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

PubMed

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

2014-01-01

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

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

PubMed

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

2007-01-01

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

Variations in the detection of anti-PEDV antibodies in serum samples using three diagnostic tests - short communication.

PubMed

Plut, Jan; Toplak, Ivan; Štukelj, Marina

2018-06-01

Over the last few years several porcine epidemic diarrhoea (PED) outbreaks have been discovered in Europe including the first PED case in Slovenia in January 2015. The aim of this study was to determine when PED virus (PEDV) infection started in Slovenia. Serum samples collected between 2012 and 2016 were tested. Three hundred and seventy-five serum samples were collected from 132 Slovenian small, one-site pig farms. Samples were tested for PEDV antibodies utilising three different serological methods: commercially-available indirect ELISA, in-house blocking ELISA test and Immunoperoxidase Monolayer Assay (IPMA) test. One hundred and seventy (45.33%) tested samples were found positive by the commercially-available ELISA test kit, and 10 (5.68%) of these 170 samples found positive were positive by the in-house blocking ELISA. Only these 10 samples were collected from a farm where clinical signs of PED infection had been observed and PEDV was confirmed by RT-PCR methodology; the other 160 samples were collected randomly. Thirty-two samples with the highest S/P value obtained with the commercial ELISA were all negative with IPMA. Reasons for the high variance in the results obtained remain unclear; more research is required to ensure higher sensitivity and specificity in terms of PEDV antibody tests and other PED diagnostic methods.

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

PubMed Central

Chaya, DR; Parija, Subhash Chandra

2014-01-01

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

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

PubMed

Chaya, Dr; Parija, Subhash Chandra

2014-01-01

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

Lack of Serologic Evidence of Neospora caninum in Humans, England

PubMed Central

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

2008-01-01

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

Serological tests for detecting Rift Valley fever viral antibodies in sheep from the Nile Delta.

PubMed Central

Scott, R M; Feinsod, F M; Allam, I H; Ksiazek, T G; Peters, C J; Botros, B A; Darwish, M A

1986-01-01

To determine the accuracy of serological methods in detecting Rift Valley fever (RVF) viral antibodies, we examined serum samples obtained from 418 sheep in the Nile Delta by using five tests. The plaque reduction neutralization test (PRNT) was considered the standard serological method against which the four other tests were compared. Twenty-four serum samples had RVF viral antibodies detected by PRNT. Hemagglutination inhibition and enzyme-linked immunosorbent assay antibodies to RVF virus were also present in the same 24 serum samples. Indirect immunofluorescence was less sensitive in comparison with PRNT, and complement fixation was the least sensitive. These results extend observations made with laboratory animals to a large field-collected group of Egyptian sheep. PMID:3533977

Comparison of Performance Characteristics of Aspergillus PCR in Testing a Range of Blood-Based Samples in Accordance with International Methodological Recommendations.

PubMed

Springer, Jan; White, P Lewis; Hamilton, Shanna; Michel, Denise; Barnes, Rosemary A; Einsele, Hermann; Löffler, Juergen

2016-03-01

Standardized methodologies for the molecular detection of invasive aspergillosis (IA) have been established by the European Aspergillus PCR Initiative for the testing of whole blood, serum, and plasma. While some comparison of the performance of Aspergillus PCR when testing these different sample types has been performed, no single study has evaluated all three using the recommended protocols. Standardized Aspergillus PCR was performed on 423 whole-blood pellets (WBP), 583 plasma samples, and 419 serum samples obtained from hematology patients according to the recommendations. This analysis formed a bicenter retrospective anonymous case-control study, with diagnosis according to the revised European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) consensus definitions (11 probable cases and 36 controls). Values for clinical performance using individual and combined samples were calculated. For all samples, PCR positivity was significantly associated with cases of IA (for plasma, P = 0.0019; for serum, P = 0.0049; and for WBP, P = 0.0089). Plasma PCR generated the highest sensitivity (91%); the sensitivities for serum and WBP PCR were 80% and 55%, respectively. The highest specificity was achieved when testing WBP (96%), which was significantly superior to the specificities achieved when testing serum (69%, P = 0.0238) and plasma (53%, P = 0.0002). No cases were PCR negative in all specimen types, and no controls were PCR positive in all specimens. This study confirms that Aspergillus PCR testing of plasma provides robust performance while utilizing commercial automated DNA extraction processes. Combining PCR testing of different blood fractions allows IA to be both confidently diagnosed and excluded. A requirement for multiple PCR-positive plasma samples provides similar diagnostic utility and is technically less demanding. Time to diagnosis may be enhanced by testing multiple contemporaneously obtained sample types. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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

PubMed Central

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

2017-01-01

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

Development and evaluation of a recombinant-glycoprotein-based latex agglutination test for rabies virus antibody assessment.

PubMed

Jemima, Ebenezer Angel; Manoharan, Seeralan; Kumanan, Kathaperumal

2014-08-01

The measurement of neutralizing antibodies induced by the glycoprotein of rabies virus is indispensable for assessing the level of neutralizing antibodies in animals or humans. A rapid fluorescent focus inhibition test (RFFIT) has been approved by WHO and is the most widely used method to measure the virus-neutralizing antibody content in serum, but a rapid test system would be of great value to screen large numbers of serum samples. To develop and evaluate a latex agglutination test (LAT) for measuring rabies virus antibodies, a recombinant glycoprotein was expressed in an insect cell system and purified, and the protein was coated onto latex beads at concentrations of 0.1, 0.25, 0.5, 0.75, and 1 mg/ml to find out the optimal concentration for coating latex beads. It was found that 0.5 mg/ml of recombinant protein was optimal for coating latex beads, and this concentration was used to sensitize the latex beads for screening of dog serum samples. Grading of LAT results was done with standard reference serum with known antibody titers. A total of 228 serum samples were tested, out of which 145 samples were positive by both RFFIT and LAT, and the specificity was found to be 100 %. In RFFIT, 151 samples were positive, the sensitivity was found to be 96.03 %, and the accuracy/concordance was found to be 97.39 %. A rapid field test-a latex agglutination test (LAT)-was developed and evaluated for rabies virus antibody assessment using recombinant glycoprotein of rabies virus expressed in an insect cell system.

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

USGS Publications Warehouse

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

2015-01-01

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

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

PubMed

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

2015-04-01

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

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

PubMed

Duque, A; Martínez, P-J; Giraldo, A; Gualtero, D-F; Ardila, C-M; Contreras, A; Duarte, S; Lafaurie, G-I

2017-07-01

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

Simultaneous detection of antibodies to five Actinobacillus pleuropneumoniae serovars using bead-based multiplex analysis.

PubMed

Berger, Sanne Schou; Lauritsen, Klara Tølbøll; Boas, Ulrik; Lind, Peter; Andresen, Lars Ole

2017-11-01

We developed and made a preliminary validation of a bead-based multiplexed immunoassay for simultaneous detection of porcine serum antibodies to Actinobacillus pleuropneumoniae serovars 1, 2, 6, 7, and 12. Magnetic fluorescent beads were coupled with A. pleuropneumoniae antigens and tested with a panel of serum samples from experimentally infected pigs and with serum samples from uninfected and naturally infected pigs. The multiplex assay was compared to in-house ELISAs and complement fixation (CF) tests, which have been used for decades as tools for herd classification in the Danish Specific Pathogen Free system. Assay specificities and sensitivities as well as the corresponding cutoff values were determined using receiver operating characteristic (ROC) curve analysis, and the A. pleuropneumoniae multiplex assay showed good correlation with the in-house ELISAs and CF tests with areas under ROC curves ≥ 0.988. Benefits of multiplexed assays compared to ELISAs and CF tests include reduced serum sample volumes needed for analysis, less labor, and shorter assay time.

Analytical Comparison of In Vitro-Spiked Human Serum and Plasma for PCR-Based Detection of Aspergillus fumigatus DNA: a Study by the European Aspergillus PCR Initiative

PubMed Central

Mengoli, Carlo; Springer, Jan; Bretagne, Stéphane; Cuenca-Estrella, Manuel; Klingspor, Lena; Lagrou, Katrien; Melchers, Willem J. G.; Morton, C. Oliver; Barnes, Rosemary A.; Donnelly, J. Peter; White, P. Lewis

2015-01-01

The use of serum or plasma for Aspergillus PCR testing facilitates automated and standardized technology. Recommendations for serum testing are available, and while serum and plasma are regularly considered interchangeable for use in fungal diagnostics, differences in galactomannan enzyme immunoassay (GM-EIA) performance have been reported and are attributed to clot formation. Therefore, it is important to assess plasma PCR testing to determine if previous recommendations for serum are applicable and also to compare analytical performance with that of serum PCR. Molecular methods testing serum and plasma were compared through multicenter distribution of quality control panels, with additional studies to investigate the effect of clot formation and blood fractionation on DNA availability. Analytical sensitivity and time to positivity (TTP) were compared, and a regression analysis was performed to identify variables that enhanced plasma PCR performance. When testing plasma, sample volume, preextraction-to-postextraction volume ratio, PCR volume, duplicate testing, and the use of an internal control for PCR were positively associated with performance. When whole-blood samples were spiked and then fractionated, the analytical sensitivity and TTP were superior when testing plasma. Centrifugation had no effect on DNA availability, whereas the presence of clot material significantly lowered the concentration (P = 0.028). Technically, there are no major differences in the molecular processing of serum and plasma, but the formation of clot material potentially reduces available DNA in serum. During disease, Aspergillus DNA burdens in blood are often at the limits of PCR performance. Using plasma might improve performance while maintaining the methodological simplicity of serum testing. PMID:26085614

Analytical Comparison of In Vitro-Spiked Human Serum and Plasma for PCR-Based Detection of Aspergillus fumigatus DNA: a Study by the European Aspergillus PCR Initiative.

PubMed

Loeffler, Juergen; Mengoli, Carlo; Springer, Jan; Bretagne, Stéphane; Cuenca-Estrella, Manuel; Klingspor, Lena; Lagrou, Katrien; Melchers, Willem J G; Morton, C Oliver; Barnes, Rosemary A; Donnelly, J Peter; White, P Lewis

2015-09-01

The use of serum or plasma for Aspergillus PCR testing facilitates automated and standardized technology. Recommendations for serum testing are available, and while serum and plasma are regularly considered interchangeable for use in fungal diagnostics, differences in galactomannan enzyme immunoassay (GM-EIA) performance have been reported and are attributed to clot formation. Therefore, it is important to assess plasma PCR testing to determine if previous recommendations for serum are applicable and also to compare analytical performance with that of serum PCR. Molecular methods testing serum and plasma were compared through multicenter distribution of quality control panels, with additional studies to investigate the effect of clot formation and blood fractionation on DNA availability. Analytical sensitivity and time to positivity (TTP) were compared, and a regression analysis was performed to identify variables that enhanced plasma PCR performance. When testing plasma, sample volume, preextraction-to-postextraction volume ratio, PCR volume, duplicate testing, and the use of an internal control for PCR were positively associated with performance. When whole-blood samples were spiked and then fractionated, the analytical sensitivity and TTP were superior when testing plasma. Centrifugation had no effect on DNA availability, whereas the presence of clot material significantly lowered the concentration (P = 0.028). Technically, there are no major differences in the molecular processing of serum and plasma, but the formation of clot material potentially reduces available DNA in serum. During disease, Aspergillus DNA burdens in blood are often at the limits of PCR performance. Using plasma might improve performance while maintaining the methodological simplicity of serum testing. Copyright © 2015 Loeffler et al.

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

PubMed Central

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

2013-01-01

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

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

PubMed

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

2013-01-01

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

Serologic Evidence of Lyssavirus Infection in Bats, Cambodia

PubMed Central

Molia, Sophie; Audry, Laurent; Hout, Sotheara; Ngin, Sopheak; Walston, Joe; Bourhy, Hervé

2004-01-01

In Cambodia, 1,303 bats of 16 species were tested for lyssavirus. No lyssavirus nucleocapsid was detected in 1,283 brains tested by immunofluorescence assay. Antibodies against lyssaviruses were detected by enzyme-linked immunosorbent assay in 144 (14.7%) of 981 serum samples. Thirty of 187 serum samples contained neutralizing antibodies against different lyssaviruses. PMID:15663870

Performances of fourth generation HIV antigen/antibody assays on filter paper for detection of early HIV infections.

PubMed

Kania, Dramane; Truong, Tam Nguyen; Montoya, Ana; Nagot, Nicolas; Van de Perre, Philippe; Tuaillon, Edouard

2015-01-01

Point-of-care testing and diagnosis of HIV acute infections play important roles in preventing transmission, but HIV rapid diagnosis tests have poor capacity to detect early infections. Filter paper can be used for capillary blood collection and HIV testing using 4th generation immunoassays. Antigen/antibody combined immunoassays were evaluated for their capacity to identify early HIV infections using filter paper in comparison with rapid test. Thirty nine serum samples collected from HIV seroconverters were spotted onto filter paper and tested by the Roche Elecsys(®) HIV Combi PT test and the DiaSorin Liaison XL Murex HIV Ab/Ag assay. Fourth generation immunoassays identified 34 out of 39 HIV early infections using dried serum spot, whereas the Determine™ HIV-1/2 rapid test detected 24 out of 39 HIV positive serum (87.2% vs 61.5% respectively, p = 0.009). p24 antigen was detected by the Liaison XL in 19 dried serum samples (48.7%). In the group characterized by a negative western blot, 7 out of 8 (87.5%) and 6 out of 8 (75.0%) samples were found positive for HIV using the Elecsys and the Liaison XL, respectively. None of these eight samples classified in this group of early acute infections were found positive by the rapid test. Fourth generation Ag/Ab immunoassays performed on dried serum spot had good performance for HIV testing during the early phases of HIV infection. This method may be useful to detect HIV early infections in hard-to-reach populations and individuals living in remote areas before rapid tests become positive. Copyright © 2014 Elsevier B.V. All rights reserved.

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

PubMed

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

2012-04-30

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

Radioimmunoassays for the serum thymic factor (FTS)

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

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

1987-01-06

This patent describes a radioimmunoassay of serum thymic factor (FTS) in a test sample, comprising: (a) contacting a first aliquot of the sample with anti-FTS antibody, with a known amount of FTS hormone standard and a known amount of radiolabeled FTS analogue; and (b) contacting a second aliquot of the sample with anti-FTS antibody and a known amount of radiolabeled FTS analogue; and (c) measuring the radioactivity of the antigen-antibody complex in each aliquot; and (d) calculating the amount of FTS in the test sample.

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

PubMed

Liu, Zhenjiang; Yuan, Zhengwei; Zhao, Qun

2014-01-01

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

Bicentric evaluation of six anti-toxoplasma immunoglobulin G (IgG) automated immunoassays and comparison to the Toxo II IgG Western blot.

PubMed

Maudry, Arnaud; Chene, Gautier; Chatelain, Rémi; Patural, Hugues; Bellete, Bahrie; Tisseur, Bernard; Hafid, Jamal; Raberin, Hélène; Beretta, Sophie; Sung, Roger Tran Manh; Belot, Georges; Flori, Pierre

2009-09-01

A comparative study of the Toxoplasma IgG(I) and IgG(II) Access (Access I and II, respectively; Beckman Coulter Inc.), AxSYM Toxo IgG (AxSYM; Abbott Diagnostics), Vidas Toxo IgG (Vidas; bioMerieux, Marcy l'Etoile, France), Immulite Toxo IgG (Immulite; Siemens Healthcare Diagnostics Inc.), and Modular Toxo IgG (Modular; Roche Diagnostics, Basel, Switzerland) tests was done with 406 consecutive serum samples. The Toxo II IgG Western blot (LDBio, Lyon, France) was used as a reference technique in the case of intertechnique discordance. Of the 406 serum samples tested, the results for 35 were discordant by the different techniques. Using the 175 serum samples with positive results, we evaluated the standardization of the titrations obtained (in IU/ml); the medians (second quartiles) obtained were 9.1 IU/ml for the AxSYM test, 21 IU/ml for the Access I test, 25.7 IU/ml for the Access II test, 32 IU/ml for the Vidas test, 34.6 IU/ml for the Immulite test, and 248 IU/ml for the Modular test. For all the immunoassays tested, the following relative sensitivity and specificity values were found: 89.7 to 100% for the Access II test, 89.7 to 99.6% for the Immulite test, 90.2 to 99.6% for the AxSYM test, 91.4 to 99.6% for the Vidas test, 94.8 to 99.6% for the Access I test, and 98.3 to 98.7% for the Modular test. Among the 406 serum samples, we did not find any false-positive values by two different tests for the same serum sample. Except for the Modular test, which prioritized sensitivity, it appears that the positive cutoff values suggested by the pharmaceutical companies are very high (either for economical or for safety reasons). This led to imperfect sensitivity, a large number of unnecessary serological follow-ups of pregnant women, and difficulty in determining the serological status of immunosuppressed individuals.

A Rapid Dot Immunoassay for the Detection of Serum Antibodies to Eastern Equine Encephalomyelitis and Saint Louis Encephalitis Viruses in Sentinel Chickens

DTIC Science & Technology

1988-01-01

tion inhibition (HI) tests .6 uses a novel membrane, polyvinylydine difluor- EEE and SLE viruses are significant public ide (PVDF), and yields a...only 3 aged values between the two tests (Fig. 3). Vari- of 6 SLE virus-inoculated birds developed an- ability in EIA titer was observed with samples... SLE samples negative by EIA were location. Four serum samples from each of 3 also negative by DotIA. EEE-positive chickens were tested by EIA and A

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

PubMed

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

2016-10-01

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

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

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

PubMed

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

2014-02-01

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

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

PubMed

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

2004-12-01

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

Nucleic Acid-Based Cross-Linking Assay for Detection and Quantification of Hepatitis B Virus DNA

PubMed Central

Lai, Vicky C. H.; Guan, Richard; Wood, Michael L.; Lo, Su Kong; Yuen, Man-Fung; Lai, Ching-Lung

1999-01-01

A nucleic acid photo-cross-linking technology was used to develop a direct assay for the quantification of hepatitis B virus (HBV) DNA levels in serum. Cross-linker-modified DNA probes complementary to the viral genomes of the major HBV subtypes were synthesized and used in an assay that could be completed in less than 6 h. The quantification range of the assay, as determined by testing serial dilutions of Eurohep HBV reference standards and cloned HBV DNA, was 5 × 105 to 3 × 109 molecules of HBV DNA/ml of serum. Within-run and between-run coefficients of variation (CVs) for the assay were 4.3 and 4.0%, respectively. The assay was used to determine HBV DNA levels in 302 serum samples, and the results were compared to those obtained after testing the same samples with the Chiron branched-DNA (bDNA) assay for HBV DNA. Of the samples tested, 218 were positive for HBV DNA by both assays and 72 gave results below the cutoff for both assays. Of the remaining 12 samples, 10 were positive for HBV DNA by the cross-linking assay only; the 2 other samples were positive by the bDNA assay only. Twenty-eight samples had to be retested by the bDNA assay (CV, >20% between the results obtained from the testing of each sample in duplicate), whereas only three samples required retesting by the cross-linking assay. The correlation between the HBV DNA levels, as measured by the two tests, was very high (r = 0.902; P = 0.01). We conclude that the cross-linking assay is a sensitive and reproducible method for the detection and quantification of HBV DNA levels in serum. PMID:9854083

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

PubMed

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

2006-12-01

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

Bayesian validation of a serum and milk ELISA for antibodies against Mycobacterium avium subspecies paratuberculosis in Greek dairy goats across lactation.

PubMed

Angelidou, E; Kostoulas, P; Leontides, L

2014-02-01

We validated a commercial (Idexx Pourquier, Montpellier, France) serum and milk indirect ELISA that detects antibodies against Mycobacterium avium ssp. paratuberculosis (MAP) in Greek dairy goats. Each goat was sampled 4 times, starting from kidding and covering early, mid, and late lactation. A total of 1,268 paired milk (or colostrum) and serum samples were collected during the 7-mo lactation period. Bayesian latent class models, which allow for the continuous interpretation of test results, were used to derive the distribution of the serum and milk ELISA response for healthy and MAP-infected individuals at each lactation stage. Both serum and milk ELISA, in all lactation stages, had average and similar overall discriminatory ability as measured by the area under the curve (AUC). For each test, the smallest overlap between the distribution of the healthy and MAP-infected does was in late lactation. At this stage, the AUC was 0.89 (95% credible interval: 0.70; 0.98) and 0.92 (0.74; 0.99) for the milk and serum ELISA, respectively. Both tests had comparable sensitivities and specificities at the recommended cutoffs across lactation. Lowering the cutoffs led to an increase in sensitivity without serious loss in specificity. In conclusion, the milk ELISA was as accurate as the serum ELISA. Therefore, it could serve as the diagnostic tool of choice, especially during the implementation of MAP control programs that require frequent testing, because milk sampling is a noninvasive, rapid, and easy process. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

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

PubMed

Howe, S E; Lynch, D M

1985-05-01

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

Detection of antibodies to egg drop syndrome virus in chicken serum using a field-based immunofiltration (flow-through) test.

PubMed

Raj, G Dhinakar; Thiagarajan, V; Nachimuthu, K

2007-09-01

A simple, user-friendly, and rapid method to detect the presence of antibodies to egg drop syndrome 76 (EDS) virus in chicken sera based on an immunofiltration (flow-through) test was developed. Purified EDS virus antigen was coated onto nitrocellulose membranes housed in a plastic module with layers of absorbent filter pads underneath. Following addition of serum to be tested and washing, monoclonal antibodies or polyclonal serum to chicken immunoglobulin G (IgG) was used as a bridge antibody to mediate binding between EDS virus-specific IgG and protein A gold conjugate. The appearance of a pink dot indicated the presence of antibodies to EDS virus in the sample tested. The results could be obtained within 5-10 min. The developed immunofiltration test could detect antibodies in the sera of experimentally vaccinated chickens from 2 wk postvaccination. With field sera samples, this test was positive in samples having hemagglutination inhibition titers of 8 and above. This test has the potential to be used as a field-based kit to assess seroconversion in EDS-vaccinated flocks.

51Cr-EDTA absorption blood test: an easy method for assessing small intestinal permeability in dogs.

PubMed

Frias, Rafael; Sankari, Satu; Westermarck, Elias

2004-01-01

The 51Cr-EDTA test is a valuable clinical tool for screening intestinal diseases in dogs. The test is performed by calculating the percentage of recovery from urine of a PO-ingested dose of 51Cr-EDTA after 6 or 24 hours. Careful urine collection is a practical limitation of this test in dogs, and our goal was to develop a simpler test that measures 51Cr-EDTA in blood. A 51Cr-EDTA absorption test was simultaneously performed on urine and serum 43 times in healthy Beagle Dogs. Timed blood samples were withdrawn, and urine was collected during a 6-hour period. Percentages of the ingested dose were then calculated in urine and serum. The mean +/- standard deviation (range) percentage in urine after 6 hours was 14.07 +/- 8.72% (3.81-34.18%), whereas results in serum from samples taken at 2, 3, 4, 5, and 6 hours were 0.49 +/- 0.45% (0.02-2.13%), 0.75 +/- 0.52% (0.03-1.89%), 0.82 +/- 0.57% (0.13-2.21%), 0.70 +/- 0.53% (0.12-1.99%), and 0.47 +/- 0.44% (0.11-1.79%), respectively. The results for blood specimens showed good concordance with those for urine, especially for the samples taken at 4 hours (r = 0.89). Moreover, the correlation between urine and blood was better when the sum of the percentages of the recovered analyte from various blood samples was compared with urine. The correlation coefficient when summing 4 blood samples was excellent (r = 0.97) and remained excellent when summing only 2 blood samples taken at 3 and 5 hours (r = 0.95) or at 3 and 4 hours (r = 0.94). We conclude that a serum 51Cr-EDTA test determined by summing successive blood samples provides an easier means of estimating small intestinal permeability in dogs and gives results comparable to those of the 6-hour urine test.

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

PubMed

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

2012-11-01

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

[The comparative study of specificity of test-systems in diagnostic of HIV-infection on categories of samples of blood serum of pregnant women].

PubMed

Sharipova, I N; Khodak, N M; Puzirev, V F; Burkov, A N; Ulanova, T I

2015-03-01

The detection of false positive serological reactions (FPSR) on HIV-infection under screening examination of pregnant women is an actual problem of practical health care. The original observations testify that under analysis of the same samples of blood serum of pregnant women using screening immune enzyme test-systems of various manufacturers the unmatched data concerning FPSR can be obtained. The purpose of this study was to implement comparative evaluation of specificity of immune enzyme test-systems of three different manufacturers: "DS-IFA-HIV-AGAT-SCREEN" ("Diagnostic Systems"), "Genscreen Ultra HIV Ag-Ab" "Bio Rad" France) and "The CombiBest HIV-1,2 AG/AT" ("Vector-Best" Novosibirsk). The sampling of 440 samples of blood serums of pregnant women from various medical institutions of Nizhnii Novgorod was analyzed. The results of the study demonstrated that FPSR were detected in all test-systems and at that spectrum of samples differed. The identical specificity of compared test-systems amounted to 98.64%. The alternative approach to FPSR to HIV issue under screening examinations of pregnant women was proposed. The proposed mode consisted of consistent application of two test-systems of fourth generation with different format of setup of reaction.

9 CFR 113.309 - Bovine Parainfluenza3 Vaccine.

Code of Federal Regulations, 2010 CFR

2010-01-01

... develop antibody titers of 1:32 or greater by day 6 ±2 days post-challenge. (8) A sequential test... parainfluenza, susceptible calves shall be used as test animals (20 vaccinates and five controls). Blood samples... negative at a 1:2 final serum dilution in a varying serum constant virus neutralization test with less than...

9 CFR 113.309 - Bovine Parainfluenza3 Vaccine.

Code of Federal Regulations, 2012 CFR

2012-01-01

... develop antibody titers of 1:32 or greater by day 6 ±2 days post-challenge. (8) A sequential test... parainfluenza, susceptible calves shall be used as test animals (20 vaccinates and five controls). Blood samples... negative at a 1:2 final serum dilution in a varying serum constant virus neutralization test with less than...

Use of Paired Serum Samples for Serodiagnosis of Typhoid Fever

PubMed Central

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

2005-01-01

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

Evaluation of fecal elastase and serum cholecystokinin in dogs with a false positive fecal elastase test.

PubMed

Steiner, J M; Rehfeld, J F; Pantchev, N

2010-01-01

An assay for the measurement of pancreatic elastase in dog feces has been introduced. The goal of this study was to evaluate the rate of false-positive fecal-elastase test results in dogs with suspected exocrine pancreatic insufficiency (EPI) and to assess serum cholecystokinin (CCK) concentrations in dogs with a false positive fecal elastase test result. Twenty-six fecal and serum samples from dogs suspected of EPI, for which samples had been submitted to a commercial laboratory (Vet Med Labor) for analysis. Prospective study. Serum trypsin-like immunoreactivity (TLI) was measured in 26 dogs with a decreased fecal elastase concentration of <10 microg/g feces. Serum CCK concentrations were measured in 21 of these dogs. Of 26 dogs with a decreased fecal elastase concentration, 6 (23%) had serum TLI concentrations within or above the reference range. Serum CCK concentrations were significantly higher in dogs with a true positive fecal elastase test result (median: 1.1 pmol/L; range: 0.1-3.3 pmol/L) than in those with a false positive fecal elastase test result (median: 0.1 pmol/L; range: 0.1-0.9 pmol/L; P value = .0163). The rate of false positive fecal elastase test results was high in this group of dogs, suggesting that diagnosis of EPI must be confirmed by other means. The decreased CCK concentration in dogs with a false positive fecal elastase test result could suggest that false positive results are because of decreased stimulation of exocrine pancreatic function caused by other conditions.

Amoeba-Resisting Bacteria and Ventilator-Associated Pneumonia

PubMed Central

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

2003-01-01

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

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

PubMed

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

2013-01-01

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

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

PubMed

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

2014-01-01

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

Comparison of the serum and salivary antibodies to detect gastric Helicobacter pylori infection in Kashan (Iran).

PubMed

Piroozmand, Ahmad; Soltani, Babak; Razavizadeh, Mohsen; Matini, Amir Hasan; Gilasi, Hamid Reza; Zavareh, Abbas Nassaji; Soltani, Siamak

2017-12-01

Helicobacter pylori ( H. pylori ) is an important and common contagious human pathogen which may cause peptic ulcer and also gastric cancer. The definite diagnosis of it is made through invasive tests. Recently, non-invasive tests including serologic tests of serum and saliva have been conducted for diagnosis of H. pylori infection. In this research, the diagnostic values of serum and salivary serology were compared together to use salivary anti- H. pylori test as an alternative method in the future. During this prospective case-control study on patients who were candidates for endoscopy and gastric biopsy from March 2015 to April 2016 in Shahid Beheshti hospital, Kashan, Iran, serum and salivary samples were obtained for measurement of Immunoglobulin G (IgG) antibody levels against H. pylori by enzyme-linked immunosorbent assay (ELISA). Histopathology was the gold standard test. Statistical analysis was performed by SPSS software version 16. Statistical tests included Kolmogorov-Smirnov, independent-samples t-test, Chi-square, Mann-Whitney U, Kruskal-Wallis, McNemar and correlation. Of 123 patients, sixty-one patients (49.6%) were H. pylori -positive according to histology. The median levels of anti- H. pylori antibodies in serum (p<0.001) and saliva (p<0.001) of H. pylori -positive cases were significantly higher than H. pylori -negative cases. Sensitivity, specificity, positive likelihood ratio, negative likelihood ratio and accuracy of serologic tests in serum were 75%, 79%, 3.5, 0.3, 77% and for saliva were 85%, 82%, 4.7, 0.18, 84% respectively. Diagnostic values of salivary ELISA are comparable to serum ELISA and can be used as an alternative modality for diagnosis of H. pylori infection.

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

PubMed

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

2015-03-01

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

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

PubMed

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

2014-04-01

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

The implementation of nucleic acid amplification technology testing for living tissue donors.

PubMed

Westby, J; Lomas, R J; Kearney, J N

2010-05-01

There is a significant requirement within the United Kingdom for tissue grafts from living donors. To ensure safety, blood samples from these donors are tested for pathogens at donation, and at 180 days post donation. Nucleic acid amplification technology (NAT) permits more sensitive detection of pathogens in blood samples than serum antigen testing. NAT testing can be applied to samples from living tissue donors to eliminate the need to re-test these donors 180 days post-donation before grafts can be implanted. This has major financial and operational advantages for a tissue bank, and this manuscript describes how NAT testing was assessed and implemented by NHSBT Tissue Services. When compared to traditional serum antigen testing, NAT testing was more cost effective, more convenient for donors and resulted in a greater proportion of donated grafts being made available for transplant.

Effect of non-surgical periodontal treatment on transferrin serum levels in patients with chronic periodontitis

PubMed Central

Shirmohamadi, Adileh; Chitsazi, Mohamad Taghi; Faramarzi, Masoumeh; Salari, Ashkan; Naser Alavi, Fereshteh; Pashazadeh, Nazila

2016-01-01

Background. Transferrin is a negative acute phase protein, which decreases during inflammation and infection. The aim of the present investigation was to evaluate changes in the transferrin serum levels subsequent to non-surgical treatment of chronic periodontal disease. Methods. Twenty patients with chronic periodontitis and 20 systemically healthy subjects without periodontal disease, who had referred to Tabriz Faculty of Dentistry, were selected. Transferrin serum levels and clinical periodontal parameters (pocket depth, clinical attachment level, gingival index, bleeding index and plaque index) were measured at baseline and 3 months after non-surgical periodontal treatment. Data were analyzed with descriptive statistical methods (means ± standard deviations). Independent samples t-test was used to compare transferrin serum levels and clinical variables between the test and control groups. Paired samples t-test was used in the test group for comparisons before and after treatment. Statistical significance was set at P < 0.05. Results. The mean transferrin serum level in patients with chronic periodontitis (213.1 ± 9.2 mg/dL) was significantly less than that in periodontally healthy subjects (307.8 ± 11.7 mg/dL). Three months after periodontal treatment, the transferrin serum level increased significantly (298.3 ± 7.6 mg/dL) and approached the levels in periodontally healthy subjects (P < 0.05). Conclusion. The decrease and increase in transferrin serum levels with periodontal disease and periodontal treatment, respectively, indicated an inverse relationship between transferrin serum levels and chronic periodontitis. PMID:27651883

9 CFR 113.300 - General requirements for live virus vaccines.

Code of Federal Regulations, 2010 CFR

2010-01-01

... control cells shall remain free of such fluorescence. (2) Serum neutralization test. The serum... fungi in accordance with the test provided in § 113.27. (2) Mycoplasma. Final container samples of... inoculated cells and uninoculated control cells. Cells shall be stained with fluorochrome conjugated specific...

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

PubMed Central

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

1998-01-01

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

Investigation of Metabolomic Blood Biomarkers for Detection of Adenocarcinoma Lung Cancer

PubMed Central

Fahrmann, Johannes F.; Kim, Kyoungmi; DeFelice, Brian C.; Taylor, Sandra L.; Gandara, David R.; Yoneda, Ken Y.; Cooke, David T.; Fiehn, Oliver; Kelly, Karen; Miyamoto, Suzanne

2015-01-01

Background Untargeted metabolomics was utilized in case control studies of adenocarcinoma (ADC) lung cancer in order to develop and test metabolite classifiers in serum and plasma as potential biomarkers for diagnosing lung cancer. Methods Serum and plasma were collected and used in two independent case-control studies (ADC1 and ADC2). Controls were frequency matched for gender, age and smoking history. There were 52 ADC cases and 31 controls in ADC1 and 43 ADC cases and 43 controls in ADC2. Metabolomics was conducted using gas chromatography time-of-flight mass spectrometry. Differential analysis was performed on ADC1 and the top candidates (FDR < 0.05) for serum and plasma used to develop individual and multiplex-classifiers that were then tested on an independent set of serum and plasma samples (ADC2). Results Aspartate provided the best accuracy (81.4%) for an individual metabolite classifier in serum whereas pyrophosphate had the best accuracy (77.9%) in plasma when independently tested. Multiplex classifiers of either 2 or 4 serum metabolites had an accuracy of 72.7% when independently tested. For plasma, a multi-metabolite classifier consisting of 8 metabolites gave an accuracy of 77.3% when independently tested. Comparison of overall diagnostic performance between the two blood matrices yielded similar performances. However, serum is most ideal given higher sensitivity for low abundant metabolites. Conclusion This study shows the potential of metabolite-based diagnostic tests for detection of lung adenocarcinoma. Further validation in a larger pool of samples is warranted. Impact These biomarkers could improve early detection and diagnosis of lung cancer. PMID:26282632

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

PubMed Central

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

2015-01-01

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

Detection of Serum microRNAs From Department of Defense Serum Repository

PubMed Central

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

2017-01-01

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

Acute phase proteins in the diagnosis of bovine subclinical mastitis.

PubMed

Safi, Shahabeddin; Khoshvaghti, Ameneh; Jafarzadeh, Seyed Reza; Bolourchi, Mahmoud; Nowrouzian, Iradj

2009-12-01

The California mastitis test (CMT) and somatic cell count (SCC) are commonly used for diagnosis of subclinical mastitis in cattle. Acute phase proteins (APPs), as alternative biomarkers of mastitis, may increase in concentration in the absence of macroscopic changes in the milk, or may precede the onset of clinical signs. The aim of this study was to compare the accuracy of APPs measured in milk and in serum with bacterial culture for the diagnosis of bovine subclinical mastitis. One hundred and seventy-five Holstein cows were randomly selected from 7 dairy farms. Quarter milk and serum samples were taken from all cows. Milk samples were analyzed using a CMT and SCC, and for haptoglobin (MHp) and amyloid A (MAA) concentrations, and were also submitted for bacterial culture. Serum samples obtained concurrently were analyzed for haptoglobin (SHp) and amyloid A (SAA). Two-sample Wilcoxon (Mann-Whitney) test was used to compare SCC, MAA, MHp, SAA, and SHp concentrations between culture-positive and culture-negative animals. Receiver-operating characteristic analysis was used to assess the performance of each test using bacterial culture as the reference method. MAA concentration was the most accurate of the 5 tests, with a sensitivity of 90.6% and specificity of 98.3% at concentrations >16.4 mg/L. MAA and MHp had significantly larger areas under the curve than the respective serum proteins, SAA and SHp. The results suggest that measuring haptoglobin and amyloid A in milk is more accurate than serum analysis for the diagnosis of subclinical mastitis in Holstein cows.

Diagnostic value of serum Golgi protein 73 for HBV-related primary hepatic carcinoma

PubMed Central

Gao, Guosheng; Dong, Feibo; Xu, Xiaozhen; Hu, Airong; Hu, Yaoren

2015-01-01

Background: Alpha-fetoprotein (AFP) levels are routinely used for diagnosis and monitoring of hepatic diseases, but it has a limited value. Golgi protein 73 (GP73) has been suggested as a new marker for hepatic diseases. Objective: To explore the clinical value of serum GP73 in different diseases associated with hepatitis B virus (HBV) infection. Method: Between January 2010 and August 2014, serum samples from 88 patients with chronic hepatitis B (CHB), 78 patients with HBV-related liver cirrhosis (LC), and 194 patients with HBV-related primary hepatic cancer (PHC) were collected. Serum samples from 30 healthy volunteers were used as controls. ELISA and microparticle enzyme immunoassay were used to measure serum GP73 and AFP levels. Receiver operating characteristic (ROC) curves were used to analyze the diagnostic value of serum GP73 and AFP for PHC. Results: For the diagnosis of PHC, GP73 showed a sensitivity of 65.5% and specificity of 66.3%, while AFP levels showed sensitivity of 64.4% and specificity of 76.5%. Serial testing (both tests are positive) could increase the specificity (sensitivity of 45.9% and specificity of 85.5%) while parallel testing (any single positive test result) could increase the sensitivity (sensitivity of 84.0% and specificity of 57.2%). Serum GP73 and AFP levels were significantly different between Child-Pugh grades (P<0.001 for GP73 and P=0.044 for AFP). Significant differences in serum GP73 and AFP were found between TNM stages (all P<0.001). Conclusion: Serum GP73 had limited diagnostic value for HBV-related PHC. The combined use of serum GP73 and AFP levels improved the diagnostic efficacy. PMID:26617863

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

PubMed Central

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

2016-01-01

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

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

PubMed

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

2016-08-01

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

Nipah Virus in Lyle's Flying Foxes, Cambodia

PubMed Central

Counor, Dorian; Ong, Sivuth; Faure, Caroline; Seng, Vansay; Molia, Sophie; Walston, Joe; Georges-Courbot, Marie Claude; Deubel, Vincent; Sarthou, Jean-Louis

2005-01-01

We conducted a survey in Cambodia in 2000 on henipavirus infection among several bat species, including flying foxes, and persons exposed to these animals. Among 1,072 bat serum samples tested by enzyme-linked immunosorbent assay, antibodies reactive to Nipah virus (NiV) antigen were detected only in Pteropus lylei species; Cynopterus sphinx, Hipposideros larvatus, Scotophilus kuhlii, Chaerephon plicata, Taphozous melanopogon, and T. theobaldi species were negative. Seroneutralization applied on a subset of 156 serum samples confirmed these results. None of the 8 human serum samples was NiV seropositive with the seroneutralization test. One virus isolate exhibiting cytopathic effect with syncytia was obtained from 769 urine samples collected at roosts of P. lylei specimens. Partial molecular characterization of this isolate demonstrated that it was closely related to NiV. These results strengthen the hypothesis that flying foxes could be the natural host of NiV. Surveillance of human cases should be implemented. PMID:16022778

Analytical validation of a new point-of-care assay for serum amyloid A in horses.

PubMed

Schwartz, D; Pusterla, N; Jacobsen, S; Christopher, M M

2018-01-17

Serum amyloid A (SAA) is a major acute phase protein in horses. A new point-of-care (POC) test for SAA (Stablelab) is available, but studies evaluating its analytical accuracy are lacking. To evaluate the analytical performance of the SAA POC test by 1) determining linearity and precision, 2) comparing results in whole blood with those in serum or plasma, and 3) comparing POC results with those obtained using a previously validated turbidimetric immunoassay (TIA). Assay validation. Analytical validation of the POC test was done in accordance with American Society of Veterinary Clinical Pathology guidelines using residual equine serum/plasma and whole blood samples from the Clinical Pathology Laboratory at the University of California-Davis. A TIA was used as the reference method. We also evaluated the effect of haematocrit (HCT). The POC test was linear for SAA concentrations of up to at least 1000 μg/mL (r = 0.991). Intra-assay CVs were 13, 18 and 15% at high (782 μg/mL), intermediate (116 μg/mL) and low (64 μg/mL) concentrations. Inter-assay (inter-batch) CVs were 45, 14 and 15% at high (1372 μg/mL), intermediate (140 μg/mL) and low (56 μg/mL) concentrations. SAA results in whole blood were significantly lower than those in serum/plasma (P = 0.0002), but were positively correlated (r = 0.908) and not affected by HCT (P = 0.261); proportional negative bias was observed in samples with SAA>500 μg/mL. The difference between methods exceeded the 95% confidence interval of the combined imprecision of both methods (15%). Analytical validation could not be performed in whole blood, the sample most likely to be used stall side. The POC test has acceptable accuracy and precision in equine serum/plasma with SAA concentrations of up to at least 1000 μg/mL. Low inter-batch precision at high concentrations may affect serial measurements, and the use of the same test batch and sample type (serum/plasma or whole blood) is recommended. Comparison of results between the POC test and the TIA is not recommended. © 2018 EVJ Ltd.

Reliability of the cloned-enzyme donor immunoassay (CEDIA) for cocaine in human serum in the range between the detection limit and the cut-off.

PubMed

Schütz, Harald; Auch, Jürgen; Erdmann, Freidoon; Weiler, Günter; Verhoff, Marcel A

2006-01-01

Immunoassays are used worldwide for the rapid screening of drugs. Despite the fact that they are highly valuable tools for the testing of legal and illicit drugs, there is a non-negligable risk of false-positive and false-negative findings and many pitfalls must be taken into account when using these tests in an uncritical manner and without valid confirmation procedures. In order to check the correlation between cloned-enzyme donor immunoassay (CEDIA) readings and exact determined gas chromatography/mass spectrometry (GC/MS) values for benzoylecgonine, a total of 472 serum samples was measured with an immunoassay (CEDIA) as well as GC/MS. As a result, it was shown that in the lower area of concentration, up to approx. 300 ng equ. benzoylecgonine/ml, there is a semiquantitative useful correlation. With higher concentrations, the values shift more and more apart and many runaways are visible. Generally the use of tests for materials other than those declared is excluded from the manufacturer's warranty and is at the user's own risk, but the authors found that many immunochemical tests with a certification only for the use in urine are well suited to the screening of serum samples, and even low readings could be confirmed by GC/MS. With blood or serum as sample, it could be regularly observed that generally very small readings could not only be confirmed with mass spectrometry but could in most cases also be quantified. This was the key factor in determining the rate of confirmation in 495 cases. In total, an incredibly high reliability for the CEDIA cocaine test (via benzoylecgonine) was shown up to the area of detection (detection limit), far below the cut-off. These results confirm the extraordinary sensitivity and specificity of this test also for blood or serum as sample material to be tested.

The Prevalence of Brucellosis in Cattle, Goats and Humans in Rural Uganda: A Comparative Study.

PubMed

Miller, R; Nakavuma, J L; Ssajjakambwe, P; Vudriko, P; Musisi, N; Kaneene, J B

2016-12-01

A cross-sectional study was conducted to determine the presence of brucellosis in cattle, goats and humans in farms from south-western Uganda and identify risk factors associated with brucellosis in these three host groups. Data and serum samples were collected from 768 cattle, 315 goats and 236 humans, with 635 samples of bovine milk, from 70 farms in two different study areas in south-western Uganda. Sera from livestock were tested with the Rose Bengal Plate test, using B. abortus and B. melitensis antigens, and human sera were tested with a commercial IgG/IgM lateral flow assay. Milk samples were tested using the OIE-approved milk ring test. Screening tests for brucellosis were positive in 14% of cattle serum, 29% of bovine milk, 17% of goat serum and 11% of human serum samples. There were significant differences in the test prevalence of brucellosis by study site, with levels higher in the study area near Lake Mburo National Park than in the study area near Queen Elizabeth National Park. Multivariable regression models identified risk factors associated with increasing test positivity at the individual and farm levels for cattle, goats and humans. Positive associations were seen between increasing seropositivity of brucellosis in goats, cattle and humans. Results of multivariable analyses suggest that improvements in farm biosecurity and hygiene may reduce the risk of brucellosis on the farm and suggest a role for ticks in bovine brucellosis. Although cattle are the focus of brucellosis control in Uganda, the significant associations between seropositivity in humans and seropositivity in goats suggest that brucellosis in goats may be an important contributor to the epidemiology of the disease on the farm. © 2015 Blackwell Verlag GmbH.

Indigenous West Nile virus infections in horses in Albania.

PubMed

Berxholi, K; Ziegler, U; Rexhepi, A; Schmidt, K; Mertens, M; Korro, K; Cuko, A; Angenvoort, J; Groschup, M H

2013-11-01

Serum samples collected from 167 equines of 12 districts in Albania were tested for West Nile virus-specific antibodies by enzyme-linked immunosorbent assay and virus neutralization assay, using WNV lineage 1 and 2. In addition, 95 bird serum samples from Albania and 29 horse samples from Kosovo were tested in ELISA. An overall seroprevalence rate of 22% was found in horses from Albania, whereas no specific antibodies were found in the equine samples from Kosovo and the bird samples. This is the first report indicating WNV infections in animals in Albania, and the first reported seroprevalence study conducted for Kosovo. These results provide evidence for widespread infections of WNV in Albania. © 2013 Blackwell Verlag GmbH.

Evaluation of serum lysyl oxidase as a blood test for colorectal cancer.

PubMed

Ward, S T; Weston, C J; Hepburn, E; Damery, S; Hejmadi, R K; Morton, D G; Middleton, G; Ismail, T; Adams, D H

2014-06-01

Lysyl oxidase (LOX) expression is elevated in colorectal cancer (CRC) tissue and associated with disease progression. A blood test may form a more acceptable diagnostic test for CRC although LOX has not previously been measured in the serum. We therefore sought to determine the clinical usefulness of a serum LOX test for CRC in a symptomatic population. Adult patients referred to a hospital colorectal clinic with bowel symptoms completed a questionnaire and provided a blood sample for serum LOX measurement. Associations between presenting symptoms, serum LOX concentrations and outcomes of investigations were tested by univariate and multivariate analyses to determine if serum LOX was clinically useful in the prediction of CRC. LOX expression in CRC and adjacent colon biopsies was evaluated by ELISA and immunohistochemistry. Thirty-one cases of colorectal cancer and 16 high-risk polyps were identified from a total of 962 participants. There was no association between serum LOX concentration and the presence of CRC, high-risk polyps or cancers at any site. LOX expression was significantly increased in CRC tissue compared to adjacent colon. Despite overexpression of LOX in CRC tissue, elevated serum levels could not be demonstrated. Serum LOX measurement is therefore not a clinically useful test for CRC. Copyright © 2013 Elsevier Ltd. All rights reserved.

The evaluation of a positive direct antiglobulin test (autocontrol) in pretransfusion testing revisited.

PubMed

Judd, W J; Barnes, B A; Steiner, E A; Oberman, H A; Averill, D B; Butch, S H

1986-01-01

Direct antiglobulin tests (DATs) using anti-IgG were performed on 65,049 blood samples from prospective transfusion recipients; 3570 tests (5.49%) were positive. Using criteria published previously (primarily excluding patients not transfused within the preceding 14 days), 778 samples from other than neonatal patients were selected for further evaluation. Eluates that did not react were obtained on 518 (66.6%) of these samples. Warm-reactive autoantibodies were apparent in 192 eluates, while 16 contained drug-related antibodies, anti-A or anti-B from prior transfusion with ABO mismatched blood components, or anti-D passively acquired from immune serum globulin. Fifty-two eluates contained alloantibodies; however, in only six of these cases did the corresponding serum lack unexpected alloantibodies, as determined by routine pretransfusion studies. Three additional weakly reactive clinically significant alloantibodies were detected solely through additional serum tests performed on DAT-positive samples. On the basis of these findings, the DAT had a low predictive value when used to detect the early manifestations of an immune response to recently transfused red cells. Elimination of the autocontrol from routine pretransfusion testing, therefore, carries minimal risk to patients yet will undoubtedly contribute to the containment of health care costs. Moreover, the risk is lower than that associated with the elimination of the antiglobulin crossmatch.

Comparison of different methods for extraction and purification of human Papillomavirus (HPV) DNA from serum samples

NASA Astrophysics Data System (ADS)

Azizah, N.; Hashim, U.; Nadzirah, Sh.; Arshad, M. K. Md; Ruslinda, A. R.; Gopinath, Subash C. B.

2017-03-01

The affectability and unwavering quality of PCR for indicative and research purposes require effective fair systems of extraction and sanitization of nucleic acids. One of the real impediments of PCR-based tests is the hindrance of the enhancement procedure by substances exhibit in clinical examples. This examination considers distinctive techniques for extraction and cleaning of viral DNA from serum tests in view of recuperation productivity as far as yield of DNA and rate recouped immaculateness of removed DNA, and rate of restraint. The best extraction strategies were the phenol/chloroform strategy and the silica gel extraction methodology for serum tests, individually. Considering DNA immaculateness, extraction technique by utilizing the phenol/chloroform strategy delivered the most tasteful results in serum tests contrasted with the silica gel, separately. The nearness of inhibitors was overcome by all DNA extraction strategies in serum tests, as confirm by semiquantitative PCR enhancement.

Long-term in vitro reactivity for HLA antibodies and comparison of detection using serum vs. plasma

PubMed Central

Norris, Philip J.; Lee, Jar-How; Carrick, Danielle M.; Gottschall, Jerome L.; Lebedeva, Mila; de Castro, B.R.; Kleinman, Steven H.; Busch, Michael P.

2010-01-01

BACKGROUND HLA antibodies are a possible cause of transfusion-related acute lung injury (TRALI), and fluorescent bead assays are often used for antibody detection. Serum is the manufacturer’s recommended sample, but plasma may be easier to obtain for studies of HLA antibody prevalence and TRALI case investigations. STUDY DESIGN AND METHODS Specimens were obtained from 44 multiparous females positive for HLA antibodies by lymphocytotoxicity testing at least 13 years prior, and from 1,000 contemporary blood donors. Screening tests were performed using a Luminex-based assay. In addition to comparing results obtained with paired plasma and serum samples, the effects of storage at 4 °C for one week and of multiple freeze-thaw cycles were evaluated. RESULTS Of 42 evaluable subjects with HLA antibodies documented >13 years earlier, only 1 showed loss of detectable antibodies, with 39 (93%) positive in the screening assay for class I and 24 (57%) positive in the screening assay for HLA class II antibodies. In 968 evaluable contemporary donors, 291 screened positive for HLA class I and 206 for HLA class II antibodies using a low assay cut-off. Screening test concordance using paired plasma and serum samples was high, particularly for subjects with higher level antibodies. Refrigeration of samples for one week did not significantly affect assay results, while repeated freeze-thaw cycles caused a decrement in signal level. CONCLUSION Serum and plasma samples gave concordant results in the majority of cases, particularly for specimens with higher-level antibodies. High-level HLA antibodies were present in most individuals for over 13 years. PMID:18980615

Brix refractometry in serum as a measure of failure of passive transfer compared to measured immunoglobulin G and total protein by refractometry in serum from dairy calves.

PubMed

Hernandez, D; Nydam, D V; Godden, S M; Bristol, L S; Kryzer, A; Ranum, J; Schaefer, D

2016-05-01

A series of trials were conducted to evaluate Brix refractometry (Brix %) for the assessment of failure of passive transfer (FPT) in dairy calves compared to: (1) serum IgG (reference standard) when measured by radial immunodiffusion (RID) or a turbidometric immunoassay (TIA), and (2) serum total protein refractometry (STP). For the serum samples tested with TIA, STP, and Brix % (n = 310; Holstein calves), the median concentrations were 21.3 g/L IgG, 58 g/L STP, and 9.2%, respectively. For the serum samples tested with RID, STP and Brix % (n = 112; Jersey calves), the mean concentrations were 38 g/L IgG, 68 g/L STP, and 10.2%, respectively. For samples tested with only Brix % and STP (n = 265; Holstein calves), median STP and Brix % were 50 g/L STP and 8.5%, respectively. Correlations between Brix % and RID, and between Brix % and TIA were equal (r = 0.79, respectively). Brix % and STP were positively correlated (r = 0.99). Brix % estimated serum IgG concentrations determined by TIA and RID (r(2) = 0.63, 0.62, respectively). When FPT was defined as serum IgG < 10 g/L, Brix % ≤ 8.5% showed optimal sensitivity (100%) and specificity (89.2%) to predict FPT. At the same IgG cut-point, an STP value of ≤ 52 g/L showed a similar sensitivity (100%) and specificity (80.4%) to predict FPT. Brix refractometry predicted successful transfer of passive immunity in dairy calves, but further evaluation as a diagnostic tool for the diagnosis of FPT is warranted. Copyright © 2016. Published by Elsevier Ltd.

Screening for chromosomal anomalies in the first trimester: does repeat maternal serum screening improve detection rates?

PubMed

Spencer, Kevin; Cuckle, Howard S

2002-10-01

To assess the within person biological variability of first trimester maternal serum biochemical markers of trisomy 21 across the 10-14 week gestational period. To evaluate whether repeat sampling and testing of free beta-hCG and PAPP-A during this period would result in an improved detection rate. Women presenting at the first trimester OSCAR clinic have blood collected prior to ultrasound dating and nuchal translucency measurement. All samples are analysed for free beta-hCG and PAPP-A before an accurate estimate of gestation is available. In 10% of cases the gestation is prior to the minimum time for NT measurement (11 weeks) and these women are rebooked for a repeat visit to the clinic at the appropriate time. Our fetal database was interrogated to obtain cases in which two maternal blood samples had been collected and analysed in the 10-14 week period. Using data from the marker correlations and statistical modelling, the impact of repeat testing on detection rate for trisomy 21 at a fixed 5% false positive rate, was assessed. 261 pairs of data were available for analysis collected over a 3 year period. The correlation between free beta-hCG in sample 1 and sample 2 was 0.890 and that for PAPP-A was 0.827. The average within person biological variation for free beta-hCG was 21% and 32% for PAPP-A. The increase in detection rate when using both sets of marker data was 3.5% when using serum biochemistry and maternal age, and 1.3% when using nuchal translucency, serum biochemistry and maternal age. Repeat sampling and testing of maternal serum biochemical markers is unlikely to substantially improve first trimester screening performance. Copyright 2002 John Wiley & Sons, Ltd.

Clustering of Toxoplasma gondii Infections Within Families of Congenitally Infected Infants

PubMed Central

Contopoulos-Ioannidis, Despina; Wheeler, Kelsey M.; Ramirez, Raymund; Press, Cindy; Mui, Ernest; Zhou, Ying; Van Tubbergen, Christine; Prasad, Sheela; Maldonado, Yvonne; Withers, Shawn; Boyer, Kenneth M.; Noble, A. Gwendolyn; Rabiah, Peter; Swisher, Charles N.; Heydemann, Peter; Wroblewski, Kristen; Karrison, Theodore; Grigg, Michael E.; Montoya, Jose G.; McLeod, Rima

2015-01-01

Background. Family clusters and epidemics of toxoplasmosis in North, Central, and South America led us to determine whether fathers of congenitally infected infants in the National Collaborative Chicago-Based Congenital Toxoplasmosis Study (NCCCTS) have a high incidence of Toxoplasma gondii infection. Methods. We analyzed serum samples collected from NCCCTS families between 1981 and 2013. Paternal serum samples were tested for T. gondii antibodies with immunoglobulin (Ig) G dye test and IgM enzyme-linked immunosorbent assay. Additional testing of paternal serum samples was performed with differential-agglutination and IgG avidity tests when T. gondii IgG and IgM results were positive and serum samples were collected by the 1-year visit of the congenitally infected child. Prevalence of paternal seropositivity and incidence of recent infection were calculated. We analyzed whether certain demographics, maternal parasite serotype, risk factors, or maternal/infant clinical manifestations were associated with paternal T. gondii infection status. Results. Serologic testing revealed a high prevalence (29 of 81; 36%) of T. gondii infection in fathers, relative to the average seropositivity rate of 9.8% for boys and men aged 12–49 years in the United States between 1994 and 2004 (P < .001). Moreover, there was a higher-than-expected incidence of recent infections among fathers with serum samples collected by the 1-year visit of their child (6 of 45; 13%; P < .001). No demographic patterns or clinical manifestations in mothers or infants were associated with paternal infections, except for sandbox exposure. Conclusions. The high prevalence of chronic and incidence of recent T. gondii infections in fathers of congenitally infected children indicates that T. gondii infections cluster within families in North America. When a recently infected person is identified, family clustering and community risk factors should be investigated for appropriate clinical management. PMID:26405150

Clustering of Toxoplasma gondii Infections Within Families of Congenitally Infected Infants.

PubMed

Contopoulos-Ioannidis, Despina; Wheeler, Kelsey M; Ramirez, Raymund; Press, Cindy; Mui, Ernest; Zhou, Ying; Van Tubbergen, Christine; Prasad, Sheela; Maldonado, Yvonne; Withers, Shawn; Boyer, Kenneth M; Noble, A Gwendolyn; Rabiah, Peter; Swisher, Charles N; Heydemann, Peter; Wroblewski, Kristen; Karrison, Theodore; Grigg, Michael E; Montoya, Jose G; McLeod, Rima

2015-12-15

Family clusters and epidemics of toxoplasmosis in North, Central, and South America led us to determine whether fathers of congenitally infected infants in the National Collaborative Chicago-Based Congenital Toxoplasmosis Study (NCCCTS) have a high incidence of Toxoplasma gondii infection. We analyzed serum samples collected from NCCCTS families between 1981 and 2013. Paternal serum samples were tested for T. gondii antibodies with immunoglobulin (Ig) G dye test and IgM enzyme-linked immunosorbent assay. Additional testing of paternal serum samples was performed with differential-agglutination and IgG avidity tests when T. gondii IgG and IgM results were positive and serum samples were collected by the 1-year visit of the congenitally infected child. Prevalence of paternal seropositivity and incidence of recent infection were calculated. We analyzed whether certain demographics, maternal parasite serotype, risk factors, or maternal/infant clinical manifestations were associated with paternal T. gondii infection status. Serologic testing revealed a high prevalence (29 of 81; 36%) of T. gondii infection in fathers, relative to the average seropositivity rate of 9.8% for boys and men aged 12-49 years in the United States between 1994 and 2004 (P < .001). Moreover, there was a higher-than-expected incidence of recent infections among fathers with serum samples collected by the 1-year visit of their child (6 of 45; 13%; P < .001). No demographic patterns or clinical manifestations in mothers or infants were associated with paternal infections, except for sandbox exposure. The high prevalence of chronic and incidence of recent T. gondii infections in fathers of congenitally infected children indicates that T. gondii infections cluster within families in North America. When a recently infected person is identified, family clustering and community risk factors should be investigated for appropriate clinical management. © The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2010 CFR

2010-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2012 CFR

2012-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2011 CFR

2011-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2013 CFR

2013-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

21 CFR 606.151 - Compatibility testing.

Code of Federal Regulations, 2014 CFR

2014-04-01

... identification. (b) The use of fresh recipient serum or plasma samples less than 3 days old for all...) Procedures to demonstrate incompatibility between the donor's cell type and the recipient's serum or plasma...

Leptospira and Brucella antibodies in collared anteaters (Tamandua tetradactyla) in Brazilian zoos.

PubMed

Sales, Indiara dos Santos; Folly, Márcio Manhães; Garcia, Luize Néli Nunes; Ramos, Tatiane Mendes Varela; da Silva, Mariana Cristina; Pereira, Martha Maria

2012-12-01

The presence of Leptospira spp. and Brucella spp. antibodies was investigated in serum samples from 28 collared anteaters (Tamandua tetradactyla) kept in seven Brazilian zoos. Sera were tested against 19 Leptospira serovars using microscopic agglutination. Samples reacted to the following serovars: two (7.14%) to Patoc, three (10.71%) to Tarrasovi, three (10.71%) to both Patoc and Tarrasovi, two (7.14%) to Wolffi, and one (3.57%) to Australis. Two (7.14%) samples reacted to the buffered Brucella antigen test, but no confirmatory reaction occurred using the 2-mercaptoethanol slow slide agglutination test. No sample was reactive in the agar gel immunodiffusion test for rugose species of Brucella. The presence of anti-leptospira agglutinins in captive T. tetradactyla serum indicates that this species may be susceptible to infection by these bacteria.

Clinical Performance of Aspergillus PCR for Testing Serum and Plasma: a Study by the European Aspergillus PCR Initiative.

PubMed

White, P Lewis; Barnes, Rosemary A; Springer, Jan; Klingspor, Lena; Cuenca-Estrella, Manuel; Morton, C Oliver; Lagrou, Katrien; Bretagne, Stéphane; Melchers, Willem J G; Mengoli, Carlo; Donnelly, J Peter; Heinz, Werner J; Loeffler, Juergen

2015-09-01

Aspergillus PCR testing of serum provides technical simplicity but with potentially reduced sensitivity compared to whole-blood testing. With diseases for which screening to exclude disease represents an optimal strategy, sensitivity is paramount. The associated analytical study confirmed that DNA concentrations were greater in plasma than those in serum. The aim of the current investigation was to confirm analytical findings by comparing the performance of Aspergillus PCR testing of plasma and serum in the clinical setting. Standardized Aspergillus PCR was performed on plasma and serum samples concurrently obtained from hematology patients in a multicenter retrospective anonymous case-control study, with cases diagnosed according to European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) consensus definitions (19 proven/probable cases and 42 controls). Clinical performance and clinical utility (time to positivity) were calculated for both kinds of samples. The sensitivity and specificity for Aspergillus PCR when testing serum were 68.4% and 76.2%, respectively, and for plasma, they were 94.7% and 83.3%, respectively. Eighty-five percent of serum and plasma PCR results were concordant. On average, plasma PCR was positive 16.8 days before diagnosis and was the earliest indicator of infection in 13 cases, combined with other biomarkers in five cases. On average, serum PCR was positive 10.8 days before diagnosis and was the earliest indicator of infection in six cases, combined with other biomarkers in three cases. These results confirm the analytical finding that the sensitivity of Aspergillus PCR using plasma is superior to that using serum. PCR positivity occurs earlier when testing plasma and provides sufficient sensitivity for the screening of invasive aspergillosis while maintaining methodological simplicity. Copyright © 2015 White et al.

Powassan virus infection in snowshoe hares (Lepus americanus).

PubMed

Zarnke, R L; Yuill, T M

1981-04-01

Sera from snowshoe hares (Lepus americanus) trapped near Rochester, Alberta, Canada were tested for Powassan virus antibody by the constant virus/serum dilution neutralization test. Of 1264 serum samples tested, 137 had an antibody titer of at least 1:4 for Powassan virus. Ten hares were inoculated with Powassan virus in the laboratory. Viremia lasted 4-5 days and ceased with the appearance of Powassan antibody in the serum. Neutralizing antibody reached a peak titer of 1:119 on day 15 post-inoculation and was still detectable 13 months post-inoculation.

Prevalence and causes of abnormal PSA recovery.

PubMed

Lautenbach, Noémie; Müntener, Michael; Zanoni, Paolo; Saleh, Lanja; Saba, Karim; Umbehr, Martin; Velagapudi, Srividya; Hof, Danielle; Sulser, Tullio; Wild, Peter J; von Eckardstein, Arnold; Poyet, Cédric

2018-01-26

Prostate-specific antigen (PSA) test is of paramount importance as a diagnostic tool for the detection and monitoring of patients with prostate cancer. In the presence of interfering factors such as heterophilic antibodies or anti-PSA antibodies the PSA test can yield significantly falsified results. The prevalence of these factors is unknown. We determined the recovery of PSA concentrations diluting patient samples with a standard serum of known PSA concentration. Based on the frequency distribution of recoveries in a pre-study on 268 samples, samples with recoveries <80% or >120% were defined as suspect, re-tested and further characterized to identify the cause of interference. A total of 1158 consecutive serum samples were analyzed. Four samples (0.3%) showed reproducibly disturbed recoveries of 10%, 68%, 166% and 4441%. In three samples heterophilic antibodies were identified as the probable cause, in the fourth anti-PSA-autoantibodies. The very low recovery caused by the latter interference was confirmed in serum, as well as heparin- and EDTA plasma of blood samples obtained 6 months later. Analysis by eight different immunoassays showed recoveries ranging between <10% and 80%. In a follow-up study of 212 random plasma samples we found seven samples with autoantibodies against PSA which however did not show any disturbed PSA recovery. About 0.3% of PSA determinations by the electrochemiluminescence assay (ECLIA) of Roche diagnostics are disturbed by heterophilic or anti-PSA autoantibodies. Although they are rare, these interferences can cause relevant misinterpretations of a PSA test result.

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

PubMed

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

2017-11-01

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

[The Application of Gold-immunochromatographic Test Strip Reader in Serum Antibody Detection in Echinococcosis].

PubMed

Xie, Gui-lin; Yin, Jun-xia; Duan, Xin-yu; Feng, Xiao-hui; Wang, Yang; Jiang, Kai; Chen, Xin-mei

2015-06-01

One hundred and fifty-nine serum samples from hydatid disease patients and 80 serum samples from patients with other liver diseases were detected by gold-immunochromatographic assay, and read by naked eyes and the gold-immunochromatographic test strip reader. The sensitivity, specificity, and accuracy of eye-based method was 92.4% (147/159), 85.0% (68/80), and 89.9% (215/239), which was lower than that of the reader detection (95.6%, 93.7%, 95.0%, respectively). While, its false negative rate (7.5%, 12/159) and false positive rate (15.0%, 12/80) was higher than that of the reader detection (4.4% and 6.3%, respectively).

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

PubMed Central

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

2012-01-01

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

Determining skeletal maturation using insulin-like growth factor I (IGF-I) test.

PubMed

Gupta, Shreya; Jain, Sandhya; Gupta, Puneet; Deoskar, Anuradha

2012-11-01

To investigate the validity of Insulin like Growth Factor -1(IGF-1) as a skeletal maturity indicator by comparing serum IGF-1 levels with the stages in cervical vertebral maturation (CVM) and in the middle phalanx of the third finger (MP3). The study population was selected by using simple random sampling technique and consisted of 30 female subjects in the age range of 8-23 years who had blood sample, cephalometric and MP3 radiographs taken on the same day. Serum IGF-I estimation was carried out on the blood samples using chemiluminescence immunoassay (CLIA) method. CVM was evaluated using method by Baccetti et al and MP3 staging was done using Rajagopal & Kansal method. Mean IGF-1 level between the stages was compared by Kruskal-Wallis and Mann Whitney test. Serum IGF-1 levels in females correlate well with skeletal maturity determined by CVM and MP3 stages and increase sharply during early pubertal stages followed by a decrease in late puberty. In addition we hypothesis that serum IGF-1 testing can be undertaken as a preliminary screening test in patients in whom the orthodontist predicts the possibility of using myofunctional appliance but in whom the chronologic age is not suggestive for a growth modification therapy. The finding of the study highlights the fact that the serum IGF-1 estimation can be a valuable tool in assessing skeletal maturation. Copyright © 2012 Società Italiana di Ortodonzia SIDO. Published by Elsevier Srl. All rights reserved.

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

PubMed

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

2015-03-01

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

Prevalence of equine Piroplasmosis and its association with tick infestation in the State of Sao Paulo, Brazil

USDA-ARS?s Scientific Manuscript database

Serum samples were collected from 582 horses from 40 stud farms in the State of São Paulo and tick (Acari: Ixodidae) infestations were evaluated on them. Serum samples were subjected to the complement fixation test (CFT) and a competitive inhibition ELISA (cELISA) for Babesia caballi and Theileria e...

Clinical Validation of a miRNA Blood Test to Identify High-Risk Individuals Eligible for Low-Dose Computed Tomography Screening for Lung Cancer Early Detection

DTIC Science & Technology

2015-10-01

screening. The project requires the use of serum samples obtained from at-risk subjects (heavy smokers with defined characteristics) undergoing...screening. The project requires the use of serum samples obtained from at-risk subjects (heavy smokers with defined characteristics) that were collected

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

PubMed

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

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

Diagnostic reliability of an immunochromatographic test for Chagas disease screening at a primary health care centre in a rural endemic area

PubMed Central

Mendicino, Diego; Stafuza, Mariana; Colussi, Carlina; del Barco, Mónica; Streiger, Mirtha; Moretti, Edgardo

2014-01-01

Many patients with Chagas disease live in remote communities that lack both equipment and trained personnel to perform a diagnosis by conventional serology (CS). Thus, reliable tests suitable for use under difficult conditions are required. In this study, we evaluated the ability of personnel with and without laboratory skills to perform immunochromatographic (IC) tests to detect Chagas disease at a primary health care centre (PHCC). We examined whole blood samples from 241 patients and serum samples from 238 patients. Then, we calculated the percentage of overall agreement (POA) between the two groups of operators for the sensitivity (S), specificity (Sp) and positive (PPV) and negative (NPV) predictive values of IC tests compared to CS tests. We also evaluated the level of agreement between ELISAs and indirect haemagglutination (IHA) tests. The readings of the IC test results showed 100% agreement (POA = 1). The IC test on whole blood showed the following values: S = 87.3%; Sp = 98.8%; PPV = 96.9% and NPV = 95.9%. Additionally, the IC test on serum displayed the following results: S = 95.7%; Sp = 100%; PPV = 100% and NPV = 98.2%. Using whole blood, the agreement with ELISA was 96.3% and the agreement with IHA was 94.1%. Using serum, the agreement with ELISA was 97.8% and the agreement with IHA was 96.6%. The IC test performance with serum samples was excellent and demonstrated its usefulness in a PHCC with minimal equipment. If the IC test S value and NPV with whole blood are improved, then this test could also be used in areas lacking laboratories or specialised personnel. PMID:25466624

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

PubMed

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

2015-01-06

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

Implications of the use of serological and molecular methods to detect infection by Leishmania spp. in urban pet dogs.

PubMed

Paz, Gustavo F; Rugani, Jeronimo M N; Marcelino, Andreza P; Gontijo, Célia M F

2018-06-01

The aim of this study was to evaluate the relationship between naturally occurring Leishmania spp. infections in dogs (Canis familiaris) and the practical implications of the use of serological and molecular methods to confirm diagnoses. The study population consisted of 96 domestic dogs in southeastern Brazil. Serum samples were tested for the presence of anti-Leishmania immunoglobulin G (IgG) antibodies using four commercial canine visceral leishmaniasis kits. Dogs confirmed positive by immunofluorescence antibody test (IFAT) were culled and samples from mesenteric lymph nodes, spleen border, bone marrow and ear skin were taken and submitted to DNA extraction. PCR reactions were performed using primers that amplify a 300-350 bp fragment of the Leishmania ribosomal internal transcribed spacer 1 (ITS1) region. The ITS1 amplified products were analyzed by PCR-RFLP using Hae III restriction endonuclease. To confirm the Leishmania species detected by PCR, each purified sample was sequenced in duplicate. Of the 96 serum samples submitted to serological assays, 8 (8.3%) tested positive for Leishmania by IFAT, 4 (4.1%) by ELISA, 2 (2.1%) by rK39 RDT and 7 (7.3%) by DPP. Four of these infected dogs (50%) were found to be infected only by Leishmania braziliensis or Leishmania amazonensis, and their serum samples tested positive by IFAT and DPP. These findings demonstrate for the first time that cross-reactivity of L. braziliensis and L. amazonensis infection in dogs can be found using the DPP serum test. This is the first record of Leishmania (Leishmania) amazonensis confirmed by a specific molecular marker in dogs (Canis familiaris) from Belo Horizonte, Brazil. Copyright © 2018 Elsevier B.V. All rights reserved.

Identification of patients with nasopharyngeal carcinoma by serum protein profiling using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry.

PubMed

Zhu, Xiao-Dong; Su, Fang; Liang, Zhong-Guo; Li, Ling; Qu, Song; Liang, Xia; Wang, Qi; Liang, Shi-Xiong; Chen, Long

2014-08-01

As diagnosis of nasopharyngeal carcinoma at an early disease stage is important, we attempted to distinguish between patients with nasopharyngeal carcinoma and noncancer controls by using serum protein profiles. Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry and CM10 protein chip were used to detect the serum proteomic patterns of 65 patients with nasopharyngeal carcinoma before radiotherapy and 93 noncancer controls. Proteomic spectra of serum samples from 50 nasopharyngeal carcinoma patients and 60 noncancer controls were used as a training set. The validity of the classification tree was then challenged with a blind test set which included another 15 patients with nasopharyngeal carcinoma and 33 noncancer controls. Biomarker Wizard 3.01 and Biomarker Pattern 5.01 were used in combination to analyze the data and to develop diagnostic models. 21 protein peaks were significantly different between nasopharyngeal carcinoma and controls. 4 mass peaks (M4182, M5343, M5913 and M8702 mass/charge ratio) were chosen automatically to construct a classification tree. The classification tree correctly determined 93.8 % (45/48) of the test samples with 93.3 % (14/15) of the nasopharyngeal carcinoma samples and 93.9 % (31/33) of the noncancer samples. Using a combination of serum protein profiles and Epstein-Barr viral capsid antigen immunoglobulin A antibody tests, the diagnostic sensitivity and specificity were increased to 100 and 97 %, respectively. Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry could correctly distinguish nasopharyngeal carcinoma from noncancer individuals and showed great potential for the development of a screening test for the detection of nasopharyngeal carcinoma.

microRNA-371a-3p as informative biomarker for the follow-up of testicular germ cell cancer patients.

PubMed

van Agthoven, Ton; Eijkenboom, Wil M H; Looijenga, Leendert H J

2017-08-01

α-fetoprotein (AFP) and human chorionic gonadotropin subunit beta (B-HCG) are informative serum biomarkers for the primary diagnosis and follow-up of testicular germ cell cancer (TGCC) patients. About 20% of TGCC patients with a non-seminoma (NS) and about 80% with a seminoma (SE) are, however, negative for these biomarkers. Embryonic stem cell microRNAs (miRs) may serve as promising alternative serum biomarkers. Here we investigated a retrospective series of serum samples from selected TGCC patients who developed a relapse in time to test the possible additional value of the serum-based ampTSmiR test compared to the conventional serum-based protein biomarkers for follow-up. We investigated 261 retrospective serum samples of six selected fully evaluated TGCC patients with a proven relapse using the ampTSmiR test for miR-371a-3p, miR-373-3p, and miR-367-3p and compared the results to those of the conventional protein biomarkers. At primary diagnosis, elevated serum B-HCG, AFP and LDH levels were found to be informative in 4/6, 3/6 and 3/6 patients, respectively. At primary diagnosis the levels of miR-371a-3p and miR-373-3p were elevated in 4/4, and miR-367-3p in 3/4 patients. For two cases no starting serum sample was available for retrospective miR analysis. Residual disease (overlooked by histopathological examination) was detected in one case by miR-371a-3p only. The miR-371a-3p level was increased in one patient two months before detection of an intracranial metastasis. B-HCG was informative in 3/4 and the ampTSmiR test in 4/4 patients with a relapse or residual disease. None of the biomarkers were informative for the detection of residual mature teratoma. The ampTSmiR test is more sensitive than the conventional TGCC protein biomarkers for the detection of residual disease and relapse, excluding mature teratoma.

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

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

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

2012-01-01

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

Cross-sectional study of serum antibodies against Sarcocystis neurona in cats tested for antibodies against Toxoplasma gondii.

PubMed

Rossano, Mary G; Murphy, Alice J; Vrable, Ruth A; Vanzo, Nicole E; Lewis, Stacy K; Sheline, Katherine D; Kaneene, John B; Mansfield, Linda S

2002-08-15

To determine apparent seroprevalence of antibodies against Sarcocystis neurona in a population of domestic cats previously tested for antibodies against Toxoplasma gondii. Cross-sectional study. Serum from 196 domestic cats. Banked serum samples submitted to the Michigan State University Animal Health Diagnostic Laboratory for T. gondii diagnostic testing were tested for antibodies against S. neurona by use of an indirect fluorescent antibody (IFA) test and a western blot test. Submission records were analyzed to determine descriptive statistics and test for associations between positive results of a test for S. neurona and other variables in the data set. 10 of 196 (5%) samples yielded positive results for antibodies against S. neurona by use of western blot analysis, whereas 27 samples yielded positive results by use of the IFA. No association was found between S. neurona western blot test results and T. gondii test results, age, sex, or the reason for T. gondii testing. The S. neurona IFA titer was positively and significantly associated with positive results of western blot analysis. Domestic cats are not likely to play a substantial role as intermediate hosts in the natural life cycle of S. neurona. Results indicate that natural infection of domestic cats may occur, and small animal practitioners should be aware of this fact when evaluating cats with neurologic disease. The S. neurona IFA test had lower specificity than western blot analysis.

Detection of African Swine Fever Virus Antibodies in Serum and Oral Fluid Specimens Using a Recombinant Protein 30 (p30) Dual Matrix Indirect ELISA.

PubMed

Giménez-Lirola, Luis G; Mur, Lina; Rivera, Belen; Mogler, Mark; Sun, Yaxuan; Lizano, Sergio; Goodell, Christa; Harris, D L Hank; Rowland, Raymond R R; Gallardo, Carmina; Sánchez-Vizcaíno, José Manuel; Zimmerman, Jeff

2016-01-01

In the absence of effective vaccine(s), control of African swine fever caused by African swine fever virus (ASFV) must be based on early, efficient, cost-effective detection and strict control and elimination strategies. For this purpose, we developed an indirect ELISA capable of detecting ASFV antibodies in either serum or oral fluid specimens. The recombinant protein used in the ELISA was selected by comparing the early serum antibody response of ASFV-infected pigs (NHV-p68 isolate) to three major recombinant polypeptides (p30, p54, p72) using a multiplex fluorescent microbead-based immunoassay (FMIA). Non-hazardous (non-infectious) antibody-positive serum for use as plate positive controls and for the calculation of sample-to-positive (S:P) ratios was produced by inoculating pigs with a replicon particle (RP) vaccine expressing the ASFV p30 gene. The optimized ELISA detected anti-p30 antibodies in serum and/or oral fluid samples from pigs inoculated with ASFV under experimental conditions beginning 8 to 12 days post inoculation. Tests on serum (n = 200) and oral fluid (n = 200) field samples from an ASFV-free population demonstrated that the assay was highly diagnostically specific. The convenience and diagnostic utility of oral fluid sampling combined with the flexibility to test either serum or oral fluid on the same platform suggests that this assay will be highly useful under the conditions for which OIE recommends ASFV antibody surveillance, i.e., in ASFV-endemic areas and for the detection of infections with ASFV isolates of low virulence.

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

PubMed

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

1993-01-01

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

The effect of sample storage on the performance and reproducibility of the galactomannan EIA test.

PubMed

Kimpton, George; White, P Lewis; Barnes, Rosemary A

2014-08-01

Galactomannan enzyme immune assay (GM EIA) is a nonculture test for detecting invasive aspergillosis (IA) forming a key part of diagnosis and management. Recent reports have questioned the reproducibility of indices after sample storage. To investigate this, 198 serum samples (72 from cases and 126 from controls) and 61 plasma samples (24 from cases and 37 from controls), initially tested between 2010 and 2013, were retested to determine any change in index. Data were also collected on circulatory protein levels for false-positive serum samples. Serum indices significantly declined on retesting (median: initial, 0.50, retest, 0.23; P < 0.0001). This was shown to be diagnosis dependent as the decline was apparent on retesting of control samples (median: initial 0.50, retest 0.12; P < 0.0001), but was not evident with case samples (median: initial, 0.80, retest, 0.80; P = 0.724). Plasma samples showed little change on reanalysis after long-term storage at 4°C. Retesting after freezing showed a decrease in index values for controls (median: initial 0.40, retest 0.26; P = 0.0505), but no significant change in cases. Circulatory proteins showed a correlation between serum albumin concentration and difference in index value on retesting. Overall, this study suggests that a lack of reproducibility in GM EIA positivity is only significant when disease is absent. Retesting after freezing helps to differentiate false-positive GM EIA results and, with consecutive positivity, could help to improve accuracy in predicting disease status. The freezing of samples prior to testing could potentially reduce false-positivity rates and the need to retest. © The Author 2014. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Bluetongue virus surveillance in the Islamic Republic of Mauritania: Is serotype 26 circulating among cattle and dromedaries?

PubMed

Lorusso, Alessio; Baba, Doumbia; Spedicato, Massimo; Teodori, Liana; Bonfini, Barbara; Marcacci, Maurilia; Di Provvido, Andrea; Isselmou, Katia; Marini, Valeria; Carmine, Irene; Scacchia, Massimo; Di Sabatino, Daria; Petrini, Antonio; Bezeid, Beyatt Ahmed; Savini, Giovanni

2016-06-01

In March 2013, EDTA-blood and serum samples were collected from 119 cattle and 159 dromedaries at the slaughterhouse of Nouakchott, the capital city of the Islamic Republic of Mauritania. Serum samples were screened for the presence of Bluetongue (BT) antibodies by competitive ELISA (cELISA). Positive samples were then tested by serum-neutralization (SN) to determine BTV serotype. RNA from blood samples was first tested by a genus-specific quantitative RT-PCR assay which is able to detect all 27 existing BTV serotypes (RT-qPCR1-27). Positive samples were further screened by a RT-qPCR assay which, instead, is able to detect the classical 24 BTV serotypes only (RT-qPCR1-24). Of the 278 serum samples tested, 177 (mean=63.7%; 95% CI: 57.9%-69.1%) resulted positive by cELISA. Of these, 69 were from cattle (mean=58.0%; 95% CI: 49.0%-66.5%) and 108 from dromedaries (mean=67.9%; 95% CI: 60.3%-74.7%). BTV-26 neutralizing antibodies were by far the most frequently found as they were detected in 146 animals with titres ranging from 1:10 to 1:80. Out of 278 blood samples, 25 (mean=9.0%; 95% CI: 6.2%-12.9%) were found positive for BTV by RT-qPCR1-27, 20 (mean=16.8%; 95% CI: 11.2%-24.6%) were from cattle and 5 (mean=3.1%; 95% CI: 1.4%-7.1%) from dromedaries. When tested by RT-qPCR1-24 the 25 BTV positive samples were negative. Unfortunately, no genetic information by molecular typing or by next generation sequencing has been obtained as for the very low levels of RNA in the blood samples. Copyright © 2016 Elsevier B.V. All rights reserved.

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

PubMed

Nyberg, G; Mårtensson, E

1986-01-01

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

Bayesian estimation of the sensitivity and specificity of individual fecal culture and Paralisa to detect Mycobacterium avium subspecies paratuberculosis infection in young farmed deer.

PubMed

Stringer, Lesley A; Jones, Geoff; Jewell, Chris P; Noble, Alasdair D; Heuer, Cord; Wilson, Peter R; Johnson, Wesley O

2013-11-01

A Bayesian latent class model was used to estimate the sensitivity and specificity of an immunoglobulin G1 serum enzyme-linked immunosorbent assay (Paralisa) and individual fecal culture to detect young deer infected with Mycobacterium avium subsp. paratuberculosis. Paired fecal and serum samples were collected, between July 2009 and April 2010, from 20 individual yearling (12-24-month-old) deer in each of 20 South Island and 18 North Island herds in New Zealand and subjected to culture and Paralisa, respectively. Two fecal samples and 16 serum samples from 356 North Island deer, and 55 fecal and 37 serum samples from 401 South Island deer, were positive. The estimate of individual fecal culture sensitivity was 77% (95% credible interval [CI] = 61-92%) with specificity of 99% (95% CI = 98-99.7%). The Paralisa sensitivity estimate was 19% (95% CI = 10-30%), with specificity of 94% (95% CI = 93-96%). All estimates were robust to variation of priors and assumptions tested in a sensitivity analysis. The data informs the use of the tests in determining infection status at the individual and herd level.

Evaluation of reactive oxygen metabolites in frozen serum samples. Effect of storage and repeated thawing.

PubMed

Cavalleri, A; Colombo, C; Venturelli, E; Miceli, R; Mariani, L; Cornelli, U; Pala, V; Berrino, F; Secreto, G

2004-01-01

Measuring the free radical activity in serum samples from prospective studies is the best way to investigate the association between oxidative stress and human diseases. Prospective studies require the analysis of serum samples that have often been stored for a long time. Our study was designed to determine the effect of storage at -30 degrees C and -80 degrees C for two years on free radical activity. We analyzed the free radical activity by measuring circulating hydroperoxides in a pool of sera at baseline and after one day, one week, one month and 25 months of storage, using a photometric method (d-ROMs test). Measurements were performed in aliquots thawed only once at each time point and in aliquots frozen and thawed repeatedly over the study period. After two years we observed a small but statistically significant 4% decrease in the hydroperoxide concentration, which was substantially unaffected by storage temperatures and repeated freeze-thaw cycles. We also carried out the d-ROMs test in sera from ten apparently healthy volunteers at 2, 8, 24, and 48 hours after collection and storage at 4 degrees C and did not observe any significant variation. In conclusion, the d-ROMs test is a simple method suitable to evaluate the free radical activity in frozen serum samples after long-term storage.

Anti-signal recognition particle autoantibody ELISA validation and clinical associations.

PubMed

Aggarwal, Rohit; Oddis, Chester V; Goudeau, Danielle; Fertig, Noreen; Metes, Ilinca; Stephens, Chad; Qi, Zengbiao; Koontz, Diane; Levesque, Marc C

2015-07-01

The aim of this study was to develop and validate a quantitative anti-signal recognition particle (SRP) autoantibody serum ELISA in patients with myositis and longitudinal association with myositis disease activity. We developed a serum ELISA using recombinant purified full-length human SRP coated on ELISA plates and a secondary antibody that bound human IgG to detect anti-SRP binding. Protein immunoprecipitation was used as the gold standard for the presence of anti-SRP. Serum samples from three groups were analysed: SRP(+) myositis subjects by immunoprecipitation, SRP(-) myositis subjects by immunoprecipitation and non-myositis controls. The ELISA's sensitivity, specificity, positive predictive value and negative predictive value were evaluated. Percentage agreement and test-retest reliability were assessed. Serial samples from seven SRP immunoprecipitation-positive subjects were also tested, along with serum muscle enzymes and manual muscle testing. Using immunoprecipitation, we identified 26 SRP(+) myositis patients and 77 SRP(-) controls (including 38 patients with necrotizing myopathy). Non-myositis control patients included SLE (n = 4) and SSc (n = 7) patients. Anti-SRP positivity by ELISA showed strong agreement (97.1%) with immunoprecipitation (κ = 0.94). The sensitivity, specificity, positive predictive value, and negative predictive value of the anti-SRP ELISA were 88, 100, 100 and 96, respectively. The area under the curve was 0.94, and test-retest reliability was strong (r = 0.91, P < 0.001). Serial samples showed that anti-SRP levels paralleled changes in muscle enzymes and manual muscle testing. We developed a quantitative ELISA for detecting serum anti-SRP autoantibodies and validated the assay in myositis. Longitudinal assessment of SRP levels by ELISA may be a useful biomarker for disease activity. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

VERY LOW INFLUENZA A VIRUS PREVALENCE IN CERVIDS IN GERMAN NATIONAL PARKS.

PubMed

Soilemetzidou, Sanatana-Eirini; Greenwood, Alex D; Czirják, Gábor Á

2018-03-01

Influenza A viruses are one of the most important and most studied pathogens in humans and domestic animals but little is known about viral prevalence in non-avian wildlife. Serum samples from three free-ranging cervid species (red [ Cervus elaphus], fallow [ Dama dama] , and roe deer [ Capreolus capreolus]) were collected from six German national parks between 2000 and 2002. The serum was tested for the presence of influenza A antibodies using a commercial competitive enzyme-linked immunosorbent assay. Only one of 137 samples tested positive.

Plasma endotoxin core antibody concentration and linear growth are unrelated in rural Malawian children aged 2-5 years.

PubMed

Benzoni, Nicole; Korpe, Poonum; Thakwalakwa, Chrissie; Maleta, Ken; Stephenson, Kevin; Manary, Micah; Manary, Mark

2015-06-24

Environmental enteropathy is subclinical inflammation of the upper gastrointestinal tract associated with reduced linear growth in developing countries. Usually investigators have used biopsy or a dual sugar absorption test to assess environmental enteropathy. Such tests are time and resource intensive, restricting their utility as screening methods. Serum endotoxin core antibody (EndoCab) concentration is a potential indicator of intestinal inflammation and integrity, and thus may be useful to predict environmental enteropathy. We analyzed the association of serum EndoCab levels versus linear growth and lactulose-mannitol assay results in 2-5 year old rural Malawian children. This was an observational study of 388 rural, asymptomatic Malawian children who had anthropometric measurements taken at least every 3 months since birth. In June and July 2011, dual sugar permeability tests were performed and serum samples were drawn for EndoCab assays. Pearson correlation, Student's t test and multivariable linear regression were used to compare ln EndoCab concentrations with height-for-age z scores (HAZ) at time of sampling and 3 months later. Identical analysis was also performed for ln EndoCab versus measurements from dual sugar permeability testing performed in conjunction with serum sampling. In a subgroup of children with anthropometric data in the months prior to serum sampling, Pearson correlation was used to estimate the relationship between ln EndoCab and recent linear growth. Ln EndoCab concentrations were not correlated with HAZ at time of measurement (B = -0.078, P = 0.14) nor change in HAZ over the subsequent 3 months HAZ (B = -0.018, P = 0.27). EndoCab concentration was not associated with %lactulose excretion (B < 0.001, P = 0.98) nor the lactulose:mannitol ratio (B = 0.021, P = 0.62). Subgroup analysis also did not reveal any significant association between EndoCab and recent growth. EndoCab titers were not correlated with measurements of growth or intestinal permeability in rural pre-school aged Malawian children.

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

Serum and Plasma Metabolomic Biomarkers for Lung Cancer.

PubMed

Kumar, Nishith; Shahjaman, Md; Mollah, Md Nurul Haque; Islam, S M Shahinul; Hoque, Md Aminul

2017-01-01

In drug invention and early disease prediction of lung cancer, metabolomic biomarker detection is very important. Mortality rate can be decreased, if cancer is predicted at the earlier stage. Recent diagnostic techniques for lung cancer are not prognosis diagnostic techniques. However, if we know the name of the metabolites, whose intensity levels are considerably changing between cancer subject and control subject, then it will be easy to early diagnosis the disease as well as to discover the drug. Therefore, in this paper we have identified the influential plasma and serum blood sample metabolites for lung cancer and also identified the biomarkers that will be helpful for early disease prediction as well as for drug invention. To identify the influential metabolites, we considered a parametric and a nonparametric test namely student׳s t-test as parametric and Kruskal-Wallis test as non-parametric test. We also categorized the up-regulated and down-regulated metabolites by the heatmap plot and identified the biomarkers by support vector machine (SVM) classifier and pathway analysis. From our analysis, we got 27 influential (p-value<0.05) metabolites from plasma sample and 13 influential (p-value<0.05) metabolites from serum sample. According to the importance plot through SVM classifier, pathway analysis and correlation network analysis, we declared 4 metabolites (taurine, aspertic acid, glutamine and pyruvic acid) as plasma biomarker and 3 metabolites (aspartic acid, taurine and inosine) as serum biomarker.

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

PubMed

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

2014-01-01

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

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

PubMed

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

2018-04-06

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

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

PubMed

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

2007-11-01

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

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

PubMed

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

2018-07-01

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

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

PubMed

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

2014-01-07

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

Serum and urine metabolomic fingerprinting in diagnostics of inflammatory bowel diseases

PubMed Central

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

2014-01-01

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

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

PubMed

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

2014-04-01

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

Evaluation of Aspergillus PCR protocols for testing serum specimens.

PubMed

White, P Lewis; Mengoli, Carlo; Bretagne, Stéphane; Cuenca-Estrella, Manuel; Finnstrom, Niklas; Klingspor, Lena; Melchers, Willem J G; McCulloch, Elaine; Barnes, Rosemary A; Donnelly, J Peter; Loeffler, Juergen

2011-11-01

A panel of human serum samples spiked with various amounts of Aspergillus fumigatus genomic DNA was distributed to 23 centers within the European Aspergillus PCR Initiative to determine analytical performance of PCR. Information regarding specific methodological components and PCR performance was requested. The information provided was made anonymous, and meta-regression analysis was performed to determine any procedural factors that significantly altered PCR performance. Ninety-seven percent of protocols were able to detect a threshold of 10 genomes/ml on at least one occasion, with 83% of protocols reproducibly detecting this concentration. Sensitivity and specificity were 86.1% and 93.6%, respectively. Positive associations between sensitivity and the use of larger sample volumes, an internal control PCR, and PCR targeting the internal transcribed spacer (ITS) region were shown. Negative associations between sensitivity and the use of larger elution volumes (≥100 μl) and PCR targeting the mitochondrial genes were demonstrated. Most Aspergillus PCR protocols used to test serum generate satisfactory analytical performance. Testing serum requires less standardization, and the specific recommendations shown in this article will only improve performance.

Evaluation of Aspergillus PCR Protocols for Testing Serum Specimens▿†

PubMed Central

White, P. Lewis; Mengoli, Carlo; Bretagne, Stéphane; Cuenca-Estrella, Manuel; Finnstrom, Niklas; Klingspor, Lena; Melchers, Willem J. G.; McCulloch, Elaine; Barnes, Rosemary A.; Donnelly, J. Peter; Loeffler, Juergen

2011-01-01

A panel of human serum samples spiked with various amounts of Aspergillus fumigatus genomic DNA was distributed to 23 centers within the European Aspergillus PCR Initiative to determine analytical performance of PCR. Information regarding specific methodological components and PCR performance was requested. The information provided was made anonymous, and meta-regression analysis was performed to determine any procedural factors that significantly altered PCR performance. Ninety-seven percent of protocols were able to detect a threshold of 10 genomes/ml on at least one occasion, with 83% of protocols reproducibly detecting this concentration. Sensitivity and specificity were 86.1% and 93.6%, respectively. Positive associations between sensitivity and the use of larger sample volumes, an internal control PCR, and PCR targeting the internal transcribed spacer (ITS) region were shown. Negative associations between sensitivity and the use of larger elution volumes (≥100 μl) and PCR targeting the mitochondrial genes were demonstrated. Most Aspergillus PCR protocols used to test serum generate satisfactory analytical performance. Testing serum requires less standardization, and the specific recommendations shown in this article will only improve performance. PMID:21940479

Persistence of Immunity Acquired after a Single Dose of Rubella Vaccine in Japan.

PubMed

Okafuji, Takao; Okafuji, Teruo; Nakayama, Tetsuo

2016-05-20

To date, Takahashi, Matsuura, and TO-336 strains of live-attenuated rubella vaccine have been used in Japan. Japan implemented a single-dose rubella vaccination program until 2006. However, few reports are available on the persistence of immunity after this vaccination program. We collected 276 serum samples from January 2009 to December 2011 at Okafuji Pediatric Clinic and assessed the immune status of these samples against rubella virus during 1-10 years after vaccination with a single dose of Takahashi rubella vaccine. Regional outbreak of rubella did not occur during 1999-2011. The collected serum samples were tested for antibodies against the rubella virus by performing a standard hemagglutination inhibition (HAI) test. Our results showed that all the tested serum samples contained antibodies against the rubella virus 10 years after the vaccination. Geometric mean titer of HAI antibodies was 1:180 and decreased to 1:68 at 10 years after the vaccination. The levels of HAI antibodies decreased logarithmically with time after the vaccination. In conclusion, vaccine-acquired immunity after vaccination with a single dose of live-attenuated Takahashi rubella vaccine was retained for at least 10 years when rubella was under regional control.

Prevalence of antileptospiral serum antibodies in dogs in Ireland

USDA-ARS?s Scientific Manuscript database

A total of 474 serum samples from client owned Irish dogs were tested for the presence of antibodies against serovars Canicola, Icterohaemorrhagiae, Bratislava, Autumnalis, Pomona, Altodouro, Grippotyphosa, Mozdok, Hardjobovis and Ballum. Six percent of dogs presented to veterinary practitioners for...

Erysipelothrix rhusiopathiae and Mycoplasma hyopneumoniae: the sensitivities of enzyme-linked immunosorbent assays for detecting vaccinated sows of unknown disease status using serum and colostrum, and the correlation of the results for sow serum, colostrum, and piglet serum.

PubMed

Jenvey, Caitlin J; Reichel, Michael P; Cockcroft, Peter D

2015-03-01

Due to relatively high concentrations of immunoglobulins, colostrum has the potential to improve the sensitivity of diagnostic tests for diseases in pigs when compared with serum. It is possible that colostrum could improve the sensitivity of the antibody enzyme-linked immunosorbent assay (ELISA) compared with serum. Colostrum is also essential for piglets, providing protection against infections in the first few weeks and months of life. The sensitivity of 2 commercially available ELISAs, one for the detection of Erysipelothrix rhusiopathiae and the second for Mycoplasma hyopneumoniae antibodies, when used with sow colostrum in comparison with serum was investigated. The correlation of maternal E. rhusiopathiae- and M. hyopneumoniae-specific antibody levels with specific-antibody serum levels in the piglet was also determined. The sensitivity was defined as the proportion of vaccinated sows that were correctly identified as vaccinated at a given cutoff point. The true disease status of the sows with regard to the 2 infections was unknown. Blood and colostrum samples were collected from 20 sows, 10 primiparous and 10 multiparous, and blood samples were also collected from the piglets of each sow, 48-72 hr post-farrowing. The sensitivities of both ELISAs were significantly improved when using colostrum compared with serum. Sow serum and colostrum optical density (OD) values were significantly correlated. The mean sow OD values for serum for E. rhusiopathiae and M. hyopneumoniae and colostrum for E. rhusiopathiae were significantly correlated with piglet serum OD levels. If the improved sensitivity of colostrum can be demonstrated in infected animals, this will increase the ability of the test to identify infected animals using both individual and pooled colostrum. Testing serum and/or colostrum using ELISA can be useful predictors of piglet disease-specific OD values. © 2015 The Author(s).

Smart management of sample dilution using an artificial neural network to achieve streamlined processes and saving resources: the automated nephelometric testing of serum free light chain as case study.

PubMed

Ialongo, Cristiano; Pieri, Massimo; Bernardini, Sergio

2017-02-01

Saving resources is a paramount issue for the modern laboratory, and new trainable as well as smart technologies can be used to allow the automated instrumentation to manage samples more efficiently in order to achieve streamlined processes. In this regard the serum free light chain (sFLC) testing represents an interesting challenge, as it usually causes using a number of assays before achieving an acceptable result within the analytical range. An artificial neural network based on the multi-layer perceptron (MLP-ANN) was used to infer the starting dilution status of sFLC samples based on the information available through the laboratory information system (LIS). After the learning phase, the MLP-ANN simulation was applied to the nephelometric testing routinely performed in our laboratory on a BN ProSpec® System analyzer (Siemens Helathcare) using the N Latex FLC kit. The MLP-ANN reduced the serum kappa free light chain (κ-FLC) and serum lambda free light chain (λ-FLC) wasted tests by 69.4% and 70.8% with respect to the naïve stepwise dilution scheme used by the automated analyzer, and by 64.9% and 66.9% compared to a "rational" dilution scheme based on a 4-step dilution. Although it was restricted to follow-up samples, the MLP-ANN showed good predictive performance, which alongside the possibility to implement it in any automated system, made it a suitable solution for achieving streamlined laboratory processes and saving resources.

Detection of cerebrospinal fluid leakage by specific measurement of transferrin glycoforms.

PubMed

Kwon, Seok-Joon; Zhang, Fuming; Dordick, Jonathan S; Sonstein, William J; Linhardt, Robert J

2015-10-01

A simple and rapid detection of cerebrospinal fluid (CSF) leakage would benefit spine surgeons making critical postoperative decisions on patient care. We have assessed novel approaches to selectively determine CSF β2-transferrin (β2TF), an asialo-transferrin (aTF) biomarker, without interference from serum sialo-transferrin (sTF) in test samples. First, we performed mild periodate oxidation to selectively generate aldehyde groups in sTF for capture with magnetic hydrazide microparticles, and selective removal with a magnetic separator. Using this protocol sTF was selectively removed from mixtures of CSF and serum containing CSF aTF (β2TF) and serum sTF, respectively. Second, a two-step enzymatic method was developed with neuraminidase and galactose oxidase for generating aldehyde groups in sTF present in CSF and serum mixtures for magnetic hydrazide microparticle capture. After selectively removing sTF from mixtures of CSF and serum, ELISA could detect significant TF signal only in CSF, while the TF signal in serum was negligible. The new approach for selective removal of only sTF in test samples will be promising for the required intervention by a spine surgeon. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

PubMed

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

2018-03-01

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

Presence of antigen and antibodies in serum and genital discharges of cows from dairy herds naturally infected with Leptospira interrogans serovar hardjo.

PubMed

Dhaliwal, G S; Murray, R D; Dobson, H; Montgomery, J; Ellis, W A

1996-03-01

Samples of cervico-vaginal mucus from 163 bulling cows (group 1) and post calving discharges from 59 newly calved cows (group 2) in five dairy herds naturally infected with Leptospira interrogans serovar hardjo were examined for the presence of antigen and IgG and IgA antibodies by using two ELISA systems which were protein or carbohydrate based. Corresponding serum samples were examined for systemic immune responses by using a microscopic agglutination test (MAT) and IgG-ELISA tests. Antigen was detected by direct immunofluorescence in six of the 163 samples of cervico-vaginal mucus. Both IgG and IgA antibodies were detected by ELISA in the genital discharges with a prevalence much higher than that obtained by the MAT but lower than that observed with the serum IgG-ELISA. Combining both groups, none of the MAT-positive cattle was negative by serum-ELISA. By using the protein or carbohydrate fraction serum IgG-ELISA assays, respectively, 29 or 41 per cent of the MAT-negative cows were positive at a titre of at least 1:40. Similarly, eight or 23 samples (10 or 27 per cent) had titres of at least 1:20 in the genital discharge ELISA for IgG and IgA antibodies, respectively. The serum IgG-ELISA was the most efficient in detecting hardjo antibodies, but in group 2 the IgG- and IgA-ELISA of the post calving discharge proved to be equally effective.

METHOD FOR MICRORNA ISOLATION FROM CLINICAL SERUM SAMPLES

PubMed Central

Li, Yu; Kowdley, Kris V.

2012-01-01

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

Evaluation of the use of nonesterified fatty acids and β-hydroxybutyrate concentrations in pooled serum samples for herd-based detection of subclinical ketosis in dairy cows during the first week after parturition.

PubMed

Borchardt, Stefan; Staufenbiel, Rudolf

2012-04-15

To evaluate the use of nonesterified fatty acids (NEFA) and β-hydroxybutyrate (BHBA) concentrations in pooled serum samples for herd-based detection of subclinical ketosis (SCK) in dairy cows after calving. Cross-sectional study. 1,100 dairy cows from 110 herds. Blood samples were collected from 10 healthy cows/herd in the first week after parturition. Aliquots of serum were mixed to create a pooled sample. Concentrations of NEFA and BHBA were measured to estimate prevalence of SCK. Pooled sample test results were compared with those obtained for individual samples. Linear regression and receiver-operating characteristic curve analysis were performed; Bland-Altman plots were used to evaluate agreement between methods. Overall prevalence of SCK was 30.7%, 19.3%, and 13.6%, as determined by use of BHBA threshold concentrations of 1,000, 1,200, and 1,400 μmol/L, respectively. Pooled sample concentrations of NEFA and BHBA were significantly correlated (r = 0.98 and 0.97, respectively) with individual sample means and with the number of cows that had NEFA (R(2) range, 0.81 to 0.84) or BHBA (R(2) range, 0.65 to 0.76) concentrations above predefined thresholds. Pooled sample concentrations of NEFA and BHBA were very accurate to highly accurate for herd-based detection of SCK. Analysis of NEFA and BHBA concentrations in pooled serum samples was useful for herd-based detection of SCK. A sample size of 10 cows/herd was deemed adequate for monitoring dairy herds for SCK. Reference criteria specific to pooled samples should be used for this type of herd-based testing.

Comparison of RFFIT tests with different standard sera and testing procedures.

PubMed

Yu, Peng-cheng; Noguchi, Akira; Inoue, Satoshi; Tang, Qing; Rayner, Simon; Liang, Guo-dong

2012-06-01

The World Health Organization (WHO) standard assay for determining antibody level is the rapid fluorescent focus inhibition test (RFFIT) and is used to determine the degree of immunity after vaccination against rabies. To compare the difference in RFFIT results between the laboratories of The National Institute of Infectious Disease in Japan (NIID) and the Chinese Centre for Disease Control (CCDC) as well the influence of the choice of standard serum (STD) for the detection, the two laboratories detection methods were simultaneously manipulated by RFFIT. The reference serums used in NIID and the WHO standard serum used in CCDC were compared in the same RFFIT detection to determine the titer of four sera samples C1, S1, S2 and S4 in parallel, and the titers of the detected sera samples were calculated using the standard formula for neutralizing antibody titer. No significant difference was found in RFFIT methods from the two laboratories and the RFFIT testing procedures of the two laboratories have good consistency. However, different titers were obtained with the tentative internal standard serum (TI-STD) produced by adjusting to 2.0 IU of WHO standard serum in NIID and the WHO STD. The titer determined with the TI-STD was higher than that determined with WHO STD, This difference appears to be significant and requires further investigation.

Easy fix for clinical laboratories for the false-positive defect with the Abbott AxSym total beta-hCG test.

PubMed

Cole, Laurence A; Khanlian, Sarah A

2004-05-01

False-positive hCG results can lead to erroneous diagnoses and needless chemotherapy and surgery. In the last 2 years, eight publications described cases involving false-positive hCG tests; all eight involved the AxSym test. We investigated the source of this abundance of cases and a simple fix that may be used by clinical laboratories. False-positive hCG was primarily identified by absence of hCG in urine and varying or negative hCG results in alternative tests. Seventeen false-positive serum samples in the AxSym test were evaluated undiluted and at twofold dilution with diluent containing excess goat serum or immunoglobulin. We identified 58 patients with false-positive hCG, 47 of 58 due to the Abbott AxSym total hCGbeta test (81%). Sixteen of 17 of these "false-positive" results (mean 100 mIU/ml) became undetectable when tested again after twofold dilution. A simple twofold dilution with this diluent containing excess goat serum or immunoglobulin completely protected 16 of 17 samples from patients having false-positive results. It is recommended that laboratories using this test use twofold dilution as a minimum to prevent false-positive results.

Development of an indirect enzyme-linked immunosorbent assay test for detecting antibodies to chicken astrovirus in chicken sera.

PubMed

Skibinska, A; Lee, A; Wylie, M; Smyth, V J; Welsh, M D; Todd, D

2015-01-01

The development of an indirect enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of Group B chicken astrovirus (CAstV) infections is described. The test was based on the use of an affinity-purified capsid antigen, specific to CAstV isolate 11672, produced as a glutathione-S-transferase N-terminal fusion protein by a recombinant baculovirus. Strongly positive ELISA signals were elicited against experimentally produced antisera raised to CAstVs from Group B (subgroups i and ii) but were negative for antisera raised to a Group A CAstV. Using a panel of 240 selected serum samples, 99% agreement was observed when the results obtained by ELISA were compared to those from an indirect immunofluorescence test for CAstV 11672. The ELISA test was applied to 68 serum sets comprising 1864 samples, which were obtained from parent and grandparent flocks originating mainly in the UK. Of the 52 sets containing ELISA-positive samples, 24 sets had >75% samples positive and nine sets had <25% samples positive and were regarded as having high and low seropositivities, respectively. Of the 1864 serum samples tested 1090 (58.5%) were ELISA positive and of these, 234 sera (21.5%) produced strongly positive signals, whereas moderately positive and weakly positive signals were produced by 562 (51.5%) and 294 (27%) sera. When used for flock screening purposes, this ELISA test can be used to (i) investigate the occurrence of first-time CAstV infections of parent flocks during lay and the possible adverse effects caused by vertically transmitted CAstV infections on broiler hatchability and performance and (ii) diagnose Group B CAstV infections within specific pathogen free flocks.

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

PubMed Central

2004-01-01

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

Ascorbic acid interference in the measurement of serum biochemical parameters: in vivo and in vitro studies.

PubMed

Martinello, Flávia; da Silva, Edson Luiz

2006-04-01

To investigate the negative interference of ascorbic acid in serum biochemical tests in relation to the dose of vitamin C intake and to the time of blood collection. Healthy volunteers (n = 18) consumed daily doses of vitamin C (0.25-4.0 g) for 1 week and serum parameters were assayed prior to the experiment and on the eighth day of consumption. Blood samples were collected 4, 12 and 24 h after vitamin C intake. Serum levels of ascorbic acid increased significantly after vitamin C ingestion inhibiting urate and total bilirubin tests 4 and 12 h after intake (P < 0.01). A significant negative interference occurred up to 24 h after consumption of 4 g vitamin C for the urate test. In contrast, ingestion of vitamin C did not show interference in glucose, triglyceride and cholesterol tests. Addition of ascorbic acid to serum inhibited the urate test to a similar extent to that observed after vitamin C intake. However, after ingesting vitamin C, the interference for the bilirubin test was greater than that of the in vitro interference. Commonly taken doses of supplementary vitamin C interfered negatively with the serum urate test based on the Trinder method, and with bilirubin metabolism.

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

PubMed

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

2018-06-14

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

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

PubMed

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

2007-11-01

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

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

PubMed

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

2006-06-01

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

Gastrointestinal parasites of feral cats from Christmas Island.

PubMed

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

2008-01-01

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

Cross-neutralization between three mumps viruses & mapping of haemagglutinin-neuraminidase (HN) epitopes.

PubMed

Vaidya, Sunil R; Dvivedi, Garima M; Jadhav, Santoshkumar M

2016-01-01

The reports from the countries where mumps vaccine is given as routine immunization suggest differences in mumps virus neutralizing antibody titres when tested with vaccine and wild type viruses. Such reports are unavailable from countries like India where mumps vaccine is not included in routine immunization. We, therefore, undertook this study to understand the cross-neutralization activity of Indian mumps viruses. By using commercial mumps IgG enzyme immunoassay (EIA) and a rapid focus reduction neutralization test (FRNT), a panel of serum samples was tested. The panel consisted of 14 acute and 14 convalescent serum samples collected during a mumps outbreak and 18 archived serum samples. Two wild types (genotypes C and G) and Leningrad-Zagreb vaccine strain (genotype N) were used for the challenge experiments and FRNT titres were determined and further compared. The HN protein sequence of three mumps viruses was analyzed for the presence of key epitopes. All serum samples effectively neutralized mumps virus wild types and a vaccine strain. However, significantly lower FRNT titres were noted to wild types than to vaccine strain (P<0.05). The comparison between EIA and FRNT results revealed 95.6 per cent agreement. No amino acid changes were seen in the epitopes in the Indian wild type strains. All potential N-linked glycosylation sites were observed in Indian strains. Good cross-neutralization activity was observed for three mumps virus strains, however, higher level of FRNT titres was detected for mumps virus vaccine strain compared to Indian wild type isolates.

Avian influenza H9N2 seroprevalence among pig population and pig farm staff in Shandong, China.

PubMed

Li, Song; Zhou, Yufa; Zhao, Yuxin; Li, Wenbo; Song, Wengang; Miao, Zengmin

2015-03-01

Shandong province of China has a large number of pig farms with the semi-enclosed houses, allowing crowds of wild birds to seek food in the pig houses. As the carriers of avian influenza virus (AIV), these wild birds can easily pass the viruses to the pigs and even the occupational swine-exposed workers. However, thus far, serological investigation concerning H9N2 AIV in pig population and pig farm staff in Shandong is sparse. To better understand the prevalence of H9N2 AIV in pig population and pig farm staff in Shandong, the serum samples of pigs and occupational pig-exposed workers were collected and tested for the antibodies for H9N2 AIV by both hemagglutination inhibition (HI) and micro-neutralization (MN) assays. When using the antibody titers ≥40 as cut-off value, 106 (HI: 106/2176, 4.87%) and 84 (MN: 84/2176, 3.86%) serum samples of pigs were tested positive, respectively; 6 (HI: 6/287, 2.09%) and 4 (MN: 4/287, 1.39%) serum samples of the pig farm staff were positive, respectively; however, serum samples from the control humans were tested negative in both HI and MN assays. These findings revealed that there were H9N2 AIV infections in pig population and pig farm