonal effects were modelled by fitting a sine function with a minimum 25(OH)D3 level on 20 January and a maximum on 21 July. The seasonal amplitude varied from centre to centre. The largest difference between winter and summer levels was found in Germany and the smallest in Poland. The model was cross-validated to determine the consistency of the predictions and the performance of the DBS method. The Pearson's correlation between the measured values and the predicted values was r 0·65, and the sd of their differences was 21·2 nmol/l. This includes the analytical variation and the biological variation within subjects. Overall, DBS obtained by unsupervised sampling of the participants at home was a viable methodology for obtaining vitamin D status information in a large nutritional study.

  10. A quantitative method for acylcarnitines and amino acids using high resolution chromatography and tandem mass spectrometry in newborn screening dried blood spot analysis.

    PubMed

    Miller, John H; Poston, Philip A; Karnes, H Thomas

    2012-08-15

    We have developed a high resolution liquid chromatographic separation with electrospray ionization (ESI) mass spectrometry detection for the combined analysis of twelve acylcarnitines and seven amino acids commonly measured in newborn screening heritable metabolic disorders. Samples were prepared by punching 3.2 mm disks out of dried blood spots and extracting with a mixture of methanol and 0.1% formic acid containing stable isotopically labeled internal standards. Analysis was performed on an UHPLC system using a HILIC amide, 2.1 mm × 50 mm, 1.7 μm column. A normal phase gradient, employing 10mM ammonium acetate in 90:10 acetonitrile/water for mobile phase B and 0.1% formic acid in water for mobile phase A, was used. Optimized multiple reaction monitoring (MRM) was used for detection of amino acids and acylcarnitines on a Waters Premier mass spectrometer. Quantification of analytes was performed using internal calibration by fortification of sodium heparin whole blood with analytes at appropriate levels to encompass the range around the reported cut-off values. The method was fully validated with respect to precision, accuracy, recovery, linearity, matrix suppression and extraction robustness. Precision and accuracy were evaluated over 3 days and determined to be acceptable with an overall precision within 10% and accuracy within 15% of theoretical for all analytes except for acetylcarnitne at one fortified level, which quantitated 21.8% lower than the expected value. This method is suitable as a second-tier test for newborn screening of specific disorders associated with abnormal levels of acylcarnitines and amino acids, potentially reducing false positive cases and shortening the time to diagnosis.

  11. Validation and Application of a Dried Blood Spot Assay for Biofilm-Active Antibiotics Commonly Used for Treatment of Prosthetic Implant Infections

    PubMed Central

    Knippenberg, Ben; Page-Sharp, Madhu; Clark, Ben; Dyer, John; Batty, Kevin T.; Davis, Timothy M. E.

    2016-01-01

    Dried blood spot (DBS) antibiotic assays can facilitate pharmacokinetic (PK)/pharmacodynamic (PD) studies in situations where venous blood sampling is logistically difficult. We sought to develop, validate, and apply a DBS assay for rifampin (RIF), fusidic acid (FUS), and ciprofloxacin (CIP). These antibiotics are considered active against organisms in biofilms and are therefore commonly used for the treatment of infections associated with prosthetic implants. A liquid chromatography-mass spectroscopy DBS assay was developed and validated, including red cell partitioning and thermal stability for each drug and the rifampin metabolite desacetyl rifampin (Des-RIF). Plasma and DBS concentrations in 10 healthy adults were compared, and the concentration-time profiles were incorporated into population PK models. The limits of quantification for RIF, Des-RIF, CIP, and FUS in DBS were 15 μg/liter, 14 μg/liter, 25 μg/liter, and 153 μg/liter, respectively. Adjusting for hematocrit, red cell partitioning, and relative recovery, DBS-predicted plasma concentrations were comparable to measured plasma concentrations for each antibiotic (r > 0.95; P < 0.0001), and Bland-Altman plots showed no significant bias. The final population PK estimates of clearance, volume of distribution, and time above threshold MICs for measured and DBS-predicted plasma concentrations were comparable. These drugs were stable in DBSs for at least 10 days at room temperature and 1 month at 4°C. The present DBS antibiotic assays are robust and can be used as surrogates for plasma concentrations to provide valid PK and PK/PD data in a variety of clinical situations, including therapeutic drug monitoring or studies of implant infections. PMID:27270283

  12. A routine method for cholesterol and 7-dehydrocholesterol analysis in dried blood spot by GC-FID to diagnose the Smith-Lemli-Opitz syndrome.

    PubMed

    Gelzo, Monica; Clericuzio, Stefano; Barone, Rosalba; D'Apolito, Oceania; Dello Russo, Antonio; Corso, Gaetano

    2012-10-15

    This work was aimed to implement a fast and simple method to quantify cholesterol (CHOL) and 7-dehydrocholesterol (7-DHC) in dried blood spot (DBS) to diagnose the Smith-Lemli-Opitz syndrome (SLOS), an inborn error of CHOL biosynthesis. We developed and validated a GC-FID method for separation and quantification of underivatized CHOL and 7-DHC using a DBS disc of 6mm with a run time of 9 min. Correlation coefficients (r) of calibration curves ranged from 0.998 to 0.999 for CHOL and from 0.997 to 0.998 for 7-DHC. Within-day and between-day imprecision (CV%), accuracy (%), carry-over, and extraction efficacy (%) were also evaluated for validation. CHOL and 7-DHC were analyzed in DBS and plasma samples from 8 SLOS patients and 30 unaffected subjects. In SLOS patients, 7-DHC/CHOL ratios in DBS and plasma samples ranged from 0.035 to 1.448 and from 0.012 to 0.926, respectively. Results from calibration curves, quality controls and patient samples reveal that the method is suitable to analyze DBS to screen patients affected by SLOS.

  13. Determination of Biological Variance and Validation of a Fluorometric Assay for Measurement of α-l-Iduronidase Activity in Dried Blood Spots Samples: The First Experience in Iran.

    PubMed

    Abdi, Mohammad; Hakhamaneshi, Mohammad Said; Alaei, Mohammad Reza; Azadi, Namam-Ali; Vakili, Rahim; Zamanfar, Daniel; Taghikhani, Mohammad; Khatami, Shohreh

    2015-07-01

    Methods for assaying lysosomal diseases in dried blood samples are very useful today due to its several advantages related to the stability of samples, its transportation, handled and analysis, and its potential use for newborn screening compared to traditional methods in leucocytes samples. For this reason, it is important to validate these assays before being used in routine laboratory. Because of different in biological markers based on ethnicity, we aimed this study to validation a DBS-based fluorometric assay for measurement of α-l-Iduronidase activity for diagnosis of MPS I patients in Iran. DBS samples were collected from 15 MPS I patients and 60 healthy age matched subjects. Diagnostic value, biological variance and α-l-Iduronidase activity were determined. DBS α-l-Iduronidase activity was significantly higher in male subjects than in female group. Using a cut-off level of 1.08 µmol/spot 20 h, sensitivity and specificity were 100 and 98 %. The linearity of test was proved and we showed that within-run and between run precision were 5.6 and 14.66 %. Measurement of α-l-Iduronidase activity in DBS samples is an accurate test for diagnosis of MPS I and because of its rapid shipping and simplicity to keeping, DBS-based enzyme activity could be considered as a useful diagnostic tool in this disease.

  14. The use of real-time PCR to detect hepatitis C virus RNA in dried blood spots from Brazilian patients infected chronically.

    PubMed

    Santos, Carlos; Reis, Alexanda; Dos Santos, Cintia Vilhena; Damas, Cristine; Silva, Mariliza Henrique; Viana, Mônica Valverde; Ferraz, Maria Lucia; Carnauba, Dimas; El-Far, Fabiane; Serra, Fernando; Diaz, Ricardo Sobhie

    2012-01-01

    Collecting and transporting samples for RNA analysis can be challenging, especially in situations where financial resources are limited. In this study, a quantitative real-time PCR (qPCR) for the analysis of HCV RNA was developed and adapted for use with dried blood spot (DBS) samples. A qPCR for HCV 5'NCR, an internal control and a calibration curve were developed, and the sensitivity, specificity and dynamic range of amplification were evaluated using a panel of viruses. Plasma and DBS samples from 100 patients who had completed four weeks of Peginterferon alfa-2b+Ribavirin treatment were collected (DBS on SS903 collection cards and transported at room temperature). After 24 weeks of treatment, samples were collected from 68 of these patients. Of the 168 samples, 2 yielded false-negative results, and 4 yielded false-positive results (sensitivity was 98%, specificity was 94.3%, positive predictive value was 96.1%, and negative predictive value was 96.9%). Additionally, 2039 DBS samples from 1114 patients currently undergoing treatment for a chronic HCV infection in a clinical trial were tested. Only 10 samples out of the 2039 yielded invalid results warranting re-collection of DBS. The detection of HCV RNA in DBS can be a cost-effective strategy for HCV treatment monitoring, especially in settings where resources are limited.

  15. Validation of biomarkers of CVD risk from dried blood spots in community-based research: Methodologies and study-specific serum equivalencies

    PubMed Central

    Samuelsson, Laura B.; Hall, Martica H.; McLean, Shakir; Porter, James H.; Berkman, Lisa; Marino, Miguel; Sembajwe, Grace; McDade, Thomas W.; Buxton, Orfeu M.

    2016-01-01

    Objective Dried blood spot (DBS) methodology offers significant advantages over venipuncture in vulnerable populations or large-scale studies, including reduced participant burden and higher response rates. Uncertainty about validity of cardiovascular risk biomarkers remains a barrier to wide-scale use. We determined the validity of DBS-derived biomarkers of CVD risk versus gold-standard assessments, and study-specific, serum-equivalency values for clinical relevance of DBS-derived values. Methods Concurrent venipuncture serum and DBS samples (n=150 adults) were assayed in CLIA-certified and DBS laboratories, respectively. Time controls of DBS standard samples were assayed single-blind along with test samples. Linear regression analyses evaluated DBS-to-serum equivalency values; agreement and bias were assessed via Bland-Altman plots. Results Linear regressions of venipuncture values on DBS-to-serum equivalencies provided R2 values for TC, HDL-C, CRP of 0.484, 0.118, 0.666, respectively. Bland-Altman plots revealed minimal systematic bias between DBS-to-serum and venipuncture values; precision worsened at higher mean values of CRP. Time controls reveal little degradation or change in analyte values for HDL-C and CRP over 30 weeks. Conclusions DBS-assessed biomarkers represent a valid alternative to venipuncture assessments. Large studies using DBS should include study-specific serum-equivalency determinations to optimize individual-level sensitivity, viability of detecting intervention effects, and generalizability in community-level, primary prevention interventions. PMID:26652683

  16. Biaxial Creep Specimen Fabrication

    SciTech Connect

    JL Bump; RF Luther

    2006-02-09

    This report documents the results of the weld development and abbreviated weld qualification efforts performed by Pacific Northwest National Laboratory (PNNL) for refractory metal and superalloy biaxial creep specimens. Biaxial creep specimens were to be assembled, electron beam welded, laser-seal welded, and pressurized at PNNL for both in-pile (JOYO reactor, O-arai, Japan) and out-of-pile creep testing. The objective of this test campaign was to evaluate the creep behavior of primary cladding and structural alloys under consideration for the Prometheus space reactor. PNNL successfully developed electron beam weld parameters for six of these materials prior to the termination of the Naval Reactors program effort to deliver a space reactor for Project Prometheus. These materials were FS-85, ASTAR-811C, T-111, Alloy 617, Haynes 230, and Nirnonic PE16. Early termination of the NR space program precluded the development of laser welding parameters for post-pressurization seal weldments.

  17. Liquid chromatography/tandem mass spectrometry sensitivity enhancement via online sample dilution and trapping: applications in microdosing and dried blood spot (DBS) bioanalysis.

    PubMed

    Li, Fumin; Zulkoski, John P; Ding, Jie; Brown, Wes; Addison, Tom

    2010-09-15

    A simple online sample dilution, enrichment, and cleanup technique was developed for sensitive microdosing and dried blood spot (DBS) liquid chromatography/tandem mass spectrometric (LC/MS/MS) bioanalysis. Samples are diluted online with water and enriched in a trap column which is subsequently switched inline with the analytical column. Excellent lansoprazole (in acetonitrile) peak shape is maintained even with an 80-microL injection. In comparison, similar chromatographic peaks were observed only when a small volume of the same solution, i.e., 1 microL, was injected on a regular high-performance liquid chromatography (HPLC) system, where an injection of 5 microL resulted in severe peak fronting. A substantial enhancement in sensitivity is realized in the trapping mode using large injection volumes. The trap column is washed at the beginning and at the end of each injection with aqueous and organic solvent respectively to remove matrix components. This ultimately leads to reduction of matrix effects and mass spectrometer noise, thus facilitating the utilization of protein precipitation as the sample preparation for plasma samples. A lower limit of quantitation (LLOQ) of 0.5 pg/mL was demonstrated for lansoprazole in human plasma with a signal-to-noise (S/N) ratio of 13 using a 100 microL injection. Excellent intra-day precision and accuracy were established for lansoprazole in human plasma with good linearity (R(2) > 0.999) from 0.5 to 500 pg/mL. This level of LLOQ makes LC/MS/MS a practical alternative for microdosing bioanalysis, where the dose is typically 100 times lower than the therapeutic dose. The same technique was applied to quantitate lansoprazole in human whole blood employing DBS technology. With a single 3-mm punch, i.e. approximately 2 microL of whole blood or approximately 1 microL plasma, a LLOQ of 0.1 ng/mL showed sufficient S/N ratio (40) for lansoprazole when 75 microL of extract was injected. In all, the online sample dilution, cleanup, and

  18. Pharmacokinetic Study of Praziquantel Enantiomers and Its Main Metabolite R-trans-4-OH-PZQ in Plasma, Blood and Dried Blood Spots in Opisthorchis viverrini-Infected Patients

    PubMed Central

    Meister, Isabel; Kovac, Jana; Duthaler, Urs; Odermatt, Peter; Huwyler, Jörg; Vanobberghen, Fiona; Sayasone, Somphou; Keiser, Jennifer

    2016-01-01

    Background Praziquantel (PZQ) is the treatment of choice for infections with the liver fluke Opisthorchis viverrini, a major health problem in Southeast Asia. However, pharmacokinetic (PK) studies investigating the disposition of PZQ enantiomers (R- and S-PZQ) and its main metabolite, R-trans-4-OH-PZQ, in diseased patients are lacking. The implementation of a dried blood spot (DBS) sampling technique would ease the performance of PK studies in remote areas without clinical facilities. The aim of the present study is to provide data on the disposition of PZQ enantiomers and R-trans-4-OH-PZQ in opisthorchiasis patients and to validate the use of DBS compared to plasma and blood sampling. Methodology/Principal Findings PZQ was administered to nine O. viverrini-infected patients at 3 oral doses of 25 mg/kg in 4 h intervals. Plasma, blood and DBS were simultaneously collected at selected time points from 0 to 24 h post-treatment. PK parameters were determined using non-compartmental analysis. Drug concentrations and areas under the curve (AUC0–24h) measured in the 3 matrices were compared using Bland-Altman analysis. We observed plasma AUC0–24hs of 1.1, 9.0 and 188.7 μg/ml*h and half-lives of 1.1, 3.3 and 6.4 h for R-PZQ, S-PZQ and R-trans-4-OH, respectively. Maximal plasma concentrations (Cmax) of 0.2, 0.9 and 13.9 μg/ml for R-PZQ, S-PQZ and R-trans-4-OH peaked at 7 h for PZQ enantiomers and at 8.7 h for the metabolite. Individual drug concentration measurements and patient AUC0–24hs displayed ratios of blood or DBS versus plasma between 79–94% for R- and S-PZQ, and between 108–122% for R-trans-4-OH. Conclusions/Significance Pharmacodynamic (PD) in vitro studies on PZQ enantiomers and R-trans-4-OH-PZQ are necessary to be able to correlate PK parameters with efficacy. DBS appears to be a valid alternative to conventional venous sampling for PK studies in PZQ-treated patients. PMID:27152952

  19. Method for thinning specimen

    SciTech Connect

    Follstaedt, David M.; Moran, Michael P.

    2005-03-15

    A method for thinning (such as in grinding and polishing) a material surface using an instrument means for moving an article with a discontinuous surface with an abrasive material dispersed between the material surface and the discontinuous surface where the discontinuous surface of the moving article provides an efficient means for maintaining contact of the abrasive with the material surface. When used to dimple specimens for microscopy analysis, a wheel with a surface that has been modified to produce a uniform or random discontinuous surface significantly improves the speed of the dimpling process without loss of quality of finish.

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

    PubMed

    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

  1. 46 CFR 4.06-20 - Specimen collection requirements.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... involved in the SMI must provide a specimen of their breath, blood, or saliva to the marine employer as... only by qualified medical personnel. (3) Collection of an individual's saliva or breath to comply...

  2. 46 CFR 4.06-20 - Specimen collection requirements.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... involved in the SMI must provide a specimen of their breath, blood, or saliva to the marine employer as... only by qualified medical personnel. (3) Collection of an individual's saliva or breath to comply...

  3. Development and Evaluation of an Affordable Real-Time Qualitative Assay for Determining HIV-1 Virological Failure in Plasma and Dried Blood Spots

    PubMed Central

    Kliphuis, Aletta; Bronze, Michelle; Wallis, Carole L.; Kityo, Cissy; Balinda, Sheila; Stevens, Wendy; Spieker, Nicole; de Oliveira, Tulio; Rinke de Wit, Tobias F.; Schuurman, Rob

    2013-01-01

    Virological failure (VF) has been identified as the earliest, most predictive determinant of HIV-1 antiretroviral treatment (ART) failure. Due to the high cost and complexity of virological monitoring, VF assays are rarely performed in resource-limited settings (RLS). Rather, ART failure is determined by clinical monitoring and to a large extent immunological monitoring. This paper describes the development and evaluation of a low-cost, dried blood spot (DBS)-compatible qualitative assay to determine VF, in accordance with current WHO guideline recommendations for therapy switching in RLS. The assay described here is an internally controlled qualitative real-time PCR targeting the conserved long terminal repeat domain of HIV-1. This assay was applied to HIV-1 subtypes A to H and further evaluated on HIV-1 clinical plasma samples from South Africa (n = 191) and Tanzania (n = 42). Field evaluation was performed in Uganda using local clinical plasma samples (n = 176). Furthermore, assay performance was evaluated for DBS. This assay is able to identify VF for all major HIV-1 group M subtypes with equal specificity and has a lower detection limit of 1.00E+03 copies/ml for plasma samples and 5.00E+03 copies/ml for DBS. Comparative testing yielded accurate VF determination for therapy switching in 89% to 96% of samples compared to gold standards. The assay is robust and flexible, allowing for “open platform” applications and producing results comparable to those of commercial assays. Assay design enables application in laboratories that can accommodate real-time PCR equipment, allowing decentralization of testing to some extent. Compatibility with DBS extends access of sampling and thus access to this test to remote settings. PMID:23596235

  4. Higher specificity of nucleic acid sequence-based amplification isothermal technology than of real-time PCR for quantification of HIV-1 RNA on dried blood spots.

    PubMed

    Mercier-Delarue, Severine; Vray, Muriel; Plantier, Jean Christophe; Maillard, Theodora; Adjout, Zidan; de Olivera, Fabienne; Schnepf, Nathalie; Maylin, Sarah; Simon, Francois; Delaugerre, Constance

    2014-01-01

    Dried blood spots (DBS) are widely proposed as a plasma surrogate for monitoring antiretroviral treatment efficacy based on the HIV-1 RNA level (viral load [VL]) in resource-limited settings. Interfering coamplification of cell-associated HIV-1 DNA during reverse transcription (RT)-PCR can be avoided by using nucleic acid sequence-based amplification (NASBA) technology, which is based on an RNA template and isothermic conditions. We analyzed VL values obtained with DBS and plasma samples by comparing isothermic NASBA (NucliSENS EasyQ HIV-1 V2.0; bioMérieux) with real-time RT-PCR (Cobas TaqMan HIV-1 V2.0; Roche). Samples from 197 HIV-1-infected patients were tested (non-B subtypes in 51% of the cases). Nucleic acid extractions were performed by use of NucliSENS EasyMAG (bioMérieux) and Cobas AmpliPrep (Roche) before the NASBA and RT-PCR quantifications, respectively. Both quantification assays have lower limits of detection of 20 (1.3) and 800 (2.9) log10 copies/ml (log) in plasma and DBS, respectively. The mean (DBS minus plasma) differences were -0.39 and -0.46 log, respectively, for RT-PCR and NASBA. RT-PCR on DBS identified virological failure in 122 of 126 patients (sensitivity, 97%) and viral suppression in 58 of 70 patients (specificity, 83%), yielding 12 false-positive results (median, 3.2 log). NASBA on DBS identified virological failure in 85 of 96 patients (sensitivity, 89%) and viral suppression in 95 of 97 patients (specificity, 98%) and yielded 2 false-positive results (3.0 log for both). Both technologies detected HIV-1 RNA in DBS at a threshold of 800 copies/ml. This higher specificity of NASBA technology could avoid overestimation of poor compliance or the emergence of resistance when monitoring antiretroviral efficacy with the DBS method.

  5. Development and evaluation of an affordable real-time qualitative assay for determining HIV-1 virological failure in plasma and dried blood spots.

    PubMed

    Aitken, Susan C; Kliphuis, Aletta; Bronze, Michelle; Wallis, Carole L; Kityo, Cissy; Balinda, Sheila; Stevens, Wendy; Spieker, Nicole; de Oliveira, Tulio; Rinke de Wit, Tobias F; Schuurman, Rob

    2013-06-01

    Virological failure (VF) has been identified as the earliest, most predictive determinant of HIV-1 antiretroviral treatment (ART) failure. Due to the high cost and complexity of virological monitoring, VF assays are rarely performed in resource-limited settings (RLS). Rather, ART failure is determined by clinical monitoring and to a large extent immunological monitoring. This paper describes the development and evaluation of a low-cost, dried blood spot (DBS)-compatible qualitative assay to determine VF, in accordance with current WHO guideline recommendations for therapy switching in RLS. The assay described here is an internally controlled qualitative real-time PCR targeting the conserved long terminal repeat domain of HIV-1. This assay was applied to HIV-1 subtypes A to H and further evaluated on HIV-1 clinical plasma samples from South Africa (n = 191) and Tanzania (n = 42). Field evaluation was performed in Uganda using local clinical plasma samples (n = 176). Furthermore, assay performance was evaluated for DBS. This assay is able to identify VF for all major HIV-1 group M subtypes with equal specificity and has a lower detection limit of 1.00E+03 copies/ml for plasma samples and 5.00E+03 copies/ml for DBS. Comparative testing yielded accurate VF determination for therapy switching in 89% to 96% of samples compared to gold standards. The assay is robust and flexible, allowing for "open platform" applications and producing results comparable to those of commercial assays. Assay design enables application in laboratories that can accommodate real-time PCR equipment, allowing decentralization of testing to some extent. Compatibility with DBS extends access of sampling and thus access to this test to remote settings.

  6. Evaluation of Mutual Drug-Drug Interaction within Geneva Cocktail for Cytochrome P450 Phenotyping using Innovative Dried Blood Sampling Method.

    PubMed

    Bosilkovska, Marija; Samer, Caroline; Déglon, Julien; Thomas, Aurélien; Walder, Bernhard; Desmeules, Jules; Daali, Youssef

    2016-09-01

    Cytochrome P450 (CYP) activity can be assessed using a 'cocktail' phenotyping approach. Recently, we have developed a cocktail (Geneva cocktail) which combines the use of low-dose probes with a low-invasiveness dried blood spots (DBS) sampling technique and a single analytical method for the phenotyping of six major CYP isoforms. We have previously demonstrated that modulation of CYP activity after pre-treatment with CYP inhibitors/inducer could be reliably predicted using Geneva cocktail. To further validate this cocktail, in this study, we have verified whether probe drugs contained in the latter cause mutual drug-drug interactions. In a randomized, four-way, Latin-square crossover study, 30 healthy volunteers received low-dose caffeine, flurbiprofen, omeprazole, dextromethorphan and midazolam (a previously validated combination with no mutual drug-drug interactions); fexofenadine alone; bupropion alone; or all seven drugs simultaneously (Geneva cocktail). Pharmacokinetic profiles of the probe drugs and their metabolites were determined in DBS samples using both conventional micropipette sampling and new microfluidic device allowing for self-sampling. The 90% confidence intervals for the geometric mean ratios of AUC metabolite/AUC probe for CYP probes administered alone or within Geneva cocktail fell within the 0.8-1.25 bioequivalence range indicating the absence of pharmacokinetic interaction. The same result was observed for the chosen phenotyping indices, that is metabolic ratios at 2 hr (CYP1A2, CYP3A) or 3 hr (CYP2B6, CYP2C9, CYP2C19, CYP2D6) post-cocktail administration. DBS sampling could successfully be performed using a new microfluidic device. In conclusion, Geneva cocktail combined with an innovative DBS sampling device can be used routinely as a test for simultaneous CYP phenotyping.

  7. Microwave-assisted on-spot derivatization for gas chromatography-mass spectrometry based determination of polar low molecular weight compounds in dried blood spots.

    PubMed

    Sadones, Nele; Van Bever, Elien; Archer, John R H; Wood, David M; Dargan, Paul I; Van Bortel, Luc; Lambert, Willy E; Stove, Christophe P

    2016-09-23

    Dried blood spot (DBS) sampling and analysis is increasingly being applied in bioanalysis. Although the use of DBS has many advantages, it is also associated with some challenges. E.g. given the limited amount of available material, highly sensitive detection techniques are often required to attain sufficient sensitivity. In gas chromatography coupled to mass spectrometry (GC-MS), derivatization can be helpful to achieve adequate sensitivity. Because this additional sample preparation step is considered as time-consuming, we introduce a new derivatization procedure, i.e. "microwave-assisted on-spot derivatization", to minimize sample preparation of DBS. In this approach the derivatization reagents are directly applied onto the DBS and derivatization takes place in a microwave instead of via conventional heating. In this manuscript we evaluated the applicability of this new concept of derivatization for the determination of two polar low molecular weight molecules, gamma-hydroxybutyric acid (GHB) and gabapentin, in DBS using a standard GC-MS configuration. The method was successfully validated for both compounds, with imprecision and bias values within acceptance criteria (<20% at LLOQ, <15% at 3 other QC levels). Calibration lines were linear over the 10-100μg/mL and 1-30μg/mL range for GHB and gabapentin, respectively. Stability studies revealed no significant decrease of gabapentin and GHB in DBS upon storage at room temperature for at least 84 days. Furthermore, DBS-specific parameters, including hematocrit and volume spotted, were evaluated. As demonstrated by the analysis of GHB and gabapentin positive samples, "microwave-assisted on-spot derivatization" proved to be reliable, fast and applicable in routine toxicology. Moreover, other polar low molecular weight compounds of interest in clinical and/or forensic toxicology, including vigabatrin, beta-hydroxybutyric acid, propylene glycol, diethylene glycol, 1,4-butanediol and 1,2-butanediol, can also be

  8. Effects of feeding propylene glycol on dry matter intake, lactation performance, energy balance and blood metabolites in early lactation dairy cows.

    PubMed

    Liu, Q; Wang, C; Yang, W Z; Zhang, W W; Yang, X M; He, D C; Dong, K H; Huang, Y X

    2009-10-01

    The objectives of this study were to evaluate effects of feeding propylene glycol (PG) on feed intake, milk yield and milk composition, blood metabolites and energy balance in Holstein dairy cows from 1 to 63 days in milk. Thirty-two multiparous cows, blocked by lactation number, previous 305-day milk production and expected calving date, were arranged into four groups in a randomized block design. Treatments were: control, low PG, medium PG and high PG with 0, 150, 300 and 450 ml PG per cow per day, respectively. The supplement of food grade PG (0.998 g/g PG) was hand-mixed into the top one-third of the daily ration. Cows were fed ad libitum a total mixed ration consisting of forage and concentrate (50 : 50, dry matter basis). Feed intake, milk yield and milk components were not affected (P > 0.05) by PG supplementation. Overall, body weight (BW) loss tended (P < 0.08) to be linearly reduced, and energy status was linearly improved with increasing PG supplementation. Concentrations of glucose in plasma were higher for cows fed PG relative to control (55.6 v. 58.9 mg/dl) and linearly increased (P < 0.01) with increasing PG supplementation. Plasma concentrations of non-esterified fatty acids and beta-hydroxybutyrate were linearly increased, but urine acetoacetate concentration was quadratically changed with the highest for control diet and the lowest for 450 ml/day of PG. These results indicated that supplementation of PG in the early lactating cow diets had minimal effects on feed intake and milk production, but may potentially reduce contents of milk fat and milk protein. Supplementation of early lactating dairy cow diets with PG is beneficial in terms of improving energy status and reducing BW loss.

  9. Targeted case finding for hepatitis B using dry blood spot testing in the British-Chinese and South Asian populations of the North-East of England.

    PubMed

    McPherson, S; Valappil, M; Moses, S E; Eltringham, G; Miller, C; Baxter, K; Chan, A; Shafiq, K; Saeed, A; Qureshi, R; Hudson, M; Bassendine, M F

    2013-09-01

    Chronic infection with the hepatitis B virus (HBV) is a frequent cause of cirrhosis and liver cancer. Targeted HBV screening is recommended by the Centre for Disease Control (CDC) and Prevention for subjects born in countries with >2% HBV prevalence. However, there are no UK guidelines. Here, we applied the (CDC) recommendations to the British-Chinese and British-South Asian community of North-East (NE) England. British-Chinese and South Asian subjects were invited to attend for HBV education and screening sessions held in community centres. Hepatitis B surface antigen (HBsAg) and hepatitis B core total antibody (HBcAb) were tested with dry blood spot tests. South Asians were also tested for hepatitis C antibody (HCVAb). A total of 1126 subjects (606 Chinese and 520 South Asian) were screened. Sixty-two (5.5%) were HBsAg positive. Ten of these reported a previous diagnosis of HBV. The prevalence of HBsAg positivity was 4.6% when previously diagnosed individuals were excluded. The HBsAg prevalence was significantly higher in the Chinese subjects compared with South Asians (8.7% VS. 1.7% P < 0.001). In Chinese subjects, HBsAg positivity was highest in subjects born in Vietnam (17.4%), followed by China (11%), Hong Kong (7.8%) and the UK (6.7%). Subjects from Pakistan had the highest HBsAg and HCV Ab prevalence in the South Asians (3.1% and 1.8%, respectively). Ten percentage of HBsAg positive patients who had follow-up assessment had active disease requiring antiviral treatment. Undiagnosed HBV infection was above the 2% threshold for screening suggested by the CDC in the British-Chinese and Pakistani community of NE England, which provides evidence for a UK HBV-targeted screening programme.

  10. Use of dried-blood-spot samples and in-house assays to identify antiretroviral drug resistance in HIV-infected children in resource-constrained settings.

    PubMed

    Ziemniak, Carrie; Mengistu, Yohannes; Ruff, Andrea; Chen, Ya-Hui; Khaki, Leila; Bedri, Abubaker; Simen, Birgitte B; Palumbo, Paul; Eshleman, Susan H; Persaud, Deborah

    2011-12-01

    Monitoring HIV drug resistance is an important component of the World Health Organization's global HIV program. HIV drug resistance testing is optimal with commercially available clinically validated test kits using plasma; however, that type of testing may not be feasible or affordable in resource-constrained settings. HIV genotyping from dried blood spots (DBS) with noncommercial (in-house) assays may facilitate the capture of HIV drug resistance outcomes in resource-constrained settings but has had varying rates of success. With in-house assays for HIV reverse transcriptase, we evaluated the yield of genotyping DBS samples collected from HIV-infected children who were enrolled in two clinical trials conducted in sub-Saharan Africa (median HIV viral load, 5.88 log(10) HIV RNA copies/ml; range, 4.04 to 6.99). Overall, HIV genotypes were obtained for 94 (89.5%) of 105 samples tested (95% and 84% from clinical trials #1 and #2, respectively); however, successful analysis of 15 (16.1%) of the 94 samples required repeat testing using a different set of primers on previously synthesized cDNA. The yield of genotyping was lower on the DBS that were stored suboptimally from clinical trial #2 (56% versus 88% for optimally stored). Concordance with plasma genotypes derived using a clinically validated, commercial kit-based assay (ViroSeq HIV-1 genotyping system) was also assessed in a subset of children with paired testing. For 34 samples with paired DBS and plasma genotypes, there was 100% concordance for major drug resistance mutations. DBS genotyping using in-house assays provides an alternative for antiretroviral drug resistance testing in children in resource-constrained regions but may require region-specific optimization before widespread use.

  11. Therapeutic drug monitoring of carbamazepine and its metabolite in children from dried blood spots using liquid chromatography and tandem mass spectrometry.

    PubMed

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

    2015-05-10

    Carbamazepine (CBZ) is a first-line drug for the treatment of different forms of epilepsy and the first choice drug for trigeminal neuralgia. CBZ is metabolized in the liver by oxidation into carbamazepine-10,11-epoxide (CBZE), its major metabolite which is equipotent and known to contribute to the pharmacological activity of CBZ. The aim of the present study was to develop and validate a reliable, selective and sensitive liquid chromatography-tandem mass spectrometry method for the simultaneous quantification of CBZ and its active metabolite in dried blood spots (DBS). The extraction process was carried out from DBS using methanol-water-formic acid (80:20:0.1, v/v/v). Chromatographic elution was achieved by using a linear gradient with a mobile phase consisting of acetonitrile-water-0.1% formic acid at a flow rate of 0.50mL/min. The method was linear over the range 1-40mg/L and 0.25-20mg/L for CBZ and CBZE, respectively. The limit of quantification was 0.75mg/L and 0.25mg/L for CBZ and CBZE. Intra-day and inter-day assay precisions were found to be lower than 5.13%, 6.46% and 11.76%, 4.72% with mean percentage accuracies of 102.1%, 97.5% and 99.2%, 97.8% for CBZ and CBZE. We successfully applied the method for determining DBS finger-prick samples in paediatric patients and confirmed the results with concentrations measured in matched plasma samples. This novel approach allows quantification of CBZ and its metabolite from only one 3.2mm DBS disc by LC-MS/MS thus combining advantages of DBS technique and LC-MS/MS in clinical practice.

  12. Incident Infection and Resistance Mutation Analysis of Dried Blood Spots Collected in a Field Study of HIV Risk Groups, 2007-2010

    PubMed Central

    Wei, Xierong; Smith, Amanda J.; Forrest, David W.; Cardenas, Gabriel A.; Beck, Dano W.; LaLota, Marlene; Metsch, Lisa R.; Sionean, Catlainn; Owen, S. Michele; Johnson, Jeffrey A.

    2016-01-01

    Objective To assess the utility of cost-effective dried blood spot (DBS) field sampling for incidence and drug resistance surveillance of persons at high risk for HIV infection. Methods We evaluated DBS collected in 2007–2010 in non-clinical settings by finger-stick from HIV-positive heterosexuals at increased risk of HIV infection (n = 124), men who have sex with men (MSM, n = 110), and persons who inject drugs (PWID, n = 58). Relative proportions of recent-infection findings among risk groups were assessed at avidity index (AI) cutoffs of ≤25%, ≤30%, and ≤35%, corresponding to an infection mean duration of recency (MDR) of 220.6, 250.4, and 278.3 days, respectively. Drug resistance mutation prevalence was compared among the risk groups and avidity indices. Results HIV antibody avidity testing of all self-reported ARV-naïve persons (n = 186) resulted in 9.7%, 11.3% and 14.0% with findings within the 221, 250, and 278-day MDRs, respectively. The proportion of ARV-naïve MSM, heterosexuals, and PWID reporting only one risk category who had findings below the suggested 30% AI was 23.1%, 6.9% and 3.6% (p<0.001), respectively. MSM had the highest prevalence of drug resistance and the only cases of transmitted multi-class resistance. Among the ARV-experienced, MSM had disproportionately more recent-infection results than did heterosexuals and PWID. Conclusions The disproportionately higher recent-infection findings for MSM as compared to PWID and heterosexuals increased as the MDR window increased. Unreported ARV use might explain greater recent-infection findings and drug resistance in this MSM population. DBS demonstrated utility in expanded HIV testing; however, optimal field handling is key to accurate recent-infection estimates. PMID:27415433

  13. Newborn screening for galactosemia by a second-tier multiplex enzyme assay using UPLC-MS/MS in dried blood spots.

    PubMed

    Ko, Dae-Hyun; Jun, Sun-Hee; Park, Kyoung Un; Song, Sang Hoon; Kim, Jin Q; Song, Junghan

    2011-04-01

    Galactosemia is one of the most important inherited metabolic disorders detected by newborn screening tests. Abnormal results during screening should be confirmed by enzyme activity assays. Recently, we developed a multiplex enzyme assay for galactosemia in erythrocytes using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). In this study, we proposed a second-tier multiplex enzyme assay for galactosemia that can be directly applied to dried blood spots (DBSs). Supernatants from two rehydrated-punched 3.2-mm DBSs were incubated with a reaction mixture containing [¹³C6]galactose, [¹³C2]galactose-1-phosphate, and UDP-glucose as substrates for three galactose-metabolizing enzymes. After a 4-hour incubation, the end products from the combined reaction mixture, [¹³C6]galactose-1-phosphate, UDP-[¹³C2]galactose, and UDP-galactose, were simultaneously measured using UPLC-MS/MS. Substrates, products, and internal standards from the mixture of the three enzyme reactions were clearly separated in the UPLC-MS/MS system, with an injection cycle time of 10 min. Intra- and inter-assay imprecisions of the UPLC-MS/MS were 8.4-14.8% and 13.2-15.7% CV, respectively. Enzyme activities in DBSs from 37 normal individuals and 10 patients with enzyme deficiencies were analyzed. DBSs from galactosemia patients showed consistently lower enzyme activities as compared to those of normal individuals. In conclusion, multiplex enzyme assays using UPLC-MS/MS can be successfully applied to DBS analysis. This method allows a fast and effective second-tier test for newborns showing abnormal screening results.

  14. Short Communication: Effect of heat stress during the dry period on gene expression in mammary tissue and peripheral blood mononuclear cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Heat stress (HT) during the dry period compromises mammary gland development, decreases future milk production, and impairs immune status of dairy cows. Our objective was to evaluate the effect of cooling HT cows during the dry period on gene expression of the mammary gland and lymphocytes. Cows wer...

  15. Radioimmunoassay of ''free thyroxin'' in dried blood spots on filter paper - preliminary observations on the effective differentiation of subjects with congenital hypothyroidism from those with subnormal thyroxin-binding globulin and normal subjects

    SciTech Connect

    Mizuta, H.; Miyai, K.; Ichihara, K.; Amino, N.; Harada, T.; Nose, O.; Tanizawa, O.

    1982-03-01

    In this sensitive, simple method for measuring ''free thyroxin'' (FT/sub 4/) in eluates of dried blood spots on filter paper by use of a radioimmunoassay kit (Amerlex Free T/sub 4/ RIA), the measurable range of FT/sub 4/ is 1.8 to 57 ng/L (equivalent to the concentration in serum), or 7 to 237 fg/tube. The mean coefficients of variation for within assay-within spots, within assay-between spots, and between assays were 5.3%, 5.0%, and 6.2%, respectively. FT/sub 4/ in blood spotted on filter paper is stable for at least a month when dried and kept at either -20/sup 0/C, 4/sup 0/C, room temperature (about 25/sup 0/C), or 37/sup 0/C. The results for FT/sub 4/ in dried blood spots correlated closely with the free-T/sub 4/ concentration in serum (r = 0.99). The method can be used to differentiate cases of primary and secondary hypothyroidism from normal subjects and those with subnormal thyroxin-binding globulin. This method may be useful in screening for congenital hypothyroidism, because sample-retesting is not necessary.

  16. 49 CFR Appendix C to Part 219 - Post-Accident Testing Specimen Collection

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    .... Collection of blood and urine specimens is required to be conducted at an independent medical facility... and transfer of blood and urine specimens for three surviving employees can be found in the FRA post.... Each shipping box contains supplies for blood/urine collections from three individuals,...

  17. 49 CFR Appendix C to Part 219 - Post-Accident Testing Specimen Collection

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    .... Collection of blood and urine specimens is required to be conducted at an independent medical facility... and transfer of blood and urine specimens for three surviving employees can be found in the FRA post.... Each shipping box contains supplies for blood/urine collections from three individuals,...

  18. 49 CFR Appendix C to Part 219 - Post-Accident Testing Specimen Collection

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    .... Collection of blood and urine specimens is required to be conducted at an independent medical facility... and transfer of blood and urine specimens for three surviving employees can be found in the FRA post.... Each shipping box contains supplies for blood/urine collections from three individuals,...

  19. 49 CFR Appendix C to Part 219 - Post-Accident Testing Specimen Collection

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    .... Collection of blood and urine specimens is required to be conducted at an independent medical facility... and transfer of blood and urine specimens for three surviving employees can be found in the FRA post.... Each shipping box contains supplies for blood/urine collections from three individuals,...

  20. 49 CFR Appendix C to Part 219 - Post-Accident Testing Specimen Collection

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    .... Collection of blood and urine specimens is required to be conducted at an independent medical facility... and transfer of blood and urine specimens for three surviving employees can be found in the FRA post.... Each shipping box contains supplies for blood/urine collections from three individuals,...

  1. The effects of a 6-month resistance training and dried plum consumption intervention on strength, body composition, blood markers of bone turnover, and inflammation in breast cancer survivors.

    PubMed

    Simonavice, Emily; Liu, Pei-Yang; Ilich, Jasminka Z; Kim, Jeong-Su; Arjmandi, Bahram; Panton, Lynn B

    2014-06-01

    The purpose of this study was to examine the effects of resistance training (RT) and dried plum (DP) consumption on strength, body composition, blood markers of bone, and inflammation in breast cancer survivors (BCS). Twenty-three BCS (RT, n = 12; RT+DP, n = 11), aged 64 ± 7 years, were evaluated at baseline and after 6 months of intervention on the following: muscular strength (chest press and leg extension) via 1-repetition maximums (1RMs); body composition, specifically bone mineral density (BMD) by dual energy X-ray absorptiometry; biochemical markers of bone turnover (bone-specific alkaline phosphatase (BAP), tartrate resistant acid phosphatase (TRAP-5b)); and inflammation (C-reactive protein (CRP)). Target RT prescription was 2 days/week of 10 exercises, including 2 sets of 8-12 repetitions at ∼60%-80% of 1RM. RT+DP also consumed 90 g of DP daily. There were no baseline differences between groups or any group-by-time interactions for any of the variables. BCS increased upper (p < 0.05) (RT: 64 ± 14 to 80 ± 17 kg; RT+DP: 72 ± 23 to 91 ± 20 kg) and lower (p < 0.05) (RT: 69 ± 20 to 87 ± 28 kg; RT+DP: 78 ± 19 to 100 ± 21 kg) body strength. Body composition and BMD improvements were not observed. TRAP-5b decreased in the RT group (p < 0.05) (4.55 ± 1.57 to 4.04 ± 1.63 U/L) and the RT+DP group (p = 0.07) (5.10 ± 2.75 to 4.27 ± 2.03 U/L). Changes in BAP and CRP were not observed. RT was effective for improving biochemical markers of bone turnover and muscular strength in BCS. A longer and higher intensity intervention may be needed to reveal the true effects of RT and DP on body composition and biochemical markers of inflammation.

  2. Reference intervals for orotic acid in urine, plasma and dried blood spot using hydrophilic interaction liquid chromatography-tandem mass spectrometry.

    PubMed

    D'Apolito, Oceania; Garofalo, Daniela; la Marca, Giancarlo; Dello Russo, Antonio; Corso, Gaetano

    2012-02-01

    Orotic acid (OA), a marker of hereditary orotic aciduria, is usually used for the differential diagnosis of some hyperammonemic inherited defects of urea cycle and of basic amino acid transporters. This study was aimed to establish age related reference intervals of OA in urine, and for the first time in plasma, and dried blood spot (DBS) from 229 apparently healthy subjects aged from three days to 40 years. The quantification of OA was performed by a previously implemented method, using a stable isotope dilution with 1,3-[(15)N(2)]-orotic acid and hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS). The method has proved to be sensitive and accurate for a quantitative analysis of OA also in DBS and plasma. According to previous studies, urinary OA levels (mmol/mol of creatinine) decrease significantly with age. The upper limits (as 99th %ile) were of 3.44 and 1.30 in groups aged from three days to 1 year (group 1) and from 1 year to 12 years (group 2), respectively; in teenagers (from 13 to 19 years; group 3) and adults (from 20 to 40 years; group 4) urinary levels became more stable and the upper limits were of 0.64 and 1.21, respectively. Furthermore, OA levels in DBS (μM) also resulted significantly higher in subjects of group 1 (upper limit of 0.89) than in subjects of groups 2, 3 and 4 (upper limits of 0.24, 0.21, and 0.29, respectively). OA levels in plasma (μM) were significantly lower in subjects of group 3 (upper limit of 0.30) than in subjects of groups 1, 2, and 4 (upper limits of 0.59, 0.48, and 0.77, respectively). This method was also employed for OA quantification in plasma and DBS of 17 newborns affected by urea cycle defects, resulting sensitive and specific enough to screen these disorders.

  3. Sensitive determination of prohibited drugs in dried blood spots (DBS) for doping controls by means of a benchtop quadrupole/Orbitrap mass spectrometer.

    PubMed

    Thomas, Andreas; Geyer, Hans; Schänzer, Wilhelm; Crone, Catharina; Kellmann, Markus; Moehring, Thomas; Thevis, Mario

    2012-05-01

    In the present study, a new type of mass spectrometer combining a quadrupole mass filter, a higher collision dissociation (HCD) cell and an Orbitrap detector, was evaluated for the analysis of dried blood spots (DBS) in doping controls. DBS analysis is characterized by the necessity to detect prohibited compounds in sub-nanogram-per-milliliter levels with high identification capacity. After extraction of DBS with an organic solvent and liquid chromatographic separation (using a regular C18-RP-analytical UHPLC-column) of target analytes, mass spectrometry is performed with a high-resolution full scan in positive and negative mode by means of electrospray ionisation. Single-product ion mass spectra are acquired using the data-dependent analysis mode (employing an inclusion list) for previously selected precursors of known prohibited compounds with fixed retention time ranges. Besides, a sensitive screening in a targeted approach, non-targeted analysis for retrospective data evaluation is thus possible. The chosen experimental design enables the determination of various drugs from different classes with one generic sample preparation which is shown for 26 selected model compounds (Δ(9)-tetrahydrocannabinol (THC), tetrahydrocannabinol-9-carboxylic acid (THC-COOH), methylhexaneamine, methylphenidate, cocaine, nikethamide, 3,4-methylenedioxyamphetamine, N-methyl-3,4-methylenedioxyamphetamine, strychnine, mesocarb, salbutamol, formoterol, clenbuterol, metandienone, stanozolol, bisoprolol, propranolol, metoprolol, anastrazole, clomiphene, exemestane, dexamethasone, budesonide, selective androgen receptor modulator (SARM) S4 (andarine), SARM S1, hydrochlorothiazide). Generally, only qualitative result interpretation was focussed upon, but for target analytes with deuterium-labelled internal standards (salbutamol, clenbuterol, cocaine, dexamethasone, THC-COOH and THC) quantitative analysis was also possible. Especially the most challenging analytes, THC and its carboxy

  4. Dry Mouth

    MedlinePlus

    ... or chewing tobacco can increase dry mouth symptoms. Methamphetamine use. Methamphetamine use can cause severe dry mouth and damage to teeth, a condition also known as "meth mouth." If you don't have enough saliva ...

  5. Dry Mouth

    MedlinePlus

    Dry mouth is the feeling that there is not enough saliva in your mouth. Everyone has a dry mouth once in a while - if they are nervous, ... under stress. But if you have a dry mouth all or most of the time, it can ...

  6. Dry Mouth? Don't Delay Treatment

    MedlinePlus

    ... Blood & Biologics Animal & Veterinary Cosmetics Tobacco Products For Consumers Home For Consumers Consumer Updates Dry Mouth? Don' ... or neck cancer. back to top Advice for Consumers If you have persistent dry mouth: Talk to ...

  7. 37 CFR 2.59 - Filing substitute specimen(s).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2014-07-01 2014-07-01 false Filing substitute specimen(s). 2.59 Section 2.59 Patents, Trademarks, and Copyrights UNITED STATES PATENT AND TRADEMARK OFFICE... mark as used on or in connection with the goods, or in the sale or advertising of the services....

  8. Effects of using ground redberry juniper and dried distillers grains with solubles in lamb feedlot diets: growth, blood serum, fecal, and wool characteristics.

    PubMed

    Whitney, T R; Lupton, C J; Muir, J P; Adams, R P; Stewart, W C

    2014-03-01

    Effects of using ground redberry juniper and dried distillers grains with solubles (DDGS) in Rambouillet lamb (n = 45) feedlot diets on growth, blood serum, fecal, and wool characteristics were evaluated. In a randomized design study with 2 feeding periods (Period 1 = 64% concentrate diet, 35 d; Period 2 = 85% concentrate diet, 56 d), lambs were individually fed 5 isonitrogenous diets: a control diet (CNTL) that contained oat hay but not DDGS or juniper or DDGS-based diets in which 0 (0JUN), 33 (33JUN), 66 (66JUN), or 100% (100JUN) of the oat hay was replaced by juniper. During Period 1, lambs fed CNTL had greater (P < 0.05) DMI and ADG and tended to have greater (P < 0.10) G:F than lambs fed 0JUN or lambs fed DDGS-based diets. Lamb DMI, ADG, and G:F quadratically increased (P < 0.008) as juniper increased in the DDGS-based diets. During Period 2, lambs fed CNTL had greater (P < 0.05) DMI than lambs fed 0JUN or lambs fed DDGS-based diets, but ADG was similar (P > 0.41). Compared to 0JUN, lambs fed CNTL had similar (P = 0.12) G:F and tended to have less G:F (P = 0.07) than lambs fed DDGS-based diets. Among lambs fed DDGS-based diets, DMI was similar (P > 0.19), ADG increased linearly (P = 0.03), and G:F tended to decrease quadratically (P = 0.06) as juniper increased in the diet. Serum IGF-1, serum urea N (SUN), and fecal N were greater (P < 0.05) and serum Ca and P and fecal P were similar (P > 0.13) for lambs fed CNTL vs. lambs fed DDGS-based diets (CNTL). Within lambs fed DDGS-based diets, SUN increased quadratically (P = 0.01) and fecal N increased linearly (P = 0.004), which can partially be attributed to increased dietary urea and condensed tannin intake. Most wool characteristics were not affected, but wool growth per kilogram of BW decreased quadratically (P = 0.04) as percentage of juniper increased in the DDGS-based diets. When evaluating the entire 91-d feeding trial, results indicated that replacing all of the ground oat hay with ground juniper leaves

  9. Quantitative elementary analysis of medico-biological specimens using a laser mass spectrometer

    SciTech Connect

    Aref'ev, I.M.; Benyaev, N.E.; Komleva, A.A.; Ramendik, G.I.; Tyurin, D.A.

    1986-06-01

    The present work is devoted to an investigation of the dependence of the value of the relative sensitivity coefficient (RSC) on the influence oflaser radiation on medico-biological specimens in order to develop a standardless quantitative analysis. The objects of the investigation were dried tissues of organs of the animals, whole blood and its components. It was shown that the values of the RSC are close to one in the range of 20% error at laser radiation flux densities of 5.10/sup 8/-1.10/sup 9/ W/cm/sup 2/. A decrease or increase in the flux density in comparison with the optimum leads to the appearance of discrimination, in the first case during atomization where the RSC depends on the thermophysical parameters of the investigated elements, and, in the second case, during ionization where the RSC depends on the ionization potentials of the elements.

  10. Grips for Lightweight Tensile Specimens

    NASA Technical Reports Server (NTRS)

    Witte, William G., Jr.; Gibson, Walter D.

    1987-01-01

    Set of grips developed for tensile testing of lightweight composite materials. Double-wedge design substantially increases gripping force and reduces slippage. Specimen held by grips made of hardened wedges. Assembly screwed into load cell in tensile-testing machine.

  11. Manufacturing of Plutonium Tensile Specimens

    SciTech Connect

    Knapp, Cameron M

    2012-08-01

    Details workflow conducted to manufacture high density alpha Plutonium tensile specimens to support Los Alamos National Laboratory's science campaigns. Introduces topics including the metallurgical challenge of Plutonium and the use of high performance super-computing to drive design. Addresses the utilization of Abaqus finite element analysis, programmable computer numerical controlled (CNC) machining, as well as glove box ergonomics and safety in order to design a process that will yield high quality Plutonium tensile specimens.

  12. Typing of Plasmodium falciparum DNA from 2 years old Giemsa-stained dried blood spots using nested polymerase chain reaction assay.

    PubMed

    Kumar, D; Dhiman, S; Rabha, B; Goswami, D; Yadav, K; Deka, M; Veer, V; Baruah, I

    2016-01-01

    A panel of 129 Giemsa-stained thick blood spots (TBS) confirmed for Plasmodium falciparum infection having different levels of parasite density were collected from a malaria endemic area. DNA was extracted and nested polymerase chain reaction (PCR) assay was performed to amplify P. falciparum DNA. Nested PCR assay successfully amplified P. falciparum DNA at a very low parasitaemia of ~10 parasites/μl of blood. Current PCR assay is very simple and can be used retrospectively to monitor the invasion and prevalence of different Plasmodium species in endemic areas.

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

    PubMed

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

    2015-09-01

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

  14. century drying

    NASA Astrophysics Data System (ADS)

    Cook, Benjamin I.; Smerdon, Jason E.; Seager, Richard; Coats, Sloan

    2014-11-01

    Global warming is expected to increase the frequency and intensity of droughts in the twenty-first century, but the relative contributions from changes in moisture supply (precipitation) versus evaporative demand (potential evapotranspiration; PET) have not been comprehensively assessed. Using output from a suite of general circulation model (GCM) simulations from phase 5 of the Coupled Model Intercomparison Project, projected twenty-first century drying and wetting trends are investigated using two offline indices of surface moisture balance: the Palmer Drought Severity Index (PDSI) and the Standardized Precipitation Evapotranspiration Index (SPEI). PDSI and SPEI projections using precipitation and Penman-Monteith based PET changes from the GCMs generally agree, showing robust cross-model drying in western North America, Central America, the Mediterranean, southern Africa, and the Amazon and robust wetting occurring in the Northern Hemisphere high latitudes and east Africa (PDSI only). The SPEI is more sensitive to PET changes than the PDSI, especially in arid regions such as the Sahara and Middle East. Regional drying and wetting patterns largely mirror the spatially heterogeneous response of precipitation in the models, although drying in the PDSI and SPEI calculations extends beyond the regions of reduced precipitation. This expansion of drying areas is attributed to globally widespread increases in PET, caused by increases in surface net radiation and the vapor pressure deficit. Increased PET not only intensifies drying in areas where precipitation is already reduced, it also drives areas into drought that would otherwise experience little drying or even wetting from precipitation trends alone. This PET amplification effect is largest in the Northern Hemisphere mid-latitudes, and is especially pronounced in western North America, Europe, and southeast China. Compared to PDSI projections using precipitation changes only, the projections incorporating both

  15. Chromosome aberrations induced in human lymphocytes by U-235 fission neutrons: I. Irradiation of human blood samples in the "dry cell" of the TRIGA Mark II nuclear reactor.

    PubMed

    Fajgelj, A; Lakoski, A; Horvat, D; Remec, I; Skrk, J; Stegnar, P

    1991-11-01

    A set-up for irradiation of biological samples in the TRIGA Mark II research reactor in Ljubljana is described. Threshold activation detectors were used for characterisation of the neutron flux, and the accompanying gamma dose was measured by TLDs. Human peripheral blood samples were irradiated "in vitro" and biological effects evaluated according to the unstable chromosomal aberrations induced. Biological effects of two types of cultivation of irradiated blood samples, the first immediately after irradiation and the second after 96 h storage, were studied. A significant difference in the incidence of chromosomal aberrations between these two types of samples was obtained, while our dose-response curve fitting coefficients alpha 1 = (7.71 +/- 0.09) x 10(-2) Gy-1 (immediate cultivation) and alpha 2 = (11.03 +/- 0.08) x 10(-2) Gy-1 (96 h delayed cultivation) are in both cases lower than could be found in the literature.

  16. Dried blood spots for the diagnosis and quantitation of HIV-1: stability studies and evaluation of sensitivity and specificity for the diagnosis of infant HIV-1 infection in Thailand.

    PubMed

    Leelawiwat, W; Young, N L; Chaowanachan, T; Ou, C Y; Culnane, M; Vanprapa, N; Waranawat, N; Wasinrapee, P; Mock, P A; Tappero, J; McNicholl, J M

    2009-02-01

    Molecular methods for HIV-1 infection using dried blood-spot (DBS) for HIV-1 CRF01_AE subtypes have not been fully optimized. In this study assays for HIV-1 diagnosis or quantitation were evaluated using infant DBS from Thailand. Paired DBS and whole blood samples from 56 HIV-1 CRF01_AE or B'-infected infants were tested for infant diagnosis using modified Amplicor DNA PCR and NucliSens RNA NASBA and an in-house real-time PCR assay. The Amplicor Monitor viral load (VL) assay, with modifications for DBS, was also evaluated. DBS VL were hematocrit corrected. Stability studies were done on DBS stored at -70 degrees C to 37 degrees C for up to 1 year. The DBS diagnostic assays were 96-100% sensitive and 100% specific for HIV-1 diagnosis. DBS HIV-1 VL were highly correlated with plasma VL when corrected using the actual or an assumed hematocrit factor (r(c)=0.88 or 0.93, respectively). HIV-1 DNA in DBS appeared to be more stable than RNA and could be detected after up to 9 months at most temperatures. DBS VL could be consistently determined when stored frozen. These results show that DBS can be used accurately instead of whole blood for the diagnosis of HIV-1 infection and VL quantitation, particularly if samples are appropriately stored.

  17. Dry cell battery poisoning

    MedlinePlus

    Batteries - dry cell ... Acidic dry cell batteries contain: Manganese dioxide Ammonium chloride Alkaline dry cell batteries contain: Sodium hydroxide Potassium hydroxide Lithium dioxide dry cell batteries ...

  18. Positron lifetime studies in thermoplastic polyimide test specimens

    NASA Technical Reports Server (NTRS)

    Singh, J. J.; Stclair, T. L.; Holt, W. H.; Mock, W., Jr.

    1982-01-01

    Positron lifetime measurements were made in two thermoplastic polyimide materials recently developed at Langley. The long component lifetime values in polyimidesulfone samples are 847 + or - 81 Ps (dry) and 764 + or - 91 Ps (saturated). The corresponding values in LARC thermoplastic imides are 1080 + or - 139 Ps (dry) and 711 + or - 96 Ps (saturated). Clearly, the presence of moisture has greater effect on positron lifetime in LARC thermoplastic imides than in the case of polyimidesulfones. This result is consistent with the photomicrographic observations made on frozen water saturated specimens of these materials.

  19. Collection & Processing of Vertebrate Specimens for Arbovirus Studies.

    ERIC Educational Resources Information Center

    Sudia, W. Daniel; And Others

    Described are techniques used by the National Communicable Disease Center in obtaining blood and tissues from man and other vertebrates for arbovirus isolation and antibody studies. Also included are techniques for capturing and handling vertebrates; banding and marking; restraining and bleeding; storing of specimens to preserve antibody and…

  20. An interlaminar tension strength specimen

    NASA Technical Reports Server (NTRS)

    Jackson, Wade C.; Martin, Roderick H.

    1992-01-01

    This paper describes a technique to determine interlaminar tension strength, sigma(sub 3c) of a fiber reinforced composite material using a curved beam. The specimen was a unidirectional curved beam, bent 90 degrees, with straight arms. Attached to each arm was a hinged loading mechanism which was held by the grips of a tensile testing machine. Geometry effects of the specimen, including the effects of loading arm length, inner radius, thickness, and width, were studied. The data sets fell into two categories: low strength corresponding to a macroscopic flaw related failure and high strength corresponding to a microscopic flaw related failure. From the data available, the loading arm length had no effect on sigma(sub 3c). The inner radius was not expected to have a significant effect on sigma(sub 3c), but this conclusion could not be confirmed because of differences in laminate quality for each curve geometry. The thicker specimens had the lowest value of sigma(sub 3c) because of poor laminate quality. Width was found to affect the value of sigma(sub 3c) only slightly. The wider specimens generally had a slightly lower strength since more material was under high stress, and hence, had a larger probability of containing a significant flaw.

  1. Effects of a Brown-midrib corn hybrid on nutrient digestibility in wethers and on dry matter intake, performance, rumen and blood variables in dairy cows.

    PubMed

    Gorniak, T; Meyer, U; Hackelsperger, F; Dänicke, S

    2014-04-01

    The aim of the present trials was to determine the effect of an experimental Brown-midrib (Bm) corn hybrid in relation to a commercial corn hybrid (Con) on digestibility in wethers and on dry matter intake (DMI), milk yield and milk composition in dairy cows. Digestibility of crude fibre (CF), neutral detergent fibre (NDFom) and acid detergent fibre (ADFom) were higher for Bm (CF Con: 57.8%; Bm: 67.2%; NDFom Con: 56.8%; Bm: 64.8%; ADFom Con: 52.0%; Bm: 63.9%), but concentration of net energy for lactation did not differ (Con: 6.4 MJ/kg DM; Bm: 6.3 MJ/kg DM). A total of 64 lactating German Holstein cows were assigned to one of the two dietary treatments Con or Bm according to milk yield, lactation number, days in milk and live weight. In Trial 1, cows were fed a total mixed ration consisting of 50% corn silage (Con or Bm) and 50% concentrate on dry matter (DM) basis. In Trial 2, the same animals were fed the respective silage for ad libitum intake and 5.3 kg of concentrate DM per animal per day. In Trial 1, DMI and milk-fat content were decreased significantly for the Bm-treatment (DMI Con: 22.5 kg/day; Bm: 21.5 kg/day; milk fat Con: 3.8%; Bm: 3.3%). In Trial 2, milk yield and fat-corrected milk (FCM) were increased significantly, whereas milk-fat% was decreased significantly (milk yield Con: 25.8 kg/day; Bm: 29.4 kg/day; FCM Con: 27.2 kg/day; Bm: 29.6 kg/day; fat Con: 4.4%; Bm: 4.0%). Diets did not influence ruminal pH or temperature. Diets, furthermore, did not influence rumination in either trial. Additional research on digestibility and rumen fermentation should, however, be carried out using dairy cows at respective intake levels as trials with wethers cannot be transferred to high-yielding ad libitum fed cows.

  2. Colorful drying.

    PubMed

    Lakio, Satu; Heinämäki, Jyrki; Yliruusi, Jouko

    2010-03-01

    Drying is one of the standard unit operations in the pharmaceutical industry and it is important to become aware of the circumstances that dominate during the process. The purpose of this study was to test microcapsulated thermochromic pigments as heat indicators in a fluid bed drying process. The indicator powders were manually granulated with alpha-lactose monohydrate resulting in three particle-size groups. Also, pellets were coated with the indicator powders. The granules and pellets were fluidized in fluid bed dryer to observe the progress of the heat flow in the material and to study the heat indicator properties of the indicator materials. A tristimulus colorimeter was used to measure CIELAB color values. Color indicator for heat detection can be utilized to test if the heat-sensitive API would go through physical changes during the pharmaceutical drying process. Both the prepared granules and pellets can be used as heat indicator in fluid bed drying process. The colored heat indicators give an opportunity to learn new aspects of the process at real time and could be exploded, for example, for scaling-up studies.

  3. X-ray microanalysis of frozen-hydrated specimens.

    PubMed

    Zierold, K

    1983-01-01

    The preparation of frozen-hydrated bulk specimens and sections for X-ray microanalysis starts with cryofixation, which is done either by rapid immersion into liquid propane, propane jet fixation or metal mirror fixation. Bulk specimens appropriate for the analysis in a scanning electron microscope (SEM) are obtained by cryofracturing the samples, coating, usually by a thin carbon layer, and cold transfer into the cold stage of the microscope. The X-ray microanalysis of bulk specimens is affected by an internal space charge which makes quantification difficult. Frozen-hydrated dry cut sections, varying in thickness between 60 and 2000 nm, are prepared by means of cryoultramicrotomy. After cold transfer into the cold stage of a scanning electron microscope or a scanning transmission electron microscope (STEM) the sections are analyzed in the frozen-hydrated and freeze-dried state. The reliability of the results with regard to structural recognizability and X-ray spectra depends considerably on the state of hydration. Particularly ultrathin sections in STEM show very low contrast and a great mass loss in the frozen-hydrated state in comparison with the freeze-dried state. In spite of the available concepts for quantification of X-ray data to obtain physiologically important wet weight concentrations of diffusible elements, the radiation damage at present turns out to be the most serious problem for X-ray microanalysis of frozen-hydrated sections.

  4. Baseline Test Specimen Machining Report

    SciTech Connect

    mark Carroll

    2009-08-01

    The Next Generation Nuclear Plant (NGNP) Project is tasked with selecting a high temperature gas reactor technology that will be capable of generating electricity and supplying large amounts of process heat. The NGNP is presently being designed as a helium-cooled high temperature gas reactor (HTGR) with a large graphite core. The graphite baseline characterization project is conducting the research and development (R&D) activities deemed necessary to fully qualify nuclear-grade graphite for use in the NGNP reactor. Establishing nonirradiated thermomechanical and thermophysical properties by characterizing lot-to-lot and billet-to-billet variations (for probabilistic baseline data needs) through extensive data collection and statistical analysis is one of the major fundamental objectives of the project. The reactor core will be made up of stacks of graphite moderator blocks. In order to gain a more comprehensive understanding of the varying characteristics in a wide range of suitable graphites, any of which can be classified as “nuclear grade,” an experimental program has been initiated to develop an extensive database of the baseline characteristics of numerous candidate graphites. Various factors known to affect the properties of graphite will be investigated, including specimen size, spatial location within a graphite billet, specimen orientation within a billet (either parallel to [P] or transverse to [T] the long axis of the as-produced billet), and billet-to-billet variations within a lot or across different production lots. Because each data point is based on a certain position within a given billet of graphite, particular attention must be paid to the traceability of each specimen and its spatial location and orientation within each billet. The evaluation of these properties is discussed in the Graphite Technology Development Plan (Windes et. al, 2007). One of the key components in the evaluation of these graphite types will be mechanical testing on

  5. Effects of feeding a spray-dried multivalent polyclonal antibody preparation on feedlot performance, feeding behavior, carcass characteristics, rumenitis, and blood gas profile of Brangus and Nellore yearling bulls.

    PubMed

    Millen, D D; Pacheco, R D L; DiLorenzo, N; Martins, C L; Marino, C T; Bastos, J P S T; Mariani, T M; Barducci, R S; Sarti, L M N; DiCostanzo, A; Rodrigues, P H M; Arrigoni, M D B

    2015-09-01

    The objective of this study was to evaluate the effects of replacing monensin (MON) with a spray-dried multivalent polyclonal antibody preparation (PAP) against several ruminal microorganisms on feedlot performance, carcass characteristics, feeding behavior, blood gas profile, and the rumenitis incidence of Brangus and Nellore yearling bulls. The study was designed as a completely randomized design with a 2 × 2 factorial arrangement, replicated 6 times (4 bulls per pen and a total of 24 pens), in which bulls ( = 48) of each biotype were fed diets containing either MON fed at 300 mg/d or PAP fed at 3 g/d. No significant feed additive main effects were observed for ADG ( = 0.27), G:F ( = 0.28), HCW ( = 0.99), or dressing percentage ( = 0.80). However, bulls receiving PAP had greater DMI ( = 0.02) and larger ( = 0.02) final LM area as well as greater ( < 0.01) blood concentrations of bicarbonate and base excess in the extracellular fluid than bulls receiving MON. Brangus bulls had greater ( < 0.01) ADG and DMI expressed in kilograms, final BW, heavier HCW, and larger initial and final LM area than Nellore bulls. However, Nellore bulls had greater daily DMI fluctuation ( < 0.01), expressed as a percentage, and greater incidence of rumenitis ( = 0.05) than Brangus bulls. In addition, Brangus bulls had greater ( < 0.01) DMI per meal and also presented lower ( < 0.01) DM and NDF rumination rates when compared with Nellore bulls. Significant interactions ( < 0.05) between biotype and feed additive were observed for SFA, unsaturated fatty acids (UFA), MUFA, and PUFA concentrations in adipose tissues. When Nellore bulls were fed PAP, fat had greater ( < 0.05) SFA and PUFA contents but less ( < 0.01) UFA and MUFA than Nellore bulls receiving MON. For Brangus bulls, MON led to greater ( < 0.05) SFA and PUFA and less ( < 0.05) UFA and MUFA than Brangus bulls fed PAP. Feeding a spray-dried PAP led to similar feedlot performance compared with that when feeding MON. Spray-dried

  6. A specific LC/ESI-MS/MS method for determination of 25-hydroxyvitamin D3 in neonatal dried blood spots containing a potential interfering metabolite, 3-epi-25-hydroxyvitamin D3.

    PubMed

    Higashi, Tatsuya; Suzuki, Masahiro; Hanai, Junji; Inagaki, Shinsuke; Min, Jun Zhe; Shimada, Kazutake; Toyo'oka, Toshimasa

    2011-04-01

    Vitamin D deficiency in an infant is associated with a wide range of adverse health outcomes in later life. A method for the quantification of 25-hydroxyvitamin D(3) [25(OH)D(3), the best-established indicator of vitamin D status] in neonatal dried blood spots (DBSs) using LC/ESI-MS/MS has been developed and validated. The method employed two steps of derivatization, a Diels-Alder reaction with 4-phenyl-1,2,4-triazoline-3,5-dione followed by acetylation, to enhance the detectability of 25(OH)D(3) in ESI-MS/MS and to separate 25(OH)D(3) from 3-epi-25-hydroxyvitamin D(3) [3-epi-25(OH)D(3)], a potent interfering metabolite. 25(OH)D(3) was extracted from two DBS punches (3  mm in diameter, equivalent to 5.3  μL of whole blood), purified using an Oasis HLB(®) cartridge, and subjected to derivatization prior to analysis with LC/ESI-MS/MS. 25-Hydroxyvitamin D(4) was used as the internal standard. This method was reproducible (intra- and inter-assay RSDs, <6.9%) and accurate (analytical recovery, 95.2-102.7%), and the LOQ was 3.0  ng/mL. The developed method enabled specific quantification of 25(OH)D(3) in neonatal DBSs and detection of vitamin D deficiency without interference from 3-epi-25(OH)D(3).

  7. Appendix A: Specimen 72275 documentation

    NASA Technical Reports Server (NTRS)

    Marvin, U. B.

    1974-01-01

    The friability of the matrix of specimen 72275 caused numerous fragments and an abundance of fines to break away from the main mass during transport from the moon and handling in the lunar receiving laboratory. Samples 72275,1 to 72275,14 were labeled during PET examination. Samples 72275,1, 4, 6, 7, 8, and 9 were placed in storage, and the remainder were distributed.

  8. Concepts of marine specimen banking.

    PubMed

    Rossbach, M; Kniewald, G

    1997-05-01

    For more than a decade environmental specimen banking (ESB) has been an established approach for monitoring and retrospective environmental survey purposes in a number of developed countries. Specimen banking is carried out on regional or national scales for various environmental materials. The ecological or problem-oriented approach, as pursued e.g. in Germany or USA has the advantages of a restricted survey and a clear political mandate. Environmental problems, however, are by no means national or regional issues, since the diversity and dispersion of hazardous substances make environmental monitoring clearly a global affair. The structuring of our environment suggests that banking should not be limited by national boundaries, but rather be based on eco-systematic principles. Such distinct banking efforts should be devoted to the monitoring of physico-chemical aspects of climatic change and air pollution, soil quality, and aquatic monitoring on a world-wide scale. As some experience already exists with specialized banking programs for marine samples, such as the National Marine Mammal Tissue Bank or the Mussel Watch Program in the United States, an international marine specimen bank, based on principles of national ESB's, is advocated to be established in due time. Following the recommendations of the 1992 Rio 'Earth Summit' to pursue sustainable development strategies, such an establishment could strongly facilitate efforts concerning pollution control and mitigation, overexploitation and mining of ocean resources on a regional or global scale.

  9. Performance of an Early Infant Diagnostic Test, AmpliSens DNA-HIV-FRT, Using Dried Blood Spots Collected from Children Born to Human Immunodeficiency Virus-Infected Mothers in Ukraine

    PubMed Central

    Shanmugam, Vedapuri; Azarskova, Marianna; Nguyen, Shon; Hurlston, Mackenzie; Sabatier, Jennifer; Zhang, Guoqing; Osmanov, Saladin; Ellenberger, Dennis; Yang, Chunfu; Vitek, Charles; Liulchuk, Maria; Nizova, Natalya

    2015-01-01

    An accurate accessible test for early infant diagnosis (EID) is crucial for identifying HIV-infected infants and linking them to treatment. To improve EID services in Ukraine, dried blood spot (DBS) samples obtained from 237 HIV-exposed children (≤18 months of age) in six regions in Ukraine in 2012 to 2013 were tested with the AmpliSens DNA-HIV-FRT assay, the Roche COBAS AmpliPrep/COBAS TaqMan (CAP/CTM) HIV-1 Qual test, and the Abbott RealTime HIV-1 Qualitative assay. In comparison with the paired whole-blood results generated from AmpliSens testing at the oblast HIV reference laboratories in Ukraine, the sensitivity was 0.99 (95% confidence interval [CI], 0.95 to 1.00) for the AmpliSens and Roche CAP/CTM Qual assays and 0.96 (95% CI, 0.90 to 0.98) for the Abbott Qualitative assay. The specificity was 1.00 (95% CI, 0.97 to 1.00) for the AmpliSens and Abbott Qualitative assays and 0.99 (95% CI, 0.96 to 1.00) for the Roche CAP/CTM Qual assay. McNemar analysis indicated that the proportions of positive results for the tests were not significantly different (P > 0.05). Cohen's kappa (0.97 to 0.99) indicated almost perfect agreement among the three tests. These results indicated that the AmpliSens DBS and whole-blood tests performed equally well and were comparable to the two commercially available EID tests. More importantly, the performance characteristics of the AmpliSens DBS test meets the World Health Organization EID test requirements; implementing AmpliSens DBS testing might improve EID services in resource-limited settings. PMID:26447114

  10. Standard Specimen Reference Set: Breast — EDRN Public Portal

    Cancer.gov

    The primary objective of this study is to assemble a well-characterized set of blood specimens to test biomarkers that, in conjunction with mammography, can detect and discriminate breast cancer. These samples will be divided to provide “sets” of specimens that can be tested in a number of different laboratories. Since tests will be performed on the same sets of samples, the data will be directly comparable and decisions regarding which biomarker or set of biomarkers have value in breast cancer detection can be made. These sets will reside at a National Cancer Institute facility at Frederick, MD.

  11. Induced Polarization Responses of the Specimen with Sulfide Ore Minerals

    NASA Astrophysics Data System (ADS)

    Park, S.; Sung, N. H.

    2012-04-01

    Basic data of the physical properties of the rocks is required to effectively interpret geologic structures and mineralized zones in study areas from the geophysical data in the field of subsurface investigations and mineral resources explorations. In this study, the spectral induced polarization (SIP) measurement system in the laboratory was constructed to obtain the IP characteristics of the specimen with sulfide ore minerals. The SIP measurement system consists of lab transmitter for electrical current transmission, and GDP-32 for current receiver. The SIP system employs 14 steps of frequencies from 0.123 to 1,024 Hz, and uses copper sulfate solution as an electrolyte. The SIP data for system verification was acquired using a measurement system of parallel circuit with fixed resistance and condenser. This measured data was in good agreement with Cole-Cole model data. First of all, the experiment on the SIP response was conducted in the laboratory with the mixture of glass beads and pyrite powders for ore grade assessment using characteristics of IP response of the rocks. The results show that the phase difference of IP response to the frequency is nearly proportional to the weight content of pyrite, and that the dominant frequency of the IP response varies with the size of the pyrite powder. Subsequently, the specimens used for SIP measurement are slate and limestone which were taken from drilling cores and outcrops of skarn ore deposits. All specimens are cylindrical in shape, with a diameter of 5 cm and a length of 10 cm. When measuring SIP of water-saturated specimens, the specimen surface is kept dry, tap water is put into the bottom of sample holder and a lid is closed. It is drawn that the SIP characteristics of the rocks show the phase difference depends on the amount of the sulfide minerals. The phase difference did not occur with frequencies applied in the absence of sulfide minerals in the rock specimens. On the contrary, the rock specimens containing

  12. Automated Tracking of Drosophila Specimens

    PubMed Central

    Chao, Rubén; Macía-Vázquez, Germán; Zalama, Eduardo; Gómez-García-Bermejo, Jaime; Perán, José-Ramón

    2015-01-01

    The fruit fly Drosophila Melanogaster has become a model organism in the study of neurobiology and behavior patterns. The analysis of the way the fly moves and its behavior is of great scientific interest for research on aspects such as drug tolerance, aggression or ageing in humans. In this article, a procedure for detecting, identifying and tracking numerous specimens of Drosophila by means of computer vision-based sensing systems is presented. This procedure allows dynamic information about each specimen to be collected at each moment, and then for its behavior to be quantitatively characterized. The proposed algorithm operates in three main steps: a pre-processing step, a detection and segmentation step, and tracking shape. The pre-processing and segmentation steps allow some limits of the image acquisition system and some visual artifacts (such as shadows and reflections) to be dealt with. The improvements introduced in the tracking step allow the problems corresponding to identity loss and swaps, caused by the interaction between individual flies, to be solved efficiently. Thus, a robust method that compares favorably to other existing methods is obtained. PMID:26258779

  13. 36 CFR 2.5 - Research specimens.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... superintendent approves a written research proposal and determines that the collection will benefit science or... Park Service National Catalog. (2) Specimens and data derived from consumed specimens will be...

  14. 36 CFR 2.5 - Research specimens.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... superintendent approves a written research proposal and determines that the collection will benefit science or... Park Service National Catalog. (2) Specimens and data derived from consumed specimens will be...

  15. Testing Biopsy and Cytology Specimens for Cancer

    MedlinePlus

    ... Exams and Tests for Cancer Testing Biopsy and Cytology Specimens for Cancer Waiting to hear a possible ... best decisions about your treatment. Testing Biopsy and Cytology Specimens for Cancer How is cancer diagnosed? Types ...

  16. Strong Correlation Between Concentrations of Tenofovir (TFV) Emtricitabine (FTC) in Hair and TFV Diphosphate and FTC Triphosphate in Dried Blood Spots in the iPrEx Open Label Extension: Implications for Pre-exposure Prophylaxis Adherence Monitoring.

    PubMed

    Gandhi, Monica; Glidden, David V; Liu, Albert; Anderson, Peter L; Horng, Howard; Defechereux, Patricia; Guanira, Juan V; Grinsztejn, Beatriz; Chariyalertsak, Suwat; Bekker, Linda-Gail; Grant, Robert M

    2015-11-01

    Self-reported adherence to pre-exposure prophylaxis (PrEP) has limitations, raising interest in pharmacologic monitoring. Drug concentrations in hair and dried blood spots (DBS) are used to assess long-term-exposure; hair shipment/storage occurs at room temperature. The iPrEx Open Label Extension collected DBS routinely, with opt-in hair collection; concentrations were measured with liquid chromatography/tandem mass spectrometry. In 806 hair-DBS pairs, tenofovir (TFV) hair levels and TFV diphosphate (DP) in DBS were strongly correlated (Spearman coefficient r = 0.734; P < .001), as were hair TFV/DBS emtricitabine (FTC) triphosphate (TP) (r = 0.781; P < .001); hair FTC/DBS TFV-DP (r = 0.74; P < .001); hair FTC/DBS FTC-TP (r = 0.587; P < .001). Drug detectability was generally concordant by matrix. Hair TFV/FTC concentrations correlate strongly with DBS levels, which are predictive of PrEP outcomes.

  17. [Use of dried blood spots in early diagnosis of HIV-1 infection in children born to HIV-infected mothers as part of the prevention of mother-to-child transmission in Benin].

    PubMed

    Tchiakpe, E; Hounto-Ogouyemi, A; Diop Ndiaye, H; Diouara, A A M; Aïssi, A K; Keke, R K; Kpangon, A A; Lafia, B; Métadokou, D; Bouraïma, B; Anthony, D; Hounsinou, A; Alao, M J; Azondekon, A; Ahouidi, A D; Bei, A K; Mbengue, M A S; Touré Kane, C; Zannou, D M

    2016-08-01

    The goal of this study was to evaluate using the molecular diagnosis, infection transmission rate of HIV in children born to HIV-1 positive mothers as part of the prevention of mother-to-child transmission (PMTCT) in Benin. The sample consisted of 524 dried blood spots (DBS) of children born to HIV-1 positive mothers, from 30 sites (PMTCT) taken between October 2009 and June 2010. The diagnosis of HIV-1 was performed by the qualitative detection of viral nucleic acids (RNA and DNA) in DBS on filter paper using the Abbott RealTime(®) HIV-1 Qualitative assay. We found that 51 DBS were positive (9.7%) and 473 were negative (90.3%). The failure rate of PMTCT among 420 mothers who received antiretroviral prophylaxis was 6.7% (28/420). This failure rate was significantly higher among children born to infected mothers on antiretroviral monotherapy than on triple therapy (HAART). The results of our study enrich the data in the literature on highly active antiretroviral chemoprophylaxis to reduce the transmission of HIV-1 from mother to child.

  18. Field evaluation of Abbott Real Time HIV-1 Qualitative test for early infant diagnosis using dried blood spots samples in comparison to Roche COBAS Ampliprep/COBAS TaqMan HIV-1 Qual test in Kenya.

    PubMed

    Chang, Joy; Omuomo, Kenneth; Anyango, Emily; Kingwara, Leonard; Basiye, Frank; Morwabe, Alex; Shanmugam, Vedapuri; Nguyen, Shon; Sabatier, Jennifer; Zeh, Clement; Ellenberger, Dennis

    2014-08-01

    Timely diagnosis and treatment of infants infected with HIV are critical for reducing infant mortality. High-throughput automated diagnostic tests like Roche COBAS AmpliPrep/COBAS TaqMan HIV-1 Qual Test (Roche CAPCTM Qual) and the Abbott Real Time HIV-1 Qualitative (Abbott Qualitative) can be used to rapidly expand early infant diagnosis testing services. In this study, the performance characteristics of the Abbott Qualitative were evaluated using two hundred dried blood spots (DBS) samples (100 HIV-1 positive and 100 HIV-1 negative) collected from infants attending the antenatal facilities in Kisumu, Kenya. The Abbott Qualitative results were compared to the diagnostic testing completed using the Roche CAPCTM Qual in Kenya. The sensitivity and specificity of the Abbott Qualitative were 99.0% (95% CI: 95.0-100.0) and 100.0% (95% CI: 96.0-100.0), respectively, and the overall reproducibility was 98.0% (95% CI: 86.0-100.0). The limits of detection for the Abbott Qualitative and Roche CAPCTM Qual were 56.5 and 6.9copies/mL at 95% CIs (p=0.005), respectively. The study findings demonstrate that the Abbott Qualitative test is a practical option for timely diagnosis of HIV in infants.

  19. Imaging of concrete specimens using inverse synthetic aperture radar

    NASA Astrophysics Data System (ADS)

    Rhim, Hong C.; Buyukozturk, Oral

    2000-05-01

    Radar Measurement results of laboratory size concrete specimens are presented in this paper. The purpose of this research work is to study various aspects of the radar method in an effort to develop an improved radar system for nondestructive testing of concrete structures. The radar system used for the study is an Inverse Synthetic Aperture Radar (ISAR), which is capable of transmitting microwaves at three different frequency ranges of 2-3.4, 3.4-5.8, and 8-12 GHz. Radar measurement setup is such that the radar is locates 14.4 m away from a concrete target to satisfy a far-field criterion. The concrete target is rotated for 20 degrees during the measurements for the generation of two-dimensional (cross-range) imagery. Concrete targets used for the measurements have the dimensions of 305 mm (width)×305 mm (height)×92 mm (thickness) with different inside configurations. Comparisons are made for dry and wet specimens, specimens with and without inclusions. Each specimen is made to model various situations that a concrete structure can have in reality. Results show that center frequency, frequency bandwidth, and polarization of the incident wave have different effects on identifying the thickness or inclusions inside concrete specimens. Results also suggest that a certain combination of measurement parameters is suitable for a specific application area. Thus, measurement parameters can be optimized for a specific problem. The findings are presented and discussed in details in the paper. Signal processing schemes implemented for imaging of the specimens are also discussed.

  20. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 1 2014-01-01 2014-01-01 false Split specimens. 26.135 Section 26.135 Energy NUCLEAR REGULATORY COMMISSION FITNESS FOR DUTY PROGRAMS Licensee Testing Facilities § 26.135 Split specimens. (a) If the FFD program follows split-specimen procedures, as described in § 26.113, the licensee...

  1. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 1 2011-01-01 2011-01-01 false Split specimens. 26.135 Section 26.135 Energy NUCLEAR REGULATORY COMMISSION FITNESS FOR DUTY PROGRAMS Licensee Testing Facilities § 26.135 Split specimens. (a) If the FFD program follows split-specimen procedures, as described in § 26.113, the licensee...

  2. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 1 2013-01-01 2013-01-01 false Split specimens. 26.135 Section 26.135 Energy NUCLEAR REGULATORY COMMISSION FITNESS FOR DUTY PROGRAMS Licensee Testing Facilities § 26.135 Split specimens. (a) If the FFD program follows split-specimen procedures, as described in § 26.113, the licensee...

  3. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 1 2012-01-01 2012-01-01 false Split specimens. 26.135 Section 26.135 Energy NUCLEAR REGULATORY COMMISSION FITNESS FOR DUTY PROGRAMS Licensee Testing Facilities § 26.135 Split specimens. (a) If the FFD program follows split-specimen procedures, as described in § 26.113, the licensee...

  4. 10 CFR 26.135 - Split specimens.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 1 2010-01-01 2010-01-01 false Split specimens. 26.135 Section 26.135 Energy NUCLEAR REGULATORY COMMISSION FITNESS FOR DUTY PROGRAMS Licensee Testing Facilities § 26.135 Split specimens. (a) If the FFD program follows split-specimen procedures, as described in § 26.113, the licensee...

  5. Blood sugar test - blood

    MedlinePlus

    ... sugar; Blood sugar level; Fasting blood sugar; Glucose test; Diabetic screening - blood sugar test; Diabetes - blood sugar test ... The test may be done in the following ways: After you have not eaten anything for at least 8 ...

  6. Freeze-Dried Human Red Blood Cells

    DTIC Science & Technology

    1991-07-12

    starting cells are lost at rehydration). Hypotonic or hypertonic lysis of red cells can disrupt the normal asymmetric distribution of phospholipids between...remove the buffy coat and plasma. The packed RBC were washed in isotonic dextrose saline according to standard washing procedures (11] using an automated...cell washer ( Model 2991, COBE, Lakewood, CO). The washed and packed RBC (about 85% hematocrit) were resuspended to about 40% in hypertonic phosphate

  7. Morphine-3-D glucuronide stability in postmortem specimens exposed to bacterial enzymatic hydrolysis.

    PubMed

    Carroll, F T; Marraccini, J V; Lewis, S; Wright, W

    2000-12-01

    Medical examiners frequently rely on the finding of free morphine present in postmortem specimens to assist in certifying deaths associated with narcotics. In vitro hydrolysis of morphine-3-D glucuronide (M3DG) to free morphine was studied using variable specimen pH, initial degree of specimen putrefaction, storage temperature and time, and the effectiveness of sodium fluoride (NaF) preservation. Reagent M3DG was added to opiate-free fresh blood and urine and to autopsy-derived blood specimens. Reagent bovine glucuronidase was also added to certain specimens. Freshly collected and refrigerated NaF-preserved blood produced minimal free morphine, whereas four of five autopsy blood specimens produced free morphine from M3DG. Increased storage time, temperature, and initial degree of putrefaction resulted in greater free morphine generation despite the absence of viable bacteria. Hydrolysis occurring during specimen storage can generate free morphine from M3DG and may result in erroneous conclusions in certifying narcotic deaths.

  8. Specimen rejection in laboratory medicine: Necessary for patient safety?

    PubMed Central

    Dikmen, Zeliha Gunnur; Pinar, Asli; Akbiyik, Filiz

    2015-01-01

    Introduction The emergency laboratory in Hacettepe University Hospitals receives specimens from emergency departments (EDs), inpatient services and intensive care units (ICUs). The samples are accepted according to the rejection criteria of the laboratory. In this study, we aimed to evaluate the sample rejection ratios according to the types of pre-preanalytical errors and collection areas. Materials and methods The samples sent to the emergency laboratory were recorded during 12 months between January to December, 2013 in which 453,171 samples were received and 27,067 specimens were rejected. Results Rejection ratios was 2.5% for biochemistry tests, 3.2% for complete blood count (CBC), 9.8% for blood gases, 9.2% for urine analysis, 13.3% for coagulation tests, 12.8% for therapeutic drug monitoring, 3.5% for cardiac markers and 12% for hormone tests. The most frequent rejection reasons were fibrin clots (28%) and inadequate volume (9%) for biochemical tests. Clotted samples (35%) and inadequate volume (13%) were the major causes for coagulation tests, blood gas analyses and CBC. The ratio of rejected specimens was higher in the EDs (40%) compared to ICUs (30%) and inpatient services (28%). The highest rejection ratio was observed in neurology ICU (14%) among the ICUs and internal medicine inpatient service (10%) within inpatient clinics. Conclusions We detected an overall specimen rejection rate of 6% in emergency laboratory. By documentation of rejected samples and periodic training of healthcare personnel, we expect to decrease sample rejection ratios below 2%, improve total quality management of the emergency laboratory and promote patient safety. PMID:26527231

  9. Use of Low Temperature Scanning Electron Microscopy to Observe Frozen Hydrated Specimens of Nematodes

    PubMed Central

    Wergin, William P.; Sayre, Richard M.; Erbe, Eric F.

    1993-01-01

    Frozen hydrated specimens of Pratylenchus agilis and dauer larvae of Steinernema carpocapsae were observed with low-temperature field emission scanning electron microscopy. This new technique provides information about the surface features of nematodes and also allows specimens to be fractured to reveal their internal structure. Furthermore, both halves of fractured specimens can be retained, examined, and photographed either as two-dimensional micrographs or as three-dimensional images for stereo observation (stereology) or quantitative measurements (stereometry). This technique avoids artifacts normally associated with procedures required to prepare nematodes for examination in the transmission and scanning electron microscopes, such as chemical fixation, dehydration, and sectioning or critical point drying. PMID:19279761

  10. [The German Environmental Specimen Bank].

    PubMed

    Schröter-Kermani, Christa; Gies, Andreas; Kolossa-Gehring, Marike

    2016-03-01

    The main objective of the German Environmental Specimen Bank (ESB) is the long-term storage of environmental and human samples under stable deep-freeze conditions for future research. The ESB is unique in providing a continuous historical record of environmental and human exposure to chemicals in Germany. ESB was started parallel to the development of the first German Chemicals Legislation in the late 1970s. In 1979, the ESB test operation began. After the Chemicals Law came into force in 1982, the ESB was established as a permanent facility in 1985. With the new European Chemicals Legislation, REACH, in 2007 responsibility for the safety of commercial chemicals and risk assessment was assigned to the industry. Since then, the ESB has become even more important in verifying the self-assessment of the industry, in evaluating the effectiveness of regulations, thus ensuring the protection of humans and the environment against adverse effects caused by exposure to chemicals. These objectives are pursued by the regular monitoring of contaminations and the assessment of temporal trends. Demonstrating the necessity of deriving exposure reduction measures, ESB results serve as key information for policy-makers. Information on preventing exposure to chemicals is available to the general public and to the public health services. The ESB is thus an important monitoring instrument of the Federal Ministry for the Environment, Nature Conservation, Building and Nuclear Safety. The Federal Environment Agency operates the ESB based on its own concepts, heads the scientific data evaluation and transfers results into the environmental policy arena and to the general public.

  11. Alfredo Dugès' type specimens of amphibians and reptiles revisited.

    PubMed

    Flores-Villela, Oscar; Ríos-Muñoz, César A; Magaña-Cota, Gloria E; Quezadas-Tapia, Néstor L

    2016-03-14

    The type specimens of amphibians and reptiles of the Museo de Historia Natural Alfredo Dugès, at the University of Guanajuato (MADUG) were reviewed following Smith & Necker's (1943) summary. Owing to this collection's eventful history and its historical importance as the oldest herpetological collection in Mexico, a review of its conservation status was needed. After many years, the collection has received proper recognition at the University of Guanajuato with a portion of the herpetological types considered "Precious Assets" of the university. We found 34 type specimens pertaining to 18 taxa; six are additional specimens to those previously reported; six herpetological types are missing, including the body of the type of Adelophis copei. All specimens are in good to reasonable condition except for the type of Rhinocheilus antonii, which has dried out completely. All specimens are illustrated to show their condition.

  12. Blood typing

    MedlinePlus

    ... ABO blood typing; Blood group; Anemia - immune hemolytic blood type; ABO blood type; A blood type; AB blood type; O blood type ... The 2 steps above can accurately determine your blood type. Rh typing uses a method similar to ABO ...

  13. Flat tensile specimen design for advanced composites

    NASA Technical Reports Server (NTRS)

    Worthem, Dennis W.

    1990-01-01

    Finite element analyses of flat, reduced gage section tensile specimens with various transition region contours were performed. Within dimensional constraints, such as maximum length, tab region width, gage width, gage length, and minimum tab length, a transition contour radius of 41.9 cm produced the lowest stress values in the specimen transition region. The stresses in the transition region were not sensitive to specimen material properties. The stresses in the tab region were sensitive to specimen composite and/or tab material properties. An evaluation of stresses with different specimen composite and tab material combinations must account for material nonlinearity of both the tab and the specimen composite. Material nonlinearity can either relieve stresses in the composite under the tab or elevate them to cause failure under the tab.

  14. Standardizing the Handling of Surgical Specimens.

    PubMed

    Shirey, Cheryl; Perrego, Kristen

    2015-11-01

    To standardize the handling of surgical specimens, the OR clinical educators in a community hospital setting devised a series of departmental changes as a quality improvement project. A newly created skill validation was reviewed in an hour-long educational meeting with all OR staff members. In addition to creating a new annual skill validation, discussions about specimens were included in the hand over, the time out, and a newly instituted debriefing tool to be used toward the end of a procedure. This interdisciplinary group devised interventions to improve the process of handling specimens such as standardizing the labeling process and changing the transparency of the specimen container. The goal was to assure standardization of specimen handling, specifically to assist novice staff members, and to harmonize inconsistencies between specialties within the practice of existing staff members. These combined methods helped to ensure accurate communication and procurement of specimens for all procedures.

  15. Dry Macular Degeneration

    MedlinePlus

    Dry macular degeneration Overview By Mayo Clinic Staff Dry macular degeneration is a common eye disorder among people over 65. ... vision in your direct line of sight. Dry macular degeneration may first develop in one eye and then ...

  16. Study of Biological Pigments by Single Specimen Derivative Spectrophotometry

    PubMed Central

    Goldstein, Jack M.

    1970-01-01

    The single specimen derivative (SSD) method provides an absolute absorption spectrum of a substance in the absence of a suitable reference. Both a reference and a measuring monochromatic beam pass through a single sample, and the specimen itself acts as its own reference. The two monochromatic beams maintain a fixed wavelength difference upon scanning, and the difference in absorbance of the two beams is determined. Thus, the resulting spectrum represents the first derivative of the conventional type absorption spectrum. Tissues and cell fractions have been examined at room and liquid N2 temperature and chromophoric molecules such as the mitochondrial cytochromes and blood pigments have been detectable in low concentrations. In the case of isolated cellular components, the observed effects of substrates and inhibitors confirm similar studies by conventional spectrophotometry. The extension of the SSD concept to the microscopic level has permitted the study of the tissue compartmentalization and function of cytochromes and other pigments within layered tissue. PMID:4392452

  17. Dual specimens increase the diagnostic accuracy and reduce the reaction duration of rapid urease test

    PubMed Central

    Hsu, Wen-Hung; Wang, Sophie SW; Kuo, Chao-Hung; Chen, Chiao-Yun; Chang, Ching-Wen; Hu, Huang-Ming; Wang, Jaw-Yuan; Yang, Yuan-Chieh; Lin, Yu-Chun; Wang, Wen-Ming; Wu, Deng-Chyang; Wu, Ming-Tsang; Kuo, Fu-Chen

    2010-01-01

    AIM: To evaluate the influence of multiple samplings during esophagogastroduodenoscopy (EGD) on the accuracy of the rapid urease test, and the validity of newly developed rapid urease tests, HelicotecUT plus test and HelicotecUT test, CLO test and ProntoDry test. METHODS: A total of 355 patients undergoing EGD for dyspepsia were included. Their Helicobacter pylori (H. pylori) treatment status was either naïve or eradicated. Six biopsy specimens from antrum and gastric body, respectively, were obtained during EGD. Single antral specimens and dual (antrum + body) specimens were compared. Infection status of H. pylori was evaluated by three different tests: culture, histology, and four different commercially available rapid urease tests (RUTs)-including the newly developed HelicotecUT plus test and HelicotecUT test, and established CLO test and ProntoDry test. H. pylori status was defined as positive when the culture was positive or if there were concordant positive results among histology, CLO test and ProntoDry test. RESULTS: When dual specimens were applied, sensitivity was enhanced and RUT reaction time was significantly reduced, regardless of their treatment status. Thirty minutes were enough to achieve an agreeable positive rate in all the RUTs. Both newly developed RUTs showed comparable sensitivity, specificity and accuracy to the established RUTs, regardless of patient treatment status, RUT reaction duration, and EGD biopsy sites. CONCLUSION: Combination of antrum and body biopsy specimens greatly enhances the sensitivity of rapid urease test and reduces the reaction duration to 30 min. PMID:20556840

  18. Retrospective determination of ceruloplasmin in newborn screening blood spots of patients with Wilson disease.

    PubMed

    Kroll, Charles A; Ferber, Matt J; Dawson, Brian D; Jacobson, Robert M; Mensink, Kara A; Lorey, Fred; Sherwin, John; Cunningham, George; Rinaldo, Piero; Matern, Dietrich; Hahn, Si Houn

    2006-01-01

    Wilson disease is an autosomal recessive disorder of copper transport, caused by the reduced or absent function of the Wilson disease gene ATP7B on chromosome 13. The disease is characterized by reduced incorporation of copper into the ceruloplasmin protein and reduced excretion of copper into the bile. Wilson disease is effectively treated if detected early. Our study goals were to determine the feasibility of a population screening for Wilson disease using dried blood spots and to characterize the base-line ceruloplasmin concentration in newborn blood spots of patients with Wilson disease. Ceruloplasmin was analyzed in dried blood spots obtained from 353 Mayo Clinic pediatric volunteers aged from 3 months to 18 years and from 1045 anonymous newborn screening specimens using a sandwich enzyme-linked immunosorbent assay. The original newborn screening blood spots were retrieved from two patients with Wilson disease along with age-matched controls for ceruloplasmin determination. The mean (+/-SD) concentration of ceruloplasmin in the pediatric blood spots was 40.0+/-14.4 mg/dL (range 13.1 to >60 mg/dL) and newborn blood spots was 47.2+/-15.5mg/dL (range 6.5 to >60 mg/dL). Ceruloplasmin in the newborn blood spots from two Wilson disease patients were 2.6 and 2.8 mg/dL, respectively. The newborns affected with Wilson disease had significantly lower ceruloplasmin levels in blood spots than unaffected newborns. These findings support that presymptomatic screening for Wilson disease using dried blood spots could be possible, even in the newborn period.

  19. Donating Blood

    MedlinePlus

    ... blood transfusion. Blood donors — especially donors with certain blood types — are always in demand. Who Can Donate Blood? ... Natural Disasters: How to Help Blood Blood Transfusions Blood Types Contact Us Print Resources Send to a Friend ...

  20. 36 CFR 2.5 - Research specimens.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... superintendent approves a written research proposal and determines that the collection will benefit science or... 36 Parks, Forests, and Public Property 1 2011-07-01 2011-07-01 false Research specimens. 2.5... PROTECTION, PUBLIC USE AND RECREATION § 2.5 Research specimens. (a) Taking plants, fish, wildlife, rocks...

  1. 37 CFR 1.93 - Specimens.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2010-07-01 2010-07-01 false Specimens. 1.93 Section 1.93 Patents, Trademarks, and Copyrights UNITED STATES PATENT AND TRADEMARK OFFICE, DEPARTMENT OF COMMERCE....93 Specimens. When the invention relates to a composition of matter, the applicant may be required...

  2. 36 CFR 2.5 - Research specimens.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 1 2010-07-01 2010-07-01 false Research specimens. 2.5... PROTECTION, PUBLIC USE AND RECREATION § 2.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or... Federal agency for the purpose of research, baseline inventories, monitoring, impact analysis, group...

  3. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 36 Parks, Forests, and Public Property 3 2010-07-01 2010-07-01 false Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... of research, baseline inventories, monitoring, impact analysis, group study, or museum display...

  4. Layered Plating Specimens For Mechanical Tests

    NASA Technical Reports Server (NTRS)

    Thompson, Linda B.; Flowers, Cecil E.

    1991-01-01

    Layered specimens readily made in standard sizes for tensile and other tests of mechanical properties. Standard specimen of metal ordinarily difficult to plate to standard grip thickness or diameter made by augmentation with easier-to-plate material followed by machining to standard size and shape.

  5. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... Patents, Trademarks, and Copyrights UNITED STATES PATENT AND TRADEMARK OFFICE, DEPARTMENT OF COMMERCE... include one specimen per class showing the mark as used on or in connection with the goods or services... associated with the goods. (2) A service mark specimen must show the mark as actually used in the sale...

  6. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 3 2013-07-01 2012-07-01 true Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... of research, baseline inventories, monitoring, impact analysis, group study, or museum display...

  7. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 36 Parks, Forests, and Public Property 3 2012-07-01 2012-07-01 false Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... of research, baseline inventories, monitoring, impact analysis, group study, or museum display...

  8. 36 CFR 1002.5 - Research specimens.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 36 Parks, Forests, and Public Property 3 2011-07-01 2011-07-01 false Research specimens. 1002.5... RECREATION § 1002.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or minerals except in... of research, baseline inventories, monitoring, impact analysis, group study, or museum display...

  9. 36 CFR 2.5 - Research specimens.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 36 Parks, Forests, and Public Property 1 2013-07-01 2013-07-01 false Research specimens. 2.5... PROTECTION, PUBLIC USE AND RECREATION § 2.5 Research specimens. (a) Taking plants, fish, wildlife, rocks or... Federal agency for the purpose of research, baseline inventories, monitoring, impact analysis, group...

  10. Specimen origin, type and testing laboratory are linked to longer turnaround times for HIV viral load testing in Malawi

    PubMed Central

    Chipungu, Geoffrey; Kim, Andrea A.; Sarr, Abdoulaye; Ali, Hammad; Mwenda, Reuben; Nkengasong, John N.; Singer, Daniel

    2017-01-01

    Background Efforts to reach UNAIDS’ treatment and viral suppression targets have increased demand for viral load (VL) testing and strained existing laboratory networks, affecting turnaround time. Longer VL turnaround times delay both initiation of formal adherence counseling and switches to second-line therapy for persons failing treatment and contribute to poorer health outcomes. Methods We utilized descriptive statistics and logistic regression to analyze VL testing data collected in Malawi between January 2013 and March 2016. The primary outcomes assessed were greater-than-median pretest phase turnaround time (days elapsed from specimen collection to receipt at the laboratory) and greater-than-median test phase turnaround time (days from receipt to testing). Results The median number of days between specimen collection and testing increased 3-fold between 2013 (8 days, interquartile range (IQR) = 6–16) and 2015 (24, IQR = 13–39) (p<0.001). Multivariable analysis indicated that the odds of longer pretest phase turnaround time were significantly higher for specimen collection districts without laboratories capable of conducting viral load tests (adjusted odds ratio (aOR) = 5.16; 95% confidence interval (CI) = 5.04–5.27) as well as for Malawi’s Northern and Southern regions. Longer test phase turnaround time was significantly associated with use of dried blood spots instead of plasma (aOR = 2.30; 95% CI = 2.23–2.37) and for certain testing months and testing laboratories. Conclusion Increased turnaround time for VL testing appeared to be driven in part by categorical factors specific to the phase of turnaround time assessed. Given the implications of longer turnaround time and the global effort to scale up VL testing, addressing these factors via increasing efficiencies, improving quality management systems and generally strengthening the VL spectrum should be considered essential components of controlling the HIV epidemic. PMID:28235013

  11. Dry mouth during cancer treatment

    MedlinePlus

    Chemotherapy - dry mouth; Radiation therapy - dry mouth; Transplant - dry mouth; Transplantation - dry mouth ... National Cancer Institute. Chemotherapy and you: support for people with cancer. Updated May 2007. ... ...

  12. Improved liquid-liquid extraction with inter-well volume replacement dilution workflow and its application to quantify BMS-927711 in rat dried blood spots by UHPLC-MS/MS.

    PubMed

    Zheng, Naiyu; Zeng, Jianing; Ji, Qin C; Angeles, Aida; Buzescu, Adela; Basdeo, Shenita; Aubry, Anne-Françoise; Trouba, Kevin; Patrone, Laura M; Peng, Qianping; Arnold, Mark E

    2014-02-01

    An UHPLC-MS/MS method was developed and validated to quantify BMS-927711, a drug candidate to treat migraine, in rat dried blood spots (DBS). The DBS samples were extracted using an improved liquid-liquid extraction (LLE) strategy involving in the sonication of DBS punches in 20% MeOH aqueous solution containing the internal standard, [(13)C2, D4]-BMS-927711, and then with a 100mM NH4OAc buffer solution, followed by an automated LLE with EtOAc-hexane (70:30, v/v). The presence of 20% MeOH as an organic modifier in the elution solution significantly improved the analyte elution efficiency and assay performance. A novel inter-well volume replacement dilution workflow was introduced for DBS sample dilution before LLE step. This was a simple two-step process, firstly a small portion of the DBS blank solution was discarded, and then the same volume of a concentrated DBS sample solution was spiked into the leftover blank solution to achieve a desired dilution. Chromatographic separation was achieved on an Acuity UPLC(®) BEH C18 column (2.1mm×50mm, 1.7μm) and the analyte was detected by selected reaction monitoring (SRM) with positive electrospray ionization on an AB Sciex Triple Quad 5500 mass spectrometer. The standard curve was linear from 5.00 to 5000ng/mL with assay precision ≤4.9% CV, and assay accuracy within ±3.1%Dev of the nominal values. Accurate sample dilution was achieved by using inter-well volume replacement with a precision of ≤4.2% CV and an accuracy of ±3.3% for dilution QC at 50,000ng/mL with 100-fold dilution (n=18). This robust UHPLC-MS/MS assay has been successfully applied to the non-clinical studies in rats. By using inter-well volume replacement workflow, accurate dilution was demonstrated using only one DBS blank sample for a typical dilution of <50-fold, and using only two blank DBS samples for a dilution of up to 625-fold. Moreover, this new workflow makes it easier to automate DBS sample dilution.

  13. NEW TRANSPORT MEDIUM FOR SHIPMENT OF CLINICAL SPECIMENS. I. FECAL SPECIMENS.

    PubMed

    CARY, S G; BLAIR, E B

    1964-07-01

    Cary, Sylvia G. (Walter Reed Army Institute of Research, Washington, D.C.), and Eugene B Blair. New transport medium for shipment of clinical specimens. I. Fecal specimens. J. Bacteriol. 88:96-98. 1964.-A new transport medium for the collection and shipment of clinical specimens is described. Preliminary studies indicate that, with initial fecal specimens, Salmonella and Shigella can be recovered for as long as 49 days, Vibrio comma for 22 days, and Pasteurella pestis for at least 75 days.

  14. BIOMASS DRYING TECHNOLOGIES

    EPA Science Inventory

    The report examines the technologies used for drying of biomass and the energy requirements of biomass dryers. Biomass drying processes, drying methods, and the conventional types of dryers are surveyed generally. Drying methods and dryer studies using superheated steam as the d...

  15. Electron microprobe analysis program for biological specimens: BIOMAP

    NASA Technical Reports Server (NTRS)

    Edwards, B. F.

    1972-01-01

    BIOMAP is a Univac 1108 compatible program which facilitates the electron probe microanalysis of biological specimens. Input data are X-ray intensity data from biological samples, the X-ray intensity and composition data from a standard sample and the electron probe operating parameters. Outputs are estimates of the weight percentages of the analyzed elements, the distribution of these estimates for sets of red blood cells and the probabilities for correlation between elemental concentrations. An optional feature statistically estimates the X-ray intensity and residual background of a principal standard relative to a series of standards.

  16. Optimal design of biaxial tensile cruciform specimens

    NASA Astrophysics Data System (ADS)

    Demmerle, S.; Boehler, J. P.

    1993-01-01

    F OR EXPERIMENTAL investigations concerning the mechanical behaviour under biaxial stress states of rolled sheet metals, mostly cruciform flat specimens are used. By means of empirical methods, different specimen geometries have been proposed in the literature. In order to evaluate the suitability of a specimen design, a mathematically well defined criterion is developed, based on the standard deviations of the values of the stresses in the test section. Applied to the finite element method, the criterion is employed to realize the shape optimization of biaxial cruciform specimens for isotropic elastic materials. Furthermore, the performance of the obtained optimized specimen design is investigated in the case of off-axes tests on anisotropic materials. Therefore, for the first time, an original testing device, consisting of hinged fixtures with knife edges at each arm of the specimen, is applied to the biaxial test. The obtained results indicate the decisive superiority of the optimized specimens for the proper performance on isotropic materials, as well as the paramount importance of the proposed off-axes testing technique for biaxial tests on anisotropic materials.

  17. Comparative Evaluation of Dimensional Accuracy and Tensile Strength of a Type IV Gypsum Using Microwave and Air Drying Methods.

    PubMed

    Sharma, Anuraag; Shetty, Manoj; Hegde, Chethan; Shetty, N Sridhar; Prasad, D Krishna

    2013-12-01

    To evaluate dimensional accuracy and tensile strength of a type IV gypsum product, at different time intervals, dried in air or a microwave oven. Eighty specimens prepared from a cylindrical mould were used for measuring tensile strength (group A). Twenty specimens from a master die mould were used for determining dimensional accuracy (group B). In group A, 40 specimens were dried in open air at room temperature (A1). The other 40 were removed after 30 min and air dried for 20 min. These were subjected to microwave oven drying for 5 min (A2). Ten specimens each were tested under diametral compression at each of the following time periods: 1, 2, 4 and 24 h after drying. In group B, ten specimens were dried in open air at room temperature (B1). Ten specimens were removed from the mould after 30 min and air dried for 20 min. These were then dried in a microwave oven for 5 min (B2). The data was statistically analyzed using students unpaired "t" test. At all time intervals, diametral tensile strength (DTS) values for specimens dried in microwave oven were significantly higher than for those dried in open air. There were no significant differences between the dimensional accuracy of the two groups. In this study, microwave oven drying had a positive effect on the DTS of a type IV gypsum and the microwave oven dried specimens were as accurate as the air dried specimens over the same time period.

  18. Evaluation of specimen preparation techniques for micro-PIXE localisation of elements in hyperaccumulating plants

    NASA Astrophysics Data System (ADS)

    Kachenko, Anthony G.; Siegele, Rainer; Bhatia, Naveen P.; Singh, Balwant; Ionescu, Mihail

    2008-04-01

    Hybanthus floribundus subsp. floribundus, a rare Australian Ni-hyperaccumulating shrub and Pityrogramma calomelanos var. austroamericana, an Australian naturalized As-hyperaccumulating fern are promising species for use in phytoremediation of contaminated sites. Micro-proton-induced X-ray emission (μ-PIXE) spectroscopy was used to map the elemental distribution of the accumulated metal(loid)s, Ca and K in leaf or pinnule tissues of the two plant species. Samples were prepared by two contrasting specimen preparation techniques: freeze-substitution in tetrahydrofuran (THF) and freeze-drying. The specimens were analysed to compare the suitability of each technique in preserving (i) the spatial elemental distribution and (ii) the tissue structure of the specimens. Further, the μ-PIXE results were compared with concentration of elements in the bulk tissue obtained by ICP-AES analysis. In H. floribundus subsp. floribundus, μ-PIXE analysis revealed Ni, Ca and K concentrations in freeze-dried leaf tissues were at par with bulk tissue concentrations. Elemental distribution maps illustrated that Ni was preferentially localised in the adaxial epidermal tissues (1% DW) and least concentration was found in spongy mesophyll tissues (0.53% DW). Conversely, elemental distribution maps of THF freeze-substituted tissues indicated significantly lower Ni, Ca and K concentrations than freeze-dried specimens and bulk tissue concentrations. Moreover, Ni concentrations were uniform across the whole specimen and no localisation was observed. In P. calomelanos var. austroamericana freeze-dried pinnule tissues, μ-PIXE revealed statistically similar As, Ca and K concentrations as compared to bulk tissue concentrations. Elemental distribution maps showed that As localisation was relatively uniform across the whole specimen. Once again, THF freeze-substituted tissues revealed a significant loss of As compared to freeze-dried specimens and the concentrations obtained by bulk tissue analysis

  19. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... exceeding these size requirements (a “bulky specimen”), the Office will create a digital facsimile of the... cassette tape recording, CD-ROM, or other appropriate medium. (4) For a TEAS submission, the specimen...

  20. A Live Specimen Cell for the Microscope.

    ERIC Educational Resources Information Center

    McNeil, D. W.

    1991-01-01

    Provides background and instructions for the assembly of a microaquarium, or specimen cell, in which the dynamic world of living microorganisms can be viewed through a microscope overextended periods of time utilizing the simplest of materials in the process. (JJK)

  1. Urine Specimens: Tips to Help Children

    MedlinePlus

    ... Tests Online. AACC is a not-for-profit organization and does not endorse non-AACC products and services. Advertising & Sponsorship: Policy | ... Stool | Throat Culture Urine Specimens Children sometimes balk at the idea ...

  2. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... goods or the sale of the goods when it is impracticable to place the mark on the goods, packaging for... cassette tape recording, CD-ROM, or other appropriate medium. (4) For a TEAS submission, the specimen...

  3. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... goods when it is impracticable to place the mark on the goods, packaging for the goods, or displays... appropriate medium. (4) For a TEAS submission, the specimen must be a digitized image in .jpg or .pdf format....

  4. Design and testing of small composite specimens

    NASA Technical Reports Server (NTRS)

    Sharma, A. V.

    1981-01-01

    The effect of specimen size on the buckling strains of laminates subjected to low velocity projectile impact was investigated. The fiber composite selected was T300/5208 graphite/epoxy system. The quasi-isotropic laminates tested had 16 and 32 plies. The results were compared with those of a 48-ply laminate. Specimens of three different lengths with length to width aspect ratios of 1, 1.5, and 2 were also studied. The results show that (1) the specimen length does not have any significant influence on the buckling strains at failure caused by the projectile impact, and (2) the influence of specimen thickness on the strains at failure would decrease as the velocity of the impacting projectile increases.

  5. National Aeronautics and Space Administration Biological Specimen Repository

    NASA Technical Reports Server (NTRS)

    McMonigal, Kathleen A.; Pietrzyk, Robert a.; Johnson, Mary Anne

    2008-01-01

    The National Aeronautics and Space Administration Biological Specimen Repository (Repository) is a storage bank that is used to maintain biological specimens over extended periods of time and under well-controlled conditions. Samples from the International Space Station (ISS), including blood and urine, will be collected, processed and archived during the preflight, inflight and postflight phases of ISS missions. This investigation has been developed to archive biosamples for use as a resource for future space flight related research. The International Space Station (ISS) provides a platform to investigate the effects of microgravity on human physiology prior to lunar and exploration class missions. The storage of crewmember samples from many different ISS flights in a single repository will be a valuable resource with which researchers can study space flight related changes and investigate physiological markers. The development of the National Aeronautics and Space Administration Biological Specimen Repository will allow for the collection, processing, storage, maintenance, and ethical distribution of biosamples to meet goals of scientific and programmatic relevance to the space program. Archiving of the biosamples will provide future research opportunities including investigating patterns of physiological changes, analysis of components unknown at this time or analyses performed by new methodologies.

  6. Specimen banks for cancer prognostic factor research.

    PubMed

    Burke, H B; Henson, D E

    1998-10-01

    Prognostic factors are necessary for determining whether a patient will require therapy, for selecting the optimal therapy, and for evaluating the effectiveness of the therapy chosen. Research in prognostic factors has been hampered by long waiting times and a paucity of outcomes. Specimen banks can solve these problems, but their implementation and use give rise to many important and complex issues. This paper presents an overview of some of the issues related to the use of specimen banks in prognostic factor research.

  7. STRESS CORROSION CRACKING IN TEAR DROP SPECIMENS

    SciTech Connect

    Lam, P; Philip Zapp, P; Jonathan Duffey, J; Kerry Dunn, K

    2009-05-01

    Laboratory tests were conducted to investigate the stress corrosion cracking (SCC) of 304L stainless steel used to construct the containment vessels for the storage of plutonium-bearing materials. The tear drop corrosion specimens each with an autogenous weld in the center were placed in contact with moist plutonium oxide and chloride salt mixtures. Cracking was found in two of the specimens in the heat affected zone (HAZ) at the apex area. Finite element analysis was performed to simulate the specimen fabrication for determining the internal stress which caused SCC to occur. It was found that the tensile stress at the crack initiation site was about 30% lower than the highest stress which had been shifted to the shoulders of the specimen due to the specimen fabrication process. This finding appears to indicate that the SCC initiation took place in favor of the possibly weaker weld/base metal interface at a sufficiently high level of background stress. The base material, even subject to a higher tensile stress, was not cracked. The relieving of tensile stress due to SCC initiation and growth in the HAZ and the weld might have foreclosed the potential for cracking at the specimen shoulders where higher stress was found.

  8. Use of a compact sandwich specimen to evaluate fracture toughness and interfacial bonding of bone.

    PubMed

    Wang, X; Lankford, J; Agrawal, C M

    1994-01-01

    The objective of the present study was to develop a reliable and statistically valid test to measure the fracture toughness of small specimens of bone, and by extension, prosthetic materials, using a compact sandwich specimen. Samples of bone were sandwiched between holders of a different material and using this specimen configuration a new technique was developed to test the fracture toughness of the bone interlayer. The effects of different specimens sizes and holder materials were investigated empirically. Using finite element analysis a correction factor was determined to account for the finite thickness of the interlayer and the analytical solutions governing the test specimen were accordingly modified. Bulk compact tension specimens of bone were tested for comparison. Both wet and dry bone were evaluated and the fracture surface morphology characterized using scanning electron microscopy. The results indicate no statistically significant differences between the fracture toughness values obtained from the compact tension and sandwich specimens. The application of this technique to the testing of interfacial bonding between bone and biomaterials is discussed.

  9. A curved beam test specimen for determining the interlaminar tensile strength of a laminated composite

    NASA Technical Reports Server (NTRS)

    Hiel, Clement C.; Sumich, Mark; Chappell, David P.

    1990-01-01

    A curved beam type of test specimen is evaluated for use in determining the through-the-thickness strength of laminated composites. Two variations of a curved beam specimen configuration (semi-circular and elliptical) were tested to failure using static and fatigue loads. The static failure load for the semi-circular specimens was found to be highly sensitive to flaw content, with the specimens falling into two distinct groups. This result supports the use of proof testing for structural validation. Static design allowables are derived based on the Weibull distribution. Fatigue data indicates no measured increase in specimen compliance prior to final fracture. All static and fatigue failures at room temperature dry conditions occurred catastrophically. The elliptical specimens demonstrated unusually high failure strengths indicating the presence of phenomena requiring further study. Results are also included for specimens exposed to a wet environment showing a matrix strength degradation due to moisture content. Further testing is under way to evaluate a fatigue methodology for matrix dominated failures based on residual static strength (wearout).

  10. A curved beam test specimen for determining the interlaminar tensile strength of a laminated composite

    NASA Technical Reports Server (NTRS)

    Hiel, Clement C.; Sumich, Mark; Chappell, David P.

    1991-01-01

    A curved beam type of test specimen is evaluated for use in determining the through-the-thickness strength of laminated composites. Two variations of a curved beam specimen configuration (semicircular and elliptical) were tested to failure using static and fatigue loads. The static failure load for the semicircular specimens was found to be highly sensitive to flaw content, with the specimens falling into two distinct groups. This result supports the use of proof testing for structural validation. Static design allowables are derived based on the Weibull distribution. Fatigue data indicates no measured increase in specimen compliance prior to final fracture. All static and fatigue failures at room temperature dry conditions occurred catastrophically. The elliptical specimens demonstrated unusually high failure strengths indicating the presence of phenomena requiring further study. Results are also included for specimens exposed to a wet environment showing a matrix strength degradation due to moisture content. Further testing is underway to evaluate a fatigue methodology for matrix dominated failures based on residual static strength (wearout).

  11. Solution Preserves Nucleic Acids in Body-Fluid Specimens

    NASA Technical Reports Server (NTRS)

    Pierson, Duane L.; Stowe, Raymond P.

    2004-01-01

    A solution has been formulated to preserve deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) in specimens of blood, saliva, and other bodily fluids. Specimens of this type are collected for diagnostic molecular pathology, which is becoming the method of choice for diagnosis of many diseases. The solution makes it possible to store such specimens at room temperature, without risk of decomposition, for subsequent analysis in a laboratory that could be remote from the sampling location. Thus, the solution could be a means to bring the benefits of diagnostic molecular pathology to geographic regions where refrigeration equipment and diagnostic laboratories are not available. The table lists the ingredients of the solution. The functions of the ingredients are the following: EDTA chelates divalent cations that are necessary cofactors for nuclease activity. In so doing, it functionally removes these cations and thereby retards the action of nucleases. EDTA also stabilizes the DNA helix. Tris serves as a buffering agent, which is needed because minor contaminants in an unbuffered solution can exert pronounced effects on pH and thereby cause spontaneous degradation of DNA. SDS is an ionic detergent that inhibits ribonuclease activity. SDS has been used in some lysis buffers and as a storage buffer for RNA after purification. The use of the solution is straightforward. For example, a sample of saliva is collected by placing a cotton roll around in the subject's mouth until it becomes saturated, then the cotton is placed in a collection tube. Next, 1.5 mL of the solution are injected directly into the cotton and the tube is capped for storage at room temperature. The effectiveness of the solution has been demonstrated in tests on specimens of saliva containing herpes simplex virus. In the tests, the viral DNA, as amplified by polymerase chain reaction, was detected even after storage for 120 days.

  12. A comparative study of blood warmer performance.

    PubMed

    Uhl, L; Pacini, D; Kruskall, M S

    1992-11-01

    Massive transfusions of refrigerator-temperature blood may induce hypothermia and life-threatening arrhythmias; for this reason a variety of devices have been developed for rapid blood warming. Blood warmers available in the United States use one of three warming technologies: dry heat, water bath, or countercurrent heat exchange. In the current study we evaluated blood warmers representative of each technology for speed and extent of heat transfer: the Fenwal blood warmer (Fenwal Laboratories; dry heat), the DW-1000 (American Pharmaseal Co.; dry heat), the FloTem IIe (DataChem Inc.; dry heat), the Hemokinetitherm (Dupaco Inc.; water bath), and the H250 and H500 (Level 1 Technologies; countercurrent heat exchange). Only one countercurrent heat instrument (the H500) was able to heat blood > or = 33 degrees C at target flow rates > or = 250 ml/min. Dry heat and water bath blood warmers were unable to warm blood > or = 33 degrees C at target flow rates > or = 100 ml/min. High resistance to flow with the proprietary tubing required for one instrument (the Hemokinetitherm) prevented tests of blood warming at rates > 150 ml/min. We found that instruments that used countercurrent technology warmed blood and saline more effectively than did blood warmers that used either dry heat or water bath technology. Our study also demonstrated the need for close control and standardization of experimental conditions in the evaluation of blood warming devices.

  13. Closeout of JOYO-1 Specimen Fabrication Efforts

    SciTech Connect

    ME Petrichek; JL Bump; RF Luther

    2005-10-31

    Fabrication was well under way for the JOYO biaxial creep and tensile specimens when the NR Space program was canceled. Tubes of FS-85, ASTAR-811C, and T-111 for biaxial creep specimens had been drawn at True Tube (Paso Robles, CA), while tubes of Mo-47.5 Re were being drawn at Rhenium Alloys (Cleveland, OH). The Mo-47.5 Re tubes are now approximately 95% complete. Their fabrication and the quantities produced will be documented at a later date. End cap material for FS-85, ASTAR-811C, and T-111 had been swaged at Pittsburgh Materials Technology, Inc. (PMTI) (Large, PA) and machined at Vangura (Clairton, PA). Cutting of tubes, pickling, annealing, and laser engraving were in process at PMTI. Several biaxial creep specimen sets of FS-85, ASTAR-811C, and T-111 had already been sent to Pacific Northwest National Laboratory (PNNL) for weld development. In addition, tensile specimens of FS-85, ASTAR-811C, T-111, and Mo-47.5 Re had been machined at Kin-Tech (North Huntington, PA). Actual machining of the other specimen types had not been initiated. Flowcharts 1-3 detail the major processing steps each piece of material has experienced. A more detailed description of processing will be provided in a separate document [B-MT(SRME)-51]. Table 1 lists the in-process materials and finished specimens. Also included are current metallurgical condition of these materials and specimens. The available chemical analyses for these alloys at various points in the process are provided in Table 2.

  14. [Disparity between cytohistopathological results of diagnostic specimens and the resected tumor piece in bronchopulmonary carcinoma].

    PubMed

    Sánchez Varilla, J M; Benítez Moya, J M; Baena Luna, M; Loscertales Abril, J; García Díaz, F; Vázquez Valiente, M; Villa Arellano, F; Márquez Varela, F; Martínez Puentes, V

    1996-04-01

    In 88 patients diagnosed and intervened of lung carcinoma, we evaluate the security of the Diagnostic method--cytology: sputum, brushed bronchial, bronchial aspiration, puncture pulmonary aspiration and histology of the bronchial biopsy--when we correlate them with the histology of the dried up tumour. From the 88 patient, in 11 (12.5%) there was a disparity between some specimen and the piece tumour. If we analyze the specimen: the bronchial biopsy, was different in 3 cases, (4.8%); the brushed and the bronchial biopsy, aspiration in 2, (2.7% and 2.3% respectively); the puncture pulmonary aspiration in 6, (26%) and the cytology of sputum in 2 cases, (2.4%). In 8 patients (9%), the outputs of some specimen were not conclusive, if we remove these we find that: the bronchial biopsy was not conclusive in 2 cases, (3.2%), the brushed bronchial in 5, (6.8%); the bronchial aspiration in 9 (10.2%) and the cytology of sputum in 2 cases, (2.4%). With this data we could infer that sometimes the positive outputs of the specimen have no correlation with the histology of the tumour dried up. This disparity has not been observed in the oat-cells. Therefore when we send the specimens to the pathologist, it would always be advisable for him to inform us if there are any different cells belonging to the principal group of the tumor.

  15. Dry eye syndrome

    MedlinePlus

    ... of dry eyes include: Dry environment or workplace (wind, air conditioning) Sun exposure Smoking or second-hand ... NOT smoke and avoid second-hand smoke, direct wind, and air conditioning. Use a humidifier, especially in ...

  16. Characterization of some biological specimens using TEM and SEM

    NASA Astrophysics Data System (ADS)

    Ghosh, Nabarun; Smith, Don W.

    2009-05-01

    The advent of novel techniques using the Transmission and Scanning Electron Microscopes improved observation on various biological specimens to characterize them. We studied some biological specimens using Transmission and Scanning Electron Microscopes. We followed negative staining technique with Phosphotungstic acid using bacterial culture of Bacillus subtilis. Negative staining is very convenient technique to view the structural morphology of different samples including bacteria, phage viruses and filaments in a cell. We could observe the bacterial cell wall and flagellum very well when trapped the negative stained biofilm from bacterial culture on a TEM grid. We cut ultra thin sections from the fixed root tips of Pisum sativum (Garden pea). Root tips were pre fixed with osmium tetroxide and post fixed with uranium acetate and placed in the BEEM capsule for block making. The ultrathin sections on the grid under TEM showed the granular chromatin in the nucleus. The protein bodies and large vacuoles with the storage materials were conspicuous. We followed fixation, critical point drying and sputter coating with gold to view the tissues with SEM after placing on stubs. SEM view of the leaf surface of a dangerous weed Tragia hispida showed the surface trichomes. These trichomes when break on touching releases poisonous content causing skin irritation. The cultured tissue from in vitro culture of Albizia lebbeck, a tree revealed the regenerative structures including leaf buds and stomata on the tissue surface. SEM and TEM allow investigating the minute details characteristic morphological features that can be used for classroom teaching.

  17. Investigations in space-related molecular biology. [cryo-electron microscopic and diffraction studies on terrestrial and extraterrestrial specimens

    NASA Technical Reports Server (NTRS)

    Fernandez-Moran, H.; Pritzker, A. N.

    1974-01-01

    Improved instrumentation and preparation techniques for high resolution, high voltage cryo-electron microscopic and diffraction studies on terrestrial and extraterrestrial specimens are reported. Computer correlated ultrastructural and biochemical work on hydrated and dried cell membranes and related biological systems provided information on membrane organization, ice crystal formation and ordered water, RNA virus linked to cancer, lunar rock samples, and organometallic superconducting compounds. Apollo 11, 12, 14, and 15 specimens were analyzed

  18. Evaluation of transport and storage techniques for isolation of Campylobacter fetus subsp. jejuni from turkey cecal specimens.

    PubMed

    Luechtefeld, N W; Wang, W L; Blaser, M J; Reller, L B

    1981-03-01

    Immediate culturing of fecal specimens is not always possible, and appropriate methods for transport and storage of Campylobacter fetus subsp. jejuni specimens have not been fully evaluated. Using nine techniques, we studied the survival of C. fetus subsp. jejuni in cecal specimens from infected turkeys. The organisms survived in specimens held without transport medium for 3 to 15 days (median, 9 days) at 4 degrees C, and 2 to 9 days (median, 4 days) at 25 degrees C. Only 20% of specimens frozen for 24 h at either -20 or -70 degrees C yielded C. fetus subsp. jejuni. Specimens dried on filter paper strips were negative for C. fetus subsp. jejuni within 1.5 h. Cary-Blair medium with decreased agar was the best of the six transport media tested, it enabled recovery of the organism from 100% (3 days) and 71% (7 days) of cecal samples held at 4 degrees C and 94% (3 days) and 85% (7 days) of cecal specimens held at 25 degrees C. In contrast, more than half of all cecal specimens held at 4 or 25 degrees C in Culturettes or buffered glycerol saline were negative by 3 days, and all were negative at 7 days. Results with the other three media studied (Campy-thio, thioglycolate medium, and alkaline peptone water) were intermediate. Overnight incubation of specimens in alkaline peptone water at 37 or 42 degrees C did not enhance recovery of C. fetus subsp. jejuni. Therefore, refrigeration without a transport medium is satisfactory for up to 3 days for recovery of C. fetus subsp. jejuni from specimens, however, we recommend the use of Cary-Blair medium with decreased agar for specimens that must be transported or stored for longer than 3 days and for rectal swabs, to prevent drying.

  19. Biological monitoring of occupational exposure to toxic chemicals. Collection, processing, and storage of specimens

    SciTech Connect

    Aitio, A.; Jaervisalo, J.

    1985-03-01

    Exposure to at least 100 different chemicals may be estimated on an individual basis from their concentrations in blood or urine. The present document reviews sources of error in the collection, processing and storage of specimens for this biological monitoring. Physiological factors cause variation in the concentration of chemicals in the body fluids. Distribution of water depends on posture. Exercise and meals cause changes in blood constituents. The urine output varies and, thus, the concentrations of dissolved chemicals change. Many toxic chemicals show short half times in the blood; thus, their concentrations depend on the timing of the specimen collection. Skin absorption may result in dramatically different chemical concentrations in different parts of the circulation. The stability of chemicals in the collected specimens is generally limited: chemical deterioration, adsorption, precipitation, and evaporation are the main causes of losses. For many chemicals, especially for trace elements, contamination of the specimen is the overwhelmingly most important source of error. As the range of the chemicals measured is wide, the relative importance of the sources of error is different for different chemicals. Information on most chemicals is at present very limited. Thus, before commencing a program on biological exposure monitoring, it is advisable to search the optimal conditions for specimen collection, processing, and storage.

  20. Genomic treasure troves: complete genome sequencing of herbarium and insect museum specimens.

    PubMed

    Staats, Martijn; Erkens, Roy H J; van de Vossenberg, Bart; Wieringa, Jan J; Kraaijeveld, Ken; Stielow, Benjamin; Geml, József; Richardson, James E; Bakker, Freek T

    2013-01-01

    Unlocking the vast genomic diversity stored in natural history collections would create unprecedented opportunities for genome-scale evolutionary, phylogenetic, domestication and population genomic studies. Many researchers have been discouraged from using historical specimens in molecular studies because of both generally limited success of DNA extraction and the challenges associated with PCR-amplifying highly degraded DNA. In today's next-generation sequencing (NGS) world, opportunities and prospects for historical DNA have changed dramatically, as most NGS methods are actually designed for taking short fragmented DNA molecules as templates. Here we show that using a standard multiplex and paired-end Illumina sequencing approach, genome-scale sequence data can be generated reliably from dry-preserved plant, fungal and insect specimens collected up to 115 years ago, and with minimal destructive sampling. Using a reference-based assembly approach, we were able to produce the entire nuclear genome of a 43-year-old Arabidopsis thaliana (Brassicaceae) herbarium specimen with high and uniform sequence coverage. Nuclear genome sequences of three fungal specimens of 22-82 years of age (Agaricus bisporus, Laccaria bicolor, Pleurotus ostreatus) were generated with 81.4-97.9% exome coverage. Complete organellar genome sequences were assembled for all specimens. Using de novo assembly we retrieved between 16.2-71.0% of coding sequence regions, and hence remain somewhat cautious about prospects for de novo genome assembly from historical specimens. Non-target sequence contaminations were observed in 2 of our insect museum specimens. We anticipate that future museum genomics projects will perhaps not generate entire genome sequences in all cases (our specimens contained relatively small and low-complexity genomes), but at least generating vital comparative genomic data for testing (phylo)genetic, demographic and genetic hypotheses, that become increasingly more horizontal

  1. Surface analysis of space telescope material specimens

    NASA Technical Reports Server (NTRS)

    Fromhold, A. T.; Daneshvar, K.

    1985-01-01

    Qualitative and quantitative data on Space Telescope materials which were exposed to low Earth orbital atomic oxygen in a controlled experiment during the 41-G (STS-17) mission were obtained utilizing the experimental techniques of Rutherford backscattering (RBS), particle induced X-ray emission (PIXE), and ellipsometry (ELL). The techniques employed were chosen with a view towards appropriateness for the sample in question, after consultation with NASA scientific personnel who provided the material specimens. A group of eight samples and their controls selected by NASA scientists were measured before and after flight. Information reported herein include specimen surface characterization by ellipsometry techniques, a determination of the thickness of the evaporated metal specimens by RBS, and a determination of trace impurity species present on and within the surface by PIXE.

  2. Blood pressure

    MedlinePlus Videos and Cool Tools

    Normal blood pressure is important for proper blood flow to the body's organs and tissues. The force of the blood on the walls of the arteries is called blood pressure. Blood pressure is measured both as the heart ...

  3. Dry deposition velocities

    SciTech Connect

    Sehmel, G.A.

    1984-03-01

    Dry deposition velocities are very difficult to predict accurately. In this article, reported values of dry deposition velocities are summarized. This summary includes values from the literature on field measurements of gas and particle dry deposition velocities, and the uncertainties inherent in extrapolating field results to predict dry deposition velocities are discussed. A new method is described for predicting dry deposition velocity using a least-squares correlation of surface mass transfer resistances evaluated in wind tunnel experiments. 14 references, 4 figures, 1 table.

  4. Structure of drying costs

    SciTech Connect

    Sztabert, Z.T.

    1996-05-01

    A knowledge of cost structure and cost behavior is necessary in the management activities, particularly in the domain of investment or production decision making, as well as in the areas of production cost planning and control. Prediction and analysis of values of cost components for different technologies of drying are important when selection of a drying method and drying equipment should be done. Cost structures of lumber and coal drying processes together with an application of the factor method for prediction of the drying cost are presented.

  5. Fracture of Composite Compact Tension Specimens

    DTIC Science & Technology

    1975-01-01

    E: lb/in.; X 10* M.: Fiber Volume, % 1002 S- glass /epoxy Unidirectional Crossply 6.9 4.7 2.3 4.7 1.0 1.1 0.28 0.14 55 MOD 1-5208...configuration used in most of the fracture experiments is shown in Fig. 1. In unidirectional S- glass /epoxy specimens the fiber direction with respect to...conducted only with 0° or 90° fiber orientation. Cross-ply specimens of both S- glass and graphite were tested with the outer plies oriented at 0°, 45

  6. Stress intensity factor in a tapered specimen

    NASA Technical Reports Server (NTRS)

    Xue-Hui, L.; Erdogan, F.

    1985-01-01

    The general problem of a tapered specimen containing an edge crack is formulated in terms of a system of singular integral equations. The equations are solved and the stress intensity factor is calculated for a compact and for a slender tapered specimen, the latter simulating the double cantilever beam. The results are obtained primarily for a pair of concentrated forces and for crack surface wedge forces. The stress intensity factors are also obtained for a long strip under uniform tension which contains inclined edge cracks.

  7. How to submit a nail specimen.

    PubMed

    Reinig, Erica; Rich, Phoebe; Thompson, Curtis T

    2015-04-01

    The scarcity of specific submission protocols for nail unit biopsies presents many challenges for appropriate specimen processing. Many nail biopsies are received fragmented or without orientation, often resulting in less-than-ideal tissue embedding and poor histologic sections, which are difficult to interpret. Methods are described for proper nail matrix/bed biopsy and plate submission that incorporate aspects of previous submission protocols and include inking the biopsy specimen along with submitting the tissue on a drawing of the nail. Also described is a technique for maintaining adherence of nail plate to glass slides, a chronic challenge in the laboratory.

  8. Recovery and susceptibility testing of Mycobacterium tuberculosis from extrapulmonary specimens by the BACTEC radiometric method.

    PubMed Central

    Fadda, G; Roe, S L

    1984-01-01

    This study was carried out to evaluate the sensitivity and rapidity of the BACTEC radiometric techniques for isolation and susceptibility testing of mycobacteria from extrapulmonary specimens. Concentrated specimens of urine, pleural fluid, and blood as well as other extrapulmonary specimens were processed for the recovery of mycobacteria and for drug susceptibility testing, employing conventional and BACTEC radiometric methods. Out of 483 specimens processed, 20 were found to be positive for Mycobacterium tuberculosis on the conventional Lowenstein -Jensen medium, and 19 were found to be positive in the BACTEC 7H12 medium. Average recovery times were 22.5 days for the conventional method and 10.9 days for the BACTEC method. When isolated cultures were tested for susceptibility to streptomycin, isoniazid, rifampin, and ethambutol, results were reported at an average time of 22 and 5.4 days for the conventional and BACTEC methods, respectively, with good correlation. PMID:6429193

  9. Marking planes of surgical excision on breast biopsy specimens: use of artists' pigments suspended in acetone.

    PubMed Central

    Paterson, D A; Davies, J D

    1988-01-01

    The performance of carbon and metallic inks, silver nitrate solution, and artists' pigments mounted in acetone was compared for marking the surface of surgical biopsy specimens. Using India ink is an unsatisfactory procedure because of slow drying, messiness, and spreading of the ink. It is concluded that use of artists' pigments has many advantages over other reagents, because of their rapid drying, resistance to tissue processing, and the ability to mark simultaneously many different planes of excision. Furthermore, the pigments are readily visible, are distinguishable from each other on microscopical examination, and the method entails little extra cost. Images Fig 1 Fig 2 PMID:3056982

  10. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 16 Commercial Practices 2 2013-01-01 2013-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  11. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 16 Commercial Practices 2 2014-01-01 2014-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  12. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 16 Commercial Practices 2 2012-01-01 2012-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  13. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 16 Commercial Practices 2 2011-01-01 2011-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  14. 10 CFR 26.165 - Testing split specimens and retesting single specimens.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... confirmatory creatinine and confirmatory specific gravity tests, when retesting an aliquot of a single specimen or testing Bottle B of a split specimen, to reconfirm that the creatinine concentration was less than.... The second laboratory may only conduct the confirmatory creatinine and specific gravity tests...

  15. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 16 Commercial Practices 2 2010-01-01 2010-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt.1610, Fig. 3 Figure 3 to Part...

  16. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 16 Commercial Practices 2 2011-01-01 2011-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt.1610, Fig. 3 Figure 3 to Part...

  17. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 16 Commercial Practices 2 2013-01-01 2013-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt. 1610, Fig. 3 Figure 3 to Part...

  18. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 16 Commercial Practices 2 2012-01-01 2012-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt.1610, Fig. 3 Figure 3 to Part...

  19. 16 CFR Figure 3 to Part 1610 - Specimen Holder Supported in Specimen Rack

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 16 Commercial Practices 2 2014-01-01 2014-01-01 false Specimen Holder Supported in Specimen Rack 3 Figure 3 to Part 1610 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION FLAMMABLE FABRICS ACT REGULATIONS STANDARD FOR THE FLAMMABILITY OF CLOTHING TEXTILES Pt. 1610, Fig. 3 Figure 3 to Part...

  20. 16 CFR Figure 6 to Subpart A of... - Dummy Specimen in Specimen Holder

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 16 Commercial Practices 2 2010-01-01 2010-01-01 false Dummy Specimen in Specimen Holder 6 Figure 6 to Subpart A of Part 1209 Commercial Practices CONSUMER PRODUCT SAFETY COMMISSION CONSUMER PRODUCT SAFETY ACT REGULATIONS INTERIM SAFETY STANDARD FOR CELLULOSE INSULATION The Standard Pt. 1209, Subpt....

  1. Strain controlled cyclic tests on miniaturized specimens

    NASA Astrophysics Data System (ADS)

    Procházka, R.; Džugan, J.

    2017-02-01

    The paper is dealing with strain controlled cyclic tests using a non-contact strain measurement based on digital image correlation techniques on proportional sizes of conventional specimens. The cyclic behaviour of 34CrNiMo6 high-strength steel was investigated on miniaturized round specimens with diameter of 2mm that were compared with specimens in accordance with ASTM E606 standards. The cycle asymmetry coefficient was R= -1. This application is intended to be used for life time assessment of in service components in future work which enables to carried out a group of mechanical tests from a limited amount of the experimental material. The attention was paid to confirm the suitability of the proposed size miniaturization geometry, testing set up and procedure. The test results obtained enabled to construct Manson-Coffin curves and assess fatigue parameters. The purpose of this study is to present differences between cyclic curves and cyclic parameters which have been evaluated based on conventional and miniaturized specimens.

  2. 7 CFR 97.8 - Specimen requirements.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 3 2010-01-01 2010-01-01 false Specimen requirements. 97.8 Section 97.8 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE (CONTINUED) COMMODITY LABORATORY TESTING PROGRAMS PLANT VARIETY AND PROTECTION...

  3. Archaeopteryx: notice of a "new" specimen.

    PubMed

    Ostrom, J H

    1970-10-30

    A fourth specimen of Archaeopteryx (cf. lithographica), the oldest known fossil bird, was recently found in the collections of the Teyler Museum in the Netherlands. Unique preservation of the horny sheaths of the manus claws provides new evidence that may be relevant to the question of the origins of avian flight. Tentative interpretation suggests a cursorial rather than arboreal origin.

  4. Vee-notch tool cuts specimens

    NASA Technical Reports Server (NTRS)

    Spier, R. A.

    1970-01-01

    Triangular cutting tool uses carbide tips for notching heat-treated or abrasive materials, and alloys subjected to high structural stresses. The tool is rigidly mounted in a slot of mating contour to prevent deflection during cutting of tensile specimens. No other expensive machine equipment is required.

  5. 37 CFR 2.56 - Specimens.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... advertising of the services. (3) A collective trademark or collective service mark specimen must show how a member uses the mark on the member's goods or in the sale or advertising of the member's services. (4) A... as actually used on or in connection with the goods, or in the sale or advertising of the...

  6. Structure of wet specimens in electron microscopy. Improved environmental chambers make it possible to examine wet specimens easily.

    PubMed

    Parsons, D F

    1974-11-01

    Several recent technological advances have increased the practicality and usefulness of the technique of electron microscopy of wet objects. (i) There have been gains in the effective penetration of high-voltage microscopes, scanning transmission microscopes, and high-voltage scanning microscopes. The extra effective penetration gives more scope for obtaining good images through film windows, gas, and liquid layers. (ii) Improved methods of obtaining contrast are available (especially dark field and inelastic filtering) that often make it possible to obtain sufficient contrast with wet unstained objects. (iii) Improved environmental chamber design makes it possible to insert and examine wet specimens as easily as dry specimens. The ultimate achievable resolution for wet objects in an environmental chamber will gradually become clear experimentally. Resolution is mainly a function of gas path, liquid and wet specimen thickness, specimen stage stability, acceleration voltage, and image mode (fixed or scanning beam) (13). Much depends on the development of the technique for controlling the thickness of extraneous water film around wet objects or the technique for depositing wet objects onto dry, hydrophobic support films. Although some loss of resolution due to water or gas scattering will always occur, an effective gain is anticipated in preserving the shape of individual molecules and preventing the partial collapse that usually occurs on drying or negative staining. The most basic question for biological electron microscopy is probably whether any living functions of cells can be observed so that the capabilities of the phase contrast and interference light microscopes can be extended. Investigators are now rapidly approaching a final answer to this question. The two limiting factors are (i) maintaining cell motility in spread cells immersed in thin layers of media and (ii) reducing beam radiation damage to an acceptable level. The use of sensitive emulsions and

  7. Simultaneous specimen and stage cleaning device for analytical electron microscope

    DOEpatents

    Zaluzec, Nestor J.

    1996-01-01

    An improved method and apparatus are provided for cleaning both a specimen stage, a specimen and an interior of an analytical electron microscope (AEM). The apparatus for cleaning a specimen stage and specimen comprising a plasma chamber for containing a gas plasma and an air lock coupled to the plasma chamber for permitting passage of the specimen stage and specimen into the plasma chamber and maintaining an airtight chamber. The specimen stage and specimen are subjected to a reactive plasma gas that is either DC or RF excited. The apparatus can be mounted on the analytical electron microscope (AEM) for cleaning the interior of the microscope.

  8. Ambient Dried Aerogels

    NASA Technical Reports Server (NTRS)

    Jones, Steven M.; Paik, Jong-Ah

    2013-01-01

    A method has been developed for creating aerogel using normal pressure and ambient temperatures. All spacecraft, satellites, and landers require the use of thermal insulation due to the extreme environments encountered in space and on extraterrestrial bodies. Ambient dried aerogels introduce the possibility of using aerogel as thermal insulation in a wide variety of instances where supercritically dried aerogels cannot be used. More specifically, thermoelectric devices can use ambient dried aerogel, where the advantages are in situ production using the cast-in ability of an aerogel. Previously, aerogels required supercritical conditions (high temperature and high pressure) to be dried. Ambient dried aerogels can be dried at room temperature and pressure. This allows many materials, such as plastics and certain metal alloys that cannot survive supercritical conditions, to be directly immersed in liquid aerogel precursor and then encapsulated in the final, dried aerogel. Additionally, the metalized Mylar films that could not survive the previous methods of making aerogels can survive the ambient drying technique, thus making multilayer insulation (MLI) materials possible. This results in lighter insulation material as well. Because this innovation does not require high-temperature or high-pressure drying, ambient dried aerogels are much less expensive to produce. The equipment needed to conduct supercritical drying costs many tens of thousands of dollars, and has associated running expenses for power, pressurized gasses, and maintenance. The ambient drying process also expands the size of the pieces of aerogel that can be made because a high-temperature, high-pressure system typically has internal dimensions of up to 30 cm in diameter and 60 cm in height. In the case of this innovation, the only limitation on the size of the aerogels produced would be in the ability of the solvent in the wet gel to escape from the gel network.

  9. Dry Snow Metamorphism

    DTIC Science & Technology

    2012-09-19

    REPORT Dry Snow Metamorphism Final Report Grant: 51065-EV 14. ABSTRACT 16. SECURITY CLASSIFICATION OF: The goal of this project was to characterize the...structural evolution of dry snow as it underwent metamorphism under either quasi-isothermal conditions or a temperature gradient, and to determine...Z39.18 - 5-Aug-2011 Dry Snow Metamorphism Final Report Grant: 51065-EV Report Title ABSTRACT The goal of this project was to characterize the structural

  10. Candida biotypes isolated from clinical specimens in Malaysia.

    PubMed

    Ng, K P; Madasamy, M; Saw, T L; Baki, A; He, J; Soo-Hoo, T S

    The distribution of Candida species was examined using 1114 yeasts isolated from various clinical specimens. The isolates were identified by germ tube test, hyphal/pseudohyphae and chlamydoconidia production and carbohydrate assimilation test using ten carbohydrates (glucose, sucrose, trehalose, cellobiose, arabinose, galactose, mannitol, raffinose, lactose and maltose). Among the 1114 isolates studied, 9 species of Candida were identified and the relative frequency of isolation was C. albicans (44.2%), C. parapsilosis (26.0%), C. tropicalis (17.7%), C. glabrata (9.6%), C. krusei (1.2%), C. rugosa (0.6%), C. guilliermondii (0.2%), C. lusitaniae (0.08%) and C. kefyr (0.08%). Non-C. albicans was the most common Candida species isolated from blood, respiratory system, urine and skin. The isolate from vaginal swabs was predominantly C. albicans. 82.2% of C. glabrata and 64.2% of C. krusei isolated in this study were from vaginal swabs.

  11. Salvia divinorum: toxicological aspects and analysis in human biological specimens.

    PubMed

    Margalho, Cláudia; Corte-Real, Francisco; López-Rivadulla, Manuel; Gallardo, Eugenia

    2016-07-01

    The identification and quantitation of the main psychoactive component of Salvia divinorum (salvinorin A) in biological specimens are crucial in forensic and clinical toxicology. Despite all the efforts made, its uncontrolled abuse has increased quickly, exposing its users' health to serious risks both in the short and long term. The use of alternative biological matrices in toxicological analyzes can be advantageous as complementary postmortem samples, or in situations when neither blood nor urine can be collected; they may be useful tools in those determinations, providing important information about prior exposure. The aim of this article is to present a brief summary of legal aspects of Salvia divinorum and salvinorin A, including the methods used for the determination of the latter in biological matrices.

  12. Testing Na+ in blood

    PubMed Central

    Lava, Sebastiano A.G.; Bianchetti, Mario G.; Milani, Gregorio P.

    2017-01-01

    Abstract Both direct potentiometry and indirect potentiometry are currently used for Na+ testing in blood. These measurement techniques show good agreement as long as protein and lipid concentrations in blood remain normal. In severely ill patients, indirect potentiometry commonly leads to relevant errors in Na+ estimation: 25% of specimens show a disagreement between direct and indirect potentiometry, which is ≥4 mmol/L (mostly spuriously elevated Na+ level due to low circulating albumin concentration). There is a need for increased awareness of the poor performance of indirect potentiometry in some clinical settings.

  13. Blood Vessel Tension Tester

    NASA Technical Reports Server (NTRS)

    1978-01-01

    In the photo, a medical researcher is using a specially designed laboratory apparatus for measuring blood vessel tension. It was designed by Langley Research Center as a service to researchers of Norfolk General Hospital and Eastern Virginia Medical School, Norfolk, Virginia. The investigators are studying how vascular smooth muscle-muscle in the walls of blood vessels-reacts to various stimulants, such as coffee, tea, alcohol or drugs. They sought help from Langley Research Center in devising a method of measuring the tension in blood vessel segments subjected to various stimuli. The task was complicated by the extremely small size of the specimens to be tested, blood vessel "loops" resembling small rubber bands, some only half a millimeter in diameter. Langley's Instrumentation Development Section responded with a miniaturized system whose key components are a "micropositioner" for stretching a length of blood vessel and a strain gage for measuring the smooth muscle tension developed. The micropositioner is a two-pronged holder. The loop of Mood vessel is hooked over the prongs and it is stretched by increasing the distance between the prongs in minute increments, fractions of a millimeter. At each increase, the tension developed is carefully measured. In some experiments, the holder and specimen are lowered into the test tubes shown, which contain a saline solution simulating body fluid; the effect of the compound on developed tension is then measured. The device has functioned well and the investigators say it has saved several months research time.

  14. Blood drop patterns: Formation and applications.

    PubMed

    Chen, Ruoyang; Zhang, Liyuan; Zang, Duyang; Shen, Wei

    2016-05-01

    The drying of a drop of blood or plasma on a solid substrate leads to the formation of interesting and complex patterns. Inter- and intra-cellular and macromolecular interactions in the drying plasma or blood drop are responsible for the final morphologies of the dried patterns. Changes in these cellular and macromolecular components in blood caused by diseases have been suspected to cause changes in the dried drop patterns of plasma and whole blood, which could be used as simple diagnostic tools to identify the health of humans and livestock. However, complex physicochemical driving forces involved in the pattern formation are not fully understood. This review focuses on the scientific development in microscopic observations and pattern interpretation of dried plasma and whole blood samples, as well as the diagnostic applications of pattern analysis. Dried drop patterns of plasma consist of intricate visible cracks in the outer region and fine structures in the central region, which are mainly influenced by the presence and concentration of inorganic salts and proteins during drying. The shrinkage of macromolecular gel and its adhesion to the substrate surface have been thought to be responsible for the formation of the cracks. Dried drop patterns of whole blood have three characteristic zones; their formation as functions of drying time has been reported in the literature. Some research works have applied engineering treatment to the evaporation process of whole blood samples. The sensitivities of the resultant patterns to the relative humidity of the environment, the wettability of the substrates, and the size of the drop have been reported. These research works shed light on the mechanisms of spreading, evaporation, gelation, and crack formation of the blood drops on solid substrates, as well as on the potential applications of dried drop patterns of plasma and whole blood in diagnosis.

  15. Effect of blood contamination on shear bond strength of brackets bonded with a self-etching primer combined with a resin-modified glass ionomer.

    PubMed

    Cacciafesta, Vittorio; Sfondrini, Maria Francesca; Scribante, Andrea; De Angelis, Marco; Klersy, Catherine

    2004-12-01

    This study assessed the effect of blood contamination on the shear bond strength and bond failure site of a resin-modified glass ionomer (Fuji Ortho LC, GC Europe, Leuven, Belgium) used with 3 enamel conditioners (10% polyacrylic acid, 37% phosphoric acid, and self-etching primer). One hundred twenty bovine permanent mandibular incisors were randomly divided into 8 groups; each group consisted of 15 specimens. Two enamel surface conditions were studied: dry and contaminated with blood. One hundred twenty stainless steel brackets were bonded with the resin-modified glass ionomer. After bonding, all samples were stored in distilled water for 24 hours and then tested in shear mode on a testing machine. The groups conditioned with self-etching primer and 37% phosphoric acid had the highest bond strengths for both dry and blood-contaminated enamel. The groups conditioned with 10% polyacrylic acid showed significantly lower shear bond strength value, and the unconditioned groups had the lowest bond strengths. For each enamel conditioner, no significant difference was reported between dry and blood-contaminated groups. Significant differences in debond locations were found among the groups bonded with the different conditioners. Blood contamination of enamel during the bonding procedure of Fuji Ortho LC did not affect its bond strength values, no matter which enamel conditioner was used.

  16. Human Blood Typing: A Forensic Science Approach. Part I: Background.

    ERIC Educational Resources Information Center

    Kobilinsky, Lawrence; Sheehan, Francis X.

    1988-01-01

    In this article, part I of a series, the forensic methods used in "typing" human blood, which as physical evidence is often found in the dried state, are outlined. Background information about individualization, antibody typing, fresh blood, dried blood, and additional systems is provided. (CW)

  17. Blood clotting

    MedlinePlus Videos and Cool Tools

    ... the external bleeding stops. Clotting factors in the blood cause strands of blood-borne material, called fibrin, to stick together and ... the inside of the wound. Eventually, the cut blood vessel heals, and the blood clot dissolves after ...

  18. Blood culture

    MedlinePlus

    Culture - blood ... A blood sample is needed . The site where blood will be drawn is first cleaned with an antiseptic such ... organism from the skin getting into (contaminating) the blood sample and causing a false-positive result (see ...

  19. Virtual blood bank.

    PubMed

    Wong, Kit Fai

    2011-01-24

    Virtual blood bank is the computer-controlled, electronically linked information management system that allows online ordering and real-time, remote delivery of blood for transfusion. It connects the site of testing to the point of care at a remote site in a real-time fashion with networked computers thus maintaining the integrity of immunohematology test results. It has taken the advantages of information and communication technologies to ensure the accuracy of patient, specimen and blood component identification and to enhance personnel traceability and system security. The built-in logics and process constraints in the design of the virtual blood bank can guide the selection of appropriate blood and minimize transfusion risk. The quality of blood inventory is ascertained and monitored, and an audit trail for critical procedures in the transfusion process is provided by the paperless system. Thus, the virtual blood bank can help ensure that the right patient receives the right amount of the right blood component at the right time.

  20. Spalling Experiments on Large Hard Rock Specimens

    NASA Astrophysics Data System (ADS)

    Jacobsson, Lars; Appelquist, Karin; Lindkvist, Jan Erik

    2015-07-01

    Specimens of coarse-grained Äspö diorite were axially compressed to observe stress-induced spalling. The specimens had a novel design characterized by two manufactured large radius notches on opposite sides. The tangential stress occurring in the notches aimed to represent the tangential loading around a circular opening. Fracture stages were monitored by acoustic emission measurements. Rock chips were formed similar to those found in situ, which indicates a similar fracture process. Slabs were cut out from the specimens and impregnated using a fluorescent material to visualize the cracks. The cracks were subsequently examined by the naked eye and by means of microscopy images, from which fracture paths could be identified and related to different minerals and their crystallographic orientations. The microscopy analyses showed how the stress field and the microstructure interact. Parallel cracks were formed 2-4 mm below the surface, sub-parallel to the direction of the maximum principal stress. The crack initiation, the roles of minerals such as feldspar, biotite and quartz and their grain boundaries and crystallographic directions are thoroughly studied and discussed in this paper. Scale effects, which relate to the stress gradient and microstructure, are discussed.

  1. A Review of Sleeve Gastrectomy Specimen Histopathology.

    PubMed

    Kinsinger, Luke A; Garber, James C; Whipple, Oliver

    2016-11-01

    With the increasing popularity of sleeve gastrectomy, many stomach specimens are being evaluated. Understanding the significance and treatment for unexpected pathology is important. This study examines the incidence of relevant histopathology of sleeve gastrectomy specimens. It evaluates previous data for each histopathology and provides recommendations for treatment. In this study, a retrospective review was performed for 241 patients who underwent sleeve gastrectomy from 2009 to 2014 at a single institution. Of the specimens, 122 had no significant histopathology, 91 had gastritis, 13 had lymphoid aggregates, 5 had hyperplasia, 3 had intestinal metaplasia, 3 had gastrointestinal stromal tumors (GISTs), and 3 had gastric polyps. Of the GISTs all had a low mitotic rate and the size of the tumor ranged from 1.5 to 4.5 cm. The findings of metaplasia may be a marker for increased risk of malignancy and may require additional surveillance. The findings of GIST may warrant interval imaging to survey for recurrence, though the likelihood of recurrence for the tumors in this study is less than 2 per cent based on previous studies.

  2. Dynamic Fracture Studies using Sleeved Taylor Specimens

    NASA Astrophysics Data System (ADS)

    Gilmore, Martin; Foster, Joseph, Jr.; Wilson, Leonard L.; Cullis, Ian

    2001-06-01

    The characterization of the inelastic response of materials to high rates of loading is a challenging engineering problem. As the load rate increases, the interpretation of the data recovered from the experiment become more difficult. At very high rates of loading, even the inertia of the test specimen must be accounted for in the interpretation of the data. The Taylor impact experiment is specifically designed to exploit the inertia of the specimen to produce very high loading rates and has been used to study the high strain (50materials for many years. Many high-rate loading problems produce failure in the material. This paper addresses the use of the Taylor impact experiment to study these failures. Continuum codes have been used to design sleeved impact specimens to study the failure of materials under high rates of loading. Ductile core materials are used as drivers to control rupture of more brittle sleeves of the material of interest. Annealed copper cores are used to drive dynamic failure a selection of steels. High rate plastic deformation data is presented for the driver and the sleeve together with the fracture data.

  3. Dynamic Fracture Studies Using Sleeved Taylor Specimens

    NASA Astrophysics Data System (ADS)

    Gilmore, Martin R.; Foster, Joseph C., Jr; Wilson, Leo L.

    2002-07-01

    The characterization of the inelastic response of materials to high rates of loading is a challenging engineering problem. As the load rate increases, the interpretation of the data recovered from the experiment become more difficult. At very high rates of loading, even the inertia of the test specimen must be accounted for in the interpretation of the data 1. The Taylor impact experiment is specifically designed to exploit the inertia of the specimen to produce very high loading rates and has been used to study the high strain (50%), high strain rate (103-4) behavior of materials for many years 2. Many high-rate loading problems produce failure in the material. Continuum codes have been used to design sleeved impact specimens to study the failure of materials under high rates of loading. Ductile core materials are used as drivers to control rupture of more brittle sleeves of the material of interest. Annealed copper cores are used to drive dynamic failure in AF1410 steel. High rate plastic deformation data are presented for the driver and the sleeve together with the fracture data.

  4. Motorized manipulator for positioning a TEM specimen

    DOEpatents

    Schmid, Andreas Karl; Andresen, Nord

    2010-12-14

    The invention relates to a motorized manipulator for positioning a TEM specimen holder with sub-micron resolution parallel to a y-z plane and rotating the specimen holder in the y-z plane, the manipulator comprising a base (2), and attachment means (30) for attaching the specimen holder to the manipulator, characterized in that the manipulator further comprises at least three nano-actuators (3.sup.a, 3.sup.b, 3.sup.c) mounted on the base, each nano-actuator showing a tip (4.sup.a, 4.sup.b, 4.sup.c), the at least three tips defining the y-z plane, each tip capable of moving with respect to the base in the y-z plane; a platform (5) in contact with the tips of the nano-actuators; and clamping means (6) for pressing the platform against the tips of the nano-actuators; as a result of which the nano-actuators can rotate the platform with respect to the base in the y-z plane and translate the platform parallel to the y-z plane.

  5. 10 CFR 26.165 - Testing split specimens and retesting single specimens.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... in Bottle B shall provide quantitative test results to the MRO and the MRO shall provide them to the... retesting specimens that have been subject to the special analysis permitted in § 26.163(a)(2). (2)...

  6. Influence of Specimen Preparation and Specimen Size on Composite Transverse Tensile Strength and Scatter

    NASA Technical Reports Server (NTRS)

    OBrien, T. Kevin; Chawan, Arun D.; DeMarco, Kevin; Paris, Isabelle

    2001-01-01

    The influence of specimen polishing, configuration, and size on the transverse tension strength of two glass-epoxy materials, and one carbon-epoxy material, loaded in three and four point bending was evaluated. Polishing machined edges, arid/or tension side failure surfaces, was detrimental to specimen strength characterization instead of yielding a higher, more accurate, strength as a result of removing inherent manufacture and handling flaws. Transverse tension strength was typically lower for longer span lengths due to the classical weakest link effect. However, strength was less sensitive to volume changes achieved by increasing specimen width. The Weibull scaling law typically over-predicted changes in transverse tension strengths in three point bend tests and under-predicted changes in transverse tension strengths in four point bend tests. Furthermore, the Weibull slope varied with specimen configuration, volume, and sample size. Hence, this scaling law was not adequate for predicting transverse tension strength of heterogeneous, fiber-reinforced, polymer matrix composites.

  7. Specimen loading list for the varying temperature experiment

    SciTech Connect

    Qualls, A.L.; Sitterson, R.G.

    1998-09-01

    The varying temperature experiment HFIR-RB-13J has been assembled and inserted in the reactor. Approximately 5300 specimens were cleaned, inspected, matched, and loaded into four specimen holders. A listing of each specimen loaded into the steady temperature holder, its position in the capsule, and the identification of the corresponding specimen loaded into the varying temperature holder is presented in this report.

  8. Axial-Gap Induction Motor For Levitated Specimens

    NASA Technical Reports Server (NTRS)

    Sridharan, Govind; Rhim, Won-Kyu; Barber, Dan; Chung, Sang

    1992-01-01

    Motor does not obscure view of specimen. Axial-gap induction motor applies torque to rotate electrostatically or electromagnetically levitated specimen of metal. Possible applications include turning specimens for uniform heating under focused laser beams and obtaining indirect measurements of resistivities or of surface tensions in molten specimens.

  9. Recommendations for collection and handling of specimens from group breast cancer clinical trials.

    PubMed

    Leyland-Jones, Brian R; Ambrosone, Christine B; Bartlett, John; Ellis, Matthew J C; Enos, Rebecca A; Raji, Adekunle; Pins, Michael R; Zujewski, Jo Anne; Hewitt, Stephen M; Forbes, John F; Abramovitz, Mark; Braga, Sofia; Cardoso, Fatima; Harbeck, Nadia; Denkert, Carsten; Jewell, Scott D

    2008-12-01

    Recommendations for specimen collection and handling have been developed for adoption across breast cancer clinical trials conducted by the Breast International Group (BIG)-sponsored Groups and the National Cancer Institute (NCI)-sponsored North American Cooperative Groups. These recommendations are meant to promote identifiable standards for specimen collection and handling within and across breast cancer trials, such that the variability in collection/handling practices that currently exists is minimized and specimen condition and quality are enhanced, thereby maximizing results from specimen-based diagnostic testing and research. Three working groups were formed from the Cooperative Group Banking Committee, BIG groups, and North American breast cancer cooperative groups to identify standards for collection and handling of (1) formalin-fixed, paraffin-embedded (FFPE) tissue; (2) blood and its components; and (3) fresh/frozen tissue from breast cancer trials. The working groups collected standard operating procedures from multiple group specimen banks, administered a survey on banking practices to those banks, and engaged in a series of discussions from 2005 to 2007. Their contributions were synthesized into this document, which focuses primarily on collection and handling of specimens to the point of shipment to the central bank, although also offers some guidance to central banks. Major recommendations include submission of an FFPE block, whole blood, and serial serum or plasma from breast cancer clinical trials, and use of one fixative and buffer type (10% neutral phosphate-buffered formalin, pH 7) for FFPE tissue across trials. Recommendations for proper handling and shipping were developed for blood, serum, plasma, FFPE, and fresh/frozen tissue.

  10. Recommendations for Collection and Handling of Specimens From Group Breast Cancer Clinical Trials

    PubMed Central

    Leyland-Jones, Brian R.; Ambrosone, Christine B.; Bartlett, John; Ellis, Matthew J.C.; Enos, Rebecca A.; Raji, Adekunle; Pins, Michael R.; Zujewski, Jo Anne; Hewitt, Stephen M.; Forbes, John F.; Abramovitz, Mark; Braga, Sofia; Cardoso, Fatima; Harbeck, Nadia; Denkert, Carsten; Jewell, Scott D.

    2008-01-01

    Recommendations for specimen collection and handling have been developed for adoption across breast cancer clinical trials conducted by the Breast International Group (BIG)-sponsored Groups and the National Cancer Institute (NCI)-sponsored North American Cooperative Groups. These recommendations are meant to promote identifiable standards for specimen collection and handling within and across breast cancer trials, such that the variability in collection/handling practices that currently exists is minimized and specimen condition and quality are enhanced, thereby maximizing results from specimen-based diagnostic testing and research. Three working groups were formed from the Cooperative Group Banking Committee, BIG groups, and North American breast cancer cooperative groups to identify standards for collection and handling of (1) formalin-fixed, paraffin-embedded (FFPE) tissue; (2) blood and its components; and (3) fresh/frozen tissue from breast cancer trials. The working groups collected standard operating procedures from multiple group specimen banks, administered a survey on banking practices to those banks, and engaged in a series of discussions from 2005 to 2007. Their contributions were synthesized into this document, which focuses primarily on collection and handling of specimens to the point of shipment to the central bank, although also offers some guidance to central banks. Major recommendations include submission of an FFPE block, whole blood, and serial serum or plasma from breast cancer clinical trials, and use of one fixative and buffer type (10% neutral phosphate-buffered formalin, pH 7) for FFPE tissue across trials. Recommendations for proper handling and shipping were developed for blood, serum, plasma, FFPE, and fresh/frozen tissue. PMID:18955459

  11. Indiana Corn Dry Mill

    SciTech Connect

    2006-09-01

    The goal of this project is to perform engineering, project design, and permitting for the creation and commercial demonstration of a corn dry mill biorefinery that will produce fuel-grade ethanol, distillers dry grain for animal feed, and carbon dioxide for industrial use.

  12. Tray Drying of Solids.

    ERIC Educational Resources Information Center

    Afacan, Artin; Masliyah, Jacob

    1984-01-01

    Describes a drying experiment useful in presenting the concept of simultaneous heat and mass transfer. Background information, equipment requirements, experimental procedures, and results are provided. The reasonably good agreement in the calculated rate of drying and that observed experimentally makes students feel confident in applying…

  13. A review of chevron-notched fracture specimens

    NASA Technical Reports Server (NTRS)

    Newman, J. C., Jr.

    1984-01-01

    The historical development of chevron notched fracture specimens is reviewed. Stress intensity factors and load line displacement solutions proposed for some of these specimens are compared. The original bend bar configurations up to the present day short rod and bar specimens are reviewed. The results of an analytical round robin that was conducted on chevron-notched specimens are presented. In the round robin, stress-intensity factors for either the chevron notched round rod or square bar specimens were calculated. The consensus stress intensity factor (compliance) solution for these specimens is assessed. The stress intensity factor solutions proposed for three and four point bend chevron notched specimens are reviewed.

  14. The fracture toughness of wood in compact specimens (CT specimens) tested for off-center tension

    SciTech Connect

    Gappoev, M.M.

    1995-07-01

    The application of fracture mechanics methods in the design of building construction requires reliable knowledge of material crack resistance characteristics. At present, no standard test techniques for crack resistance in wood materials have been developed. In testing wood for crack resistance, it has been recommended that single-notched beam specimens (of the SENB type) subjected to three-point bending be used. The recommended method is rather simple; however, it has disadvantages that impair its accuracy. In particular, errors may arise from not directly accounting for the proper mass of a specimen. To evaluate these errors and to lend support to our previous results obtained on beam specimens, we tested compact specimens for off-center tension. Methods for determining the crack resistance characteristics (G{sub Ic,}, G{sub IF}, K{sub Ic}) of compact wood specimens (CT-specimens), tested for off-center tension, are described. The behavior of the fracture energy G{sub IF} during crack propagation is discussed.

  15. Preparation of Regular Specimens for Atom Probes

    NASA Technical Reports Server (NTRS)

    Kuhlman, Kim; Wishard, James

    2003-01-01

    A method of preparation of specimens of non-electropolishable materials for analysis by atom probes is being developed as a superior alternative to a prior method. In comparison with the prior method, the present method involves less processing time. Also, whereas the prior method yields irregularly shaped and sized specimens, the present developmental method offers the potential to prepare specimens of regular shape and size. The prior method is called the method of sharp shards because it involves crushing the material of interest and selecting microscopic sharp shards of the material for use as specimens. Each selected shard is oriented with its sharp tip facing away from the tip of a stainless-steel pin and is glued to the tip of the pin by use of silver epoxy. Then the shard is milled by use of a focused ion beam (FIB) to make the shard very thin (relative to its length) and to make its tip sharp enough for atom-probe analysis. The method of sharp shards is extremely time-consuming because the selection of shards must be performed with the help of a microscope, the shards must be positioned on the pins by use of micromanipulators, and the irregularity of size and shape necessitates many hours of FIB milling to sharpen each shard. In the present method, a flat slab of the material of interest (e.g., a polished sample of rock or a coated semiconductor wafer) is mounted in the sample holder of a dicing saw of the type conventionally used to cut individual integrated circuits out of the wafers on which they are fabricated in batches. A saw blade appropriate to the material of interest is selected. The depth of cut and the distance between successive parallel cuts is made such that what is left after the cuts is a series of thin, parallel ridges on a solid base. Then the workpiece is rotated 90 and the pattern of cuts is repeated, leaving behind a square array of square posts on the solid base. The posts can be made regular, long, and thin, as required for samples

  16. Dry imaging cameras.

    PubMed

    Indrajit, Ik; Alam, Aftab; Sahni, Hirdesh; Bhatia, Mukul; Sahu, Samaresh

    2011-04-01

    Dry imaging cameras are important hard copy devices in radiology. Using dry imaging camera, multiformat images of digital modalities in radiology are created from a sealed unit of unexposed films. The functioning of a modern dry camera, involves a blend of concurrent processes, in areas of diverse sciences like computers, mechanics, thermal, optics, electricity and radiography. Broadly, hard copy devices are classified as laser and non laser based technology. When compared with the working knowledge and technical awareness of different modalities in radiology, the understanding of a dry imaging camera is often superficial and neglected. To fill this void, this article outlines the key features of a modern dry camera and its important issues that impact radiology workflow.

  17. Packaged kiln dried firewood

    SciTech Connect

    Cutrara, A.

    1986-07-01

    A process is described for kiln drying firewood consisting of essentially uniform lengths of split firewood pieces, the process comprising splitting essentially uniform lengths of green tree logs to form firewood pieces, placing the firewood pieces in open mesh bags to provide a plurality of bags of firewood, placing the plurality of bags of green firewood pieces in a kiln drying oven, kiln drying the pieces at temperatures in excess of 150/sup 0/F. by moving heated air over the pieces until the pieces have an overall moisture content ranging from 15% up to 30% by weight, operating the kiln at a temperature below a level which would render the structural characteristics of the bag useless and removing the kiln dried firewood pieces in the plurality of bags from the kiln drying oven.

  18. Process and apparatus for analyzing specimens for the presence of microorganisms therein

    NASA Technical Reports Server (NTRS)

    Aldridge, Jr., Clifton (Inventor); Jones, Paul W. (Inventor); Gibson, Sandra F. (Inventor); Vannest, Richard D. (Inventor); Holen, James T. (Inventor); Keyser, George F. (Inventor); Meyer, Michael C. (Inventor)

    1980-01-01

    Microorganisms in a specimen are detected, identified, and enumerated by introducing the specimen into a sampling cartridge and diluting the specimen with a known volume of water within the cartridge. The cartridge has a manifold and several cassettes attached to the manifold. Each cassette contains a serpentine flow channel having a series of filters therein and a detection cell located downstream from each filter. The flow channel in each cassette also contains a culture medium which is freeze dried and is highly selective in the sense that it promotes the growth of one type of microorganism, but not others. The mixture of the specimen and water flows from the manifold into the flow channel of each cassette where it rehydrates the culture medium therein and further flows through the filters. Each filter removes a known proportion of the microorganisms from the mixture of specimen, water and medium, thereby effecting a serial dilution. After the cassettes are heated to incubate the microoganisms, the detection cells are observed for growth of the microorganisms therein which is manifested in a change in the light transmitting characteristics of the mixtures within the cells.

  19. Measurement of vasoactive intestinal peptide using a competitive fluorescent microsphere immunoassay or ELISA in human blood samples.

    PubMed

    Song, Eun Young; VanDunk, Cassandra; Kuddo, Thea; Nelson, Phillip G

    2005-05-01

    The concentration of Vasoactive Intestinal Peptide (VIP) as measured by recycling immunoaffinity chromatography (RIC) has been reported to be elevated in the blood of patients with autism as compared with normal subjects. In this study, we have developed a "Competitive Fluorescent Microsphere Immunoassay" (cFMI) in which VIP competes with biotinylated VIP in binding to polyclonal antibodies on microspheres. The results were obtained using the Luminex100 system. We measured VIP in serum, plasma, and material eluted from dried blood spots on filter paper with both the cFMI and an ELISA procedure. We found that a purification procedure was necessary for obtaining useful results from plasma and serum, however, a preincubation step was required with the blood eluates. This newly developed cFMI was more sensitive (2.5 vs. 20.0 pg/ml), and more reproducible than the ELISA. To get accurate measurements of VIP in eluted material high sensitivity is especially important. Thus, the cFMI using the Luminex system has definite advantages over a conventional ELISA including the possibility that samples can be assayed at higher dilutions. We have determined that the VIP concentrations of serum, plasma, and dried blood spot eluate specimens as measured with the cFMI assay system were similar to those measured with ELISA. Thus, the new cFMI using Luminex system may be useful for detection of VIP in human blood samples.

  20. Detection of iso-α-acids to confirm beer consumption in postmortem specimens.

    PubMed

    Rodda, Luke N; Gerostamoulos, Dimitri; Drummer, Olaf H

    2015-01-01

    Iso-α-acids (IAAs) can be used as markers for the consumption of beer. Postmortem specimens from a range of coronial cases were analyzed for IAAs in order to determine the prevalence of beer consumption and any correlation to blood alcohol concentrations (BAC). A total of 130 cases were included in this study including those where beer was mentioned in the case circumstances, cases where beer was not mentioned specifically but alcohol was detected, and cases where neither beer was mentioned nor a positive BAC was present. Available blood, serum, vitreous humour and urine specimens were analyzed. Of the 50 cases where beer was mentioned, 86% had one or more IAAs detected. In cases that only had a positive BAC (n = 60), 57% of these cases also showed the presence of these beer markers. IAAs were detected in specimens obtained from traumatized, burnt, and decomposed cases with a mention of beer consumption or where BAC was positive in blood. No IAAs were detected in cases where BAC was negative. There was little or no correlation between blood IAA concentrations and BAC. This study demonstrates the possible detection of IAAs as a marker for beer consumption.

  1. 21 CFR 640.5 - Testing the blood.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 7 2010-04-01 2010-04-01 false Testing the blood. 640.5 Section 640.5 Food and... ADDITIONAL STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Whole Blood § 640.5 Testing the blood. All laboratory tests shall be made on a specimen of blood taken from the donor at the time of collecting the unit...

  2. 21 CFR 640.5 - Testing the blood.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 7 2014-04-01 2014-04-01 false Testing the blood. 640.5 Section 640.5 Food and... ADDITIONAL STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Whole Blood § 640.5 Testing the blood. All laboratory tests shall be made on a specimen of blood taken from the donor at the time of collecting the unit...

  3. 21 CFR 640.5 - Testing the blood.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 7 2012-04-01 2012-04-01 false Testing the blood. 640.5 Section 640.5 Food and... ADDITIONAL STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Whole Blood § 640.5 Testing the blood. All laboratory tests shall be made on a specimen of blood taken from the donor at the time of collecting the unit...

  4. 21 CFR 640.5 - Testing the blood.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 7 2013-04-01 2013-04-01 false Testing the blood. 640.5 Section 640.5 Food and... ADDITIONAL STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Whole Blood § 640.5 Testing the blood. All laboratory tests shall be made on a specimen of blood taken from the donor at the time of collecting the unit...

  5. 21 CFR 640.5 - Testing the blood.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 7 2011-04-01 2010-04-01 true Testing the blood. 640.5 Section 640.5 Food and... ADDITIONAL STANDARDS FOR HUMAN BLOOD AND BLOOD PRODUCTS Whole Blood § 640.5 Testing the blood. All laboratory tests shall be made on a specimen of blood taken from the donor at the time of collecting the unit...

  6. [Real-time PCR detection of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae DNA in clinical specimens].

    PubMed

    Vacková, Z; Lžičařová, D; Stock, N K; Kozáková, J

    2015-10-01

    The study aim was to implement a molecular real-time polymerase chain reaction (PCR) assay recommended by the CDC (Centers for Disease Control and Prevention) for the detection of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae in clinical (culture negative) specimens from patients with suspected invasive bacterial disease. Clinical specimens are referred to the National Reference Laboratory (NRL) for Meningococcal Infections, Unit for Airborne Bacterial Infections, Centre for Epidemiology and Microbiology, National Institute of Public Health from various regions of the Czech Republic. Clinical specimens are, in particular, cerebrospinal fluid, anti-coagulated blood or serum and, exceptionally, post-mortem specimens. The NRL has implemented molecular diagnosis of these bacterial pathogens involved in meningitis and sepsis from clinical specimens since 1999. The first diagnostic method was semi-nested PCR followed by electrophoretic analysis. In 2014, a molecular qualitative real-time PCR assay was implemented.

  7. Characterization of Brevibacterium spp. from clinical specimens.

    PubMed Central

    Gruner, E; Pfyffer, G E; von Graevenitz, A

    1993-01-01

    Nonfermenting coryneform bacteria identified as Brevibacterium spp. were isolated from routine clinical specimens. Four strains were derived from peritoneal fluid and has presumably been involved in the pathogenesis of continuous ambulatory peritoneal dialysis peritonitis. Another five isolates most probably represented skin contaminants. Cell wall and lipid analyses confirmed the genus identification. Strains in this taxon are difficult to distinguish from other biochemically inactive and nonmotile coryneform species but show characteristics cellular fatty acid profiles. In vitro susceptibilities to commonly used antibiotics were determined. PMID:8314980

  8. Gas permeation measurements on small polymer specimens

    NASA Technical Reports Server (NTRS)

    Burns, Karen S.; Vannorman, John D.

    1988-01-01

    Mass spectrometry was used to measure oxygen and nitrogen permeabilities while polarography was used to measure oxygen permeabilities for several contact lens materials. Applicable sample holders were designed and fabricated to accommodate curved and flat specimens. A prepared standard was used to calibrate the mass spectrometric analyses. The oxygen permeability values determined by mass spectrometry were significantly greater than those determined by polarography. This was attributed to the phase boundary phenomena and the limiting oxygen permeance of water inherent in the polarographic technique. Polarographic values determined were in good agreement with proprietary values obtained by polarography, with the exception of one material.

  9. Controlled crack growth specimen for brittle systems

    NASA Technical Reports Server (NTRS)

    Calomino, Anthony M.; Brewer, David N.

    1992-01-01

    A pure Mode 1 fracture specimen and test procedure has been developed which provides extended, stable, through-thickness crack growth in ceramics and other brittle, nonmetallic materials. Fixed displacement loading, applied at the crack mouth, promotes stable crack extension by reducing the stored elastic strain energy. Extremely fine control of applied displacements is achieved by utilizing the Poisson's expansion of a compressively loaded cylindrical pin. Stable cracks were successfully grown in soda-lime glass and monolithic Al2O3 for lengths in excess of 2O mm without uncontrollable catastrophic failure.

  10. Controlled crack growth specimen for brittle systems

    NASA Technical Reports Server (NTRS)

    Calomino, Anthony M.; Brewer, David N.

    1990-01-01

    A pure Mode 1 fracture specimen and test procedure has been developed which provides extended, stable, through-thickness crack growth in ceramics and other brittle, nonmetallic materials. Fixed displacement loading, applied at the crack mouth, promotes stable crack extension by reducing the stored elastic strain energy. Extremely fine control of applied displacements is achieved by utilizing the Poisson's expansion of a compressively loaded cylindrical pin. Stable cracks were successfully grown in soda-lime glass and monolithic Al2O3 for lengths in excess of 20 mm without uncontrollable catastrophic failure.

  11. Influence of Water Content on Mechanical Properties of Rock in Both Saturation and Drying Processes

    NASA Astrophysics Data System (ADS)

    Zhou, Zilong; Cai, Xin; Cao, Wenzhuo; Li, Xibing; Xiong, Cheng

    2016-08-01

    Water content has a pronounced influence on the properties of rock materials, which is responsible for many rock engineering hazards, such as landslides and karst collapse. Meanwhile, water injection is also used for the prevention of some engineering disasters like rock-bursts. To comprehensively investigate the effect of water content on mechanical properties of rocks, laboratory tests were carried out on sandstone specimens with different water contents in both saturation and drying processes. The Nuclear Magnetic Resonance technique was applied to study the water distribution in specimens with variation of water contents. The servo-controlled rock mechanics testing machine and Split Hopkinson Pressure Bar technique were used to conduct both compressive and tensile tests on sandstone specimens with different water contents. From the laboratory tests, reductions of the compressive and tensile strength of sandstone under static and dynamic states in different saturation processes were observed. In the drying process, all of the saturated specimens could basically regain their mechanical properties and recover its strength as in the dry state. However, for partially saturated specimens in the saturation and drying processes, the tensile strength of specimens with the same water content was different, which could be related to different water distributions in specimens.

  12. Development of fatigue life evaluation method using small specimen

    NASA Astrophysics Data System (ADS)

    Nogami, Shuhei; Nishimura, Arata; Wakai, Eichi; Tanigawa, Hiroyasu; Itoh, Takamoto; Hasegawa, Akira

    2013-10-01

    For developing the fatigue life evaluation method using small specimen, the effect of specimen size and shape on the fatigue life of the reduced activation ferritic/martensitic steels (F82H-IEA, F82H-BA07 and JLF-1) was investigated by the fatigue test at room temperature in air using round-bar and hourglass specimens with various specimen sizes (test section diameter: 0.85-10 mm). The round-bar specimen showed no specimen size and no specimen shape effects on the fatigue life, whereas the hourglass specimen showed no specimen size effect and obvious specimen shape effect on it. The shorter fatigue life of the hourglass specimen observed under low strain ranges could be attributed to the shorter micro-crack initiation life induced by the stress concentration dependent on the specimen shape. On the basis of this study, the small round-bar specimen was an acceptable candidate for evaluating the fatigue life using small specimen.

  13. Ophthalmic use of blood-derived products.

    PubMed

    Nugent, Ryan B; Lee, Graham A

    2015-01-01

    There is a wide spectrum of blood-derived products that have been used in many different medical and surgical specialties with success. Blood-derived products for clinical use can be extracted from autologous or allogeneic specimens of blood, but recombinant products are also commonly used. A number of blood derivatives have been used for a wide range of ocular conditions, from the ocular surface to the retina. With stringent preparation guidelines, the potential risk of transmission of blood-borne diseases is minimized. We review blood-derived products and how they are improving the management of ocular disease.

  14. Cranial Pathologies in a Specimen of Pachycephalosaurus

    PubMed Central

    Peterson, Joseph E.; Vittore, Christopher P.

    2012-01-01

    Background A frontoparietal dome of a large pachycephalosaurid collected from the Upper Cretaceous Hell Creek Formation in 2001 is identified as Pachycephalosaurus wyomingensis. The specimen features two large oval depressions on the dorsal surface, accompanied by numerous circular pits on the margin and inner surface of the larger depressions. Methodology/Principal Findings In order to identify the origin of these structures, computed tomography (CT) data and morphological characteristics of the specimen are analyzed and compared with similar osteological structures in fossil and extant archosaurs caused by taphonomic processes, non-pathologic bone resorption, and traumatic infection/inflammatory origins. The results of these analyses suggest that the structures are pathologic lesions likely resulting from a traumatic injury and followed by secondary infection at the site. Conclusions/Significance The presence of lesions on a frontoparietal dome, and the exclusivity of their distribution along the dorsal dome surface, offers further insight into frontoparietal dome function and supports previously hypothesized agonistic behavior in pachycephalosaurids. PMID:22558394

  15. Wildlife specimen collection, preservation, and shipment

    USGS Publications Warehouse

    White, C. LeAnn; Dusek, Robert J.; Franson, J. Christian; Friend, Milton; Gibbs, Samantha E.J.; Wild, Margaret A.

    2015-01-01

    Prior to collecting samples, it is important to determine the capabilities and submission criteria of the laboratory receiving the samples. Some laboratories may specialize in a limited number of tests, be equipped to accept only certain types of tissues (instead of entire carcasses), or specialize in particular species or group of animals (e.g., reptiles, birds, mammals). Diagnostic laboratories have specific requirements regarding preparation, labeling, and shipping of samples. Adherence to these requirements helps ensure the usefulness of any submitted specimens. Although laboratories may vary in the cost and turnaround times for diagnostic tests, some laboratories may be able to prioritize samples and accommodate accelerated time frames if communicated at the time of submission. Keeping a prepacked kit with basic carcass-collection supplies, including a paper copy of the specimen history form (available for download from the Web sites of most diagnostic laboratories), in the office or vehicle will decrease the chances of forgetting an essential item and decrease response time for arriving at an event.

  16. Is Shape of a Fresh and Dried Leaf the Same?

    PubMed Central

    Tomaszewski, Dominik; Górzkowska, Angelika

    2016-01-01

    Plants kept as dried herbarium specimens share many characteristics with their living counterparts, but there are some substantial differences between them. Due to dehydration, leaves of herbarium specimens change not only their mass and colour, but in many cases change their dimensions, too. The present study aimed to determine whether leaf shape changes during the drying process. A total of 794 pairs of fresh and dried leaves or leaflets of 22 plant taxa were studied. The shape of the blades was quantified using elliptic Fourier analysis combined with principal component analysis. In addition, area and mass of the leaves were measured. Statistical tests were applied for comparing fresh and dried leaves. The results indicate that the preservation process of pressing and drying plants for herbarium purposes causes changes in leaf shape. In general, the shape changes were directional. As the shape of fresh and dried plants is different, it is strongly recommended that shape analyses should be performed on datasets containing either of the leaf types. PMID:27045956

  17. Freeze drying method

    DOEpatents

    Coppa, Nicholas V.; Stewart, Paul; Renzi, Ernesto

    1999-01-01

    The present invention provides methods and apparatus for freeze drying in which a solution, which can be a radioactive salt dissolved within an acid, is frozen into a solid on vertical plates provided within a freeze drying chamber. The solid is sublimated into vapor and condensed in a cold condenser positioned above the freeze drying chamber and connected thereto by a conduit. The vertical positioning of the cold condenser relative to the freeze dryer helps to help prevent substances such as radioactive materials separated from the solution from contaminating the cold condenser. Additionally, the system can be charged with an inert gas to produce a down rush of gas into the freeze drying chamber to also help prevent such substances from contaminating the cold condenser.

  18. Freeze drying apparatus

    DOEpatents

    Coppa, Nicholas V.; Stewart, Paul; Renzi, Ernesto

    2001-01-01

    The present invention provides methods and apparatus for freeze drying in which a solution, which can be a radioactive salt dissolved within an acid, is frozen into a solid on vertical plates provided within a freeze drying chamber. The solid is sublimated into vapor and condensed in a cold condenser positioned above the freeze drying chamber and connected thereto by a conduit. The vertical positioning of the cold condenser relative to the freeze dryer helps to help prevent substances such as radioactive materials separated from the solution from contaminating the cold condenser. Additionally, the system can be charged with an inert gas to produce a down rush of gas into the freeze drying chamber to also help prevent such substances from contaminating the cold condenser.

  19. Acoustoconvection Drying of Meat

    NASA Astrophysics Data System (ADS)

    Zhilin, A. A.; Fedorov, A. V.

    2016-03-01

    The dynamics of moisture extraction from meat samples by the acoustoconvection and thermoconvection methods has been investigated. To describe the dynamics of moisture extraction from meat, we propose a simple relaxation model with a relaxation time of 8-10 min in satisfactorily describing experimental data on acoustoconvection drying of meat. For thermoconvection drying the relaxation time is thereby 30 and 45 min for the longitudinal and transverse positions of fibers, respectively.

  20. Characterization of Mechanical Properties of Nuclear Graphite Using Subsize Specimens and Reusing Tested Specimens

    SciTech Connect

    Ji Hyun, Yoon; Byun, Thak Sang; Strizak, Joe P; Snead, Lance Lewis

    2011-01-01

    The mechanical properties of NBG-18 nuclear grade graphite have been characterized using small specimen test techniques and statistical treatment on the test results. New fracture strength and toughness test techniques were developed to use subsize cylindrical specimens with glued heads and to reuse their broken halves. Three sets of subsize cylindrical specimens with the different diameters of 4 mm, 8 mm, and 12 mm were tested to obtain tensile fracture strength. The longer piece of the broken halves was cracked from side surfaces and tested under three-point bend loading to obtain fracture toughness. Both the strength and fracture toughness data were analyzed using Weibull distribution models focusing on size effect. The mean fracture strength decreased from 22.9 MPa to 21.5 MPa as the diameter increased from 4 mm to 12 mm, and the mean strength of 15.9 mm diameter standard specimen, 20.9 MPa, was on the extended trend line. These fracture strength data indicate that in the given diameter range the size effect is not significant and much smaller than that predicted by the Weibull statistics-based model. Further, no noticeable size effect existed in the fracture toughness data, whose mean values were in a narrow range of 1.21 1.26 MPa. The Weibull moduli measured for fracture strength and fracture toughness datasets were around 10. It is therefore believed that the small or negligible size effect enables to use the subsize specimens and that the new fracture toughness test method to reuse the broken specimens to help minimize irradiation space and radioactive waste.

  1. Microwave drying of high strength dental stone: effects on dimensional accuracy.

    PubMed

    Yap, Adrian U J; Yap, S H; Teo, Jason C K; Tay, C M; Ng, K L; Thean, Hilary P Y

    2003-01-01

    High-strength dental stone is widely used to produce dies for the fabrication of restorations with the lost-wax technique. It is normal to wait at least 24 hours for casts to dry and gain sufficient strength prior to initiating laboratory procedures. This waiting time may be greatly reduced by using microwave drying. This study determined the optimum microwave energy density for preserving working die accuracy of a Type IV high-strength dental stone (Silky Rock; Whipmix). Cylindrical die specimens were fabricated according to manufacturer's instructions and allowed to set for one hour. The specimens were subsequently treated as follows: Group I (Control group)--air dried; Group II--microwaved at 700W for 40 seconds; Group III--microwaved at 490W for 60 seconds. The percentage weight loss of cylindrical specimens (n = 6) and the percentage dimensional change (n = 7) of die specimens in three axes (x, y and z) were determined at 30 minutes, 1 hour and 24 hours after air drying/microwaving. Weight loss was measured using an electronic digital balance, while dimensional changes were assessed using image analysis software. Data was subject to ANOVA/Scheffe's tests at significance level 0.05. No significant difference in percentage weight loss was observed between air drying for 24 hours and microwaved specimens at all time intervals. Although no significant difference in percentage dimensional changes was observed between specimens microwaved at 490W for 60 seconds and specimens air dried for 24 hours, significant changes in x, y and z dimensions were observed after microwaving at 700W for 40 seconds at various time intervals. Microwave radiation at 490W for 60 seconds is recommended for drying Type IV high-strength dental stone. Further investigations are required to determine changes in physical properties associated with the aforementioned microwave power density.

  2. Drying SDS-Polyacrylamide Gels.

    PubMed

    Sambrook, Joseph; Russell, David W

    2006-09-01

    INTRODUCTIONThis protocol describes a method for drying SDS-polyacrylamide gels. Gels containing proteins radiolabeled with (35)S-labeled amino acids must be dried before autoradiographic images can be obtained. Nonradioactive gels can also be preserved by drying.

  3. Blood Types

    MedlinePlus

    ... Loss Surgery? A Week of Healthy Breakfasts Shyness Blood Types KidsHealth > For Teens > Blood Types Print A A ... or straight hair instead of curly. ...Make Eight Blood Types The different markers that can be found in ...

  4. Blood Sugar

    MedlinePlus

    Blood sugar, or glucose, is the main sugar found in your blood. It comes from the food you eat, and is your body's main source of energy. Your blood carries glucose to all of your body's cells to use ...

  5. Blood Transfusion

    MedlinePlus

    ... notice a decrease in red blood cell levels. Iron overload If you receive multiple blood transfusions, you may end up with too much iron in your blood. Iron overload (hemochromatosis) can damage ...

  6. Donating Blood

    MedlinePlus

    ... can't get an infection or disease from giving blood. The needles and other equipment used are sterile ... part of blood (plasma) within 72 hours after giving blood. It generally takes about 4–8 weeks to ...

  7. Blood Types

    MedlinePlus

    ... cause exceptions to the above patterns. ABO blood typing is not sufficient to prove or disprove paternity or maternity. ... and Complications History of Blood Transfusion Iron and Blood Donation Iron ...

  8. Disinclusion of unerupted teeth by mean of self-ligating brackets: Effect of blood contamination on shear bond strength

    PubMed Central

    Sfondrini, Maria F.; Gatti, Sara; Gandini, Paola

    2013-01-01

    Objectives: The aim of this study was to assess the effect of blood contamination on the shear bond strength and failure site of three different orthodontic self-ligating brackets. Study Design: 240 bovine permanent mandibular incisors were randomly divided into 12 groups of 20 specimens each. Orthodontic self-ligating brackets were tested under four different enamel surface conditions: a) dry, b) blood contamination before priming, c) blood contamination after priming, d) blood contamination before and after priming. Brackets were bonded to the teeth and subsequently tested using a Instron universal testing machine. Shear bond strength values and adhesive failure rate were recorded. Statistical analysis was performed using ANOVA and Tukey tests (strength values), and Chi squared test (ARI Scores). Results: Non-contaminated enamel surfaces showed highest bond strengths for all self ligating brackets. Under blood-contamination shear bond strengths lowered for all brackets tested. Groups contaminated before and after primer application showed the lowest shear bond strength. Significant differences in debond locations were found among the groups under the various enamel surface conditions. Conclusions: Blood contamination of enamel during the bonding procedure lowers bond strength values of self ligating brackets, expecially when contamination occur in different times of the bonding procedure. Key words:Disinclusion, self ligating brackets, blood, contamination, enamel, orthodontics, oral surgery. PMID:23229253

  9. In-vacuum micro-PIXE analysis of biological specimens in frozen-hydrated state

    NASA Astrophysics Data System (ADS)

    Tylko, G.; Mesjasz-Przybyłowicz, J.; Przybyłowicz, W. J.

    2007-07-01

    The first in-vacuum measurements of biological specimens in frozen-hydrated state using proton microbeam have been performed at Materials Research Group, iThemba LABS, South Africa. A commercially available cryotransfer system used in electron microscopy has been adapted for this purpose. The analyzed material was frozen in propane cooled by liquid nitrogen, fractured, carbon coated and transferred onto the cold stage (100 K) in the nuclear microprobe chamber. Micro-PIXE and simultaneous proton backscattering was performed using 3 MeV proton beam. Monitoring of water vapour composition during the proton bombardment showed good stability of the analyzed material. Quantitative results were obtained by standardless method, tested using 20% gelatine standards with added PbCl 2. The differences between the concentrations obtained for frozen-hydrated and next freeze-dried specimens and the value calculated on the basis of weighted mass of PbCl 2 added to gelatine were statistically insignificant. Leaf tissue of Senecio anomalochrous and larvae of Chysolina pardalina were used as examples of plant and animal tissue. Quantitative elemental mapping of frozen-hydrated specimens compared with subsequent analysis of the same areas after freeze-drying revealed the same distribution pattern in both cases.

  10. Diagnostic dry bone histology in human paleopathology.

    PubMed

    de Boer, H H Hans; Van der Merwe, A E Lida

    2016-10-01

    Paleopathology is the study of trauma and disease as may be observed in ancient (human) remains. In contrast to its central role in current medical practice, microscopy plays a rather modest role in paleopathology. This is at least partially due to the differences between fresh and decomposed (i.e., skeletonized or "dry bone") tissue samples. This review discusses these differences and describes how they affect the histological analysis of paleopathological specimens. First, we provide a summary of some general challenges related to the histological analysis of palaeopathological specimens. Second, the reader is introduced in bone tissue histology and bone tissue dynamics. The remainder of the paper is dedicated to the diagnostic value of dry bone histology. Its value and limitations are illustrated by comparing several well-studied paleopathological cases with similar contemporary, clinical cases. This review illustrates that due to post-mortem loss of soft tissue, a limited number of disorders display pathognomonic features during histological analysis of skeletonized human remains. In the remainder of cases, histology may help to narrow down the differential diagnosis or is diagnostically unspecific. A comprehensive, multidisciplinary diagnostic approach therefore remains essential. Clin. Anat. 29:831-843, 2016. © 2016 Wiley Periodicals, Inc.

  11. Dissolution of bulk specimens of silicon nitride

    NASA Technical Reports Server (NTRS)

    Davis, W. F.; Merkle, E. J.

    1981-01-01

    An accurate chemical characterization of silicon nitride has become important in connection with current efforts to incorporate components of this material into advanced heat engines. However, there are problems concerning a chemical analysis of bulk silicon nitride. Current analytical methods require the pulverization of bulk specimens. A pulverization procedure making use of grinding media, on the other hand, will introduce contaminants. A description is given of a dissolution procedure which overcomes these difficulties. It has been found that up to at least 0.6 g solid pieces of various samples of hot pressed and reaction bonded silicon nitride can be decomposed in a mixture of 3 mL hydrofluoric acid and 1 mL nitric acid overnight at 150 C in a Parr bomb. High-purity silicon nitride is completely soluble in nitric acid after treatment in the bomb. Following decomposition, silicon and hydrofluoric acid are volatilized and insoluble fluorides are converted to a soluble form.

  12. A Biorepository for Ophthalmic Surgical Specimens

    PubMed Central

    Skeie, Jessica M.; Tsang, Stephen H.; Zande, Ryan Vande; Fickbohm, Macy M.; Shah, Shaival S.; Vallone, John G.; Mahajan, Vinit B.

    2014-01-01

    Biorepositories are collections of surgically obtained human tissues for current and future investigations of disease mechanisms, therapeutics, and diagnostics. In ophthalmology, a critical challenge is how to interface the operating room with the laboratory. To attain standards required for basic research, clinical and research teams must cooperate to collect, annotate, and store specimens that yield consistent results required for advanced molecular techniques. We developed an efficient platform for obtaining vitreous and other eye tissues from the operating room and transferring them to the lab. The platform includes a mobile lab cart for on-site tissue processing, a multi-user, web-based database for point-of-care phenotypic capture, and an integrated data tracking system for long-term storage. These biorepository instruments have proven essential for our studies in ophthalmic disease proteomics. This system can be implemented in other operating rooms and laboratories for a variety of biological tissues. PMID:24115637

  13. Recording and submitting specimen history data

    USGS Publications Warehouse

    Franson, J. Christian

    1987-01-01

    Webster defines history as "a chronological record of significant events." In wildlife disease investigations, determining the history or background of a problem is the first significant step in establishing a diagnosis. You can greatly assist the diagnostic process by providing a thorough history with specimens yo submit. This information is also of value in understanding the natural history of disease outbreaks, and is difficult if not impossible to obtain after the event has occurred. Detailed field observations during the course of a die-off and investigation of significant events preceding it also provide valuable information on which to base corrective actions. Remember, the most helpful information is that which obtained at the time of the event by a sensitive and aware observer.

  14. Tensile test of dumbbell-shaped specimen in thickness direction

    NASA Astrophysics Data System (ADS)

    Iizuka, Takashi

    2016-10-01

    Sheet metal forming is widely used in manufacturing shops, and evaluation of forming limit for sheet metal is important. However, specimen shape influe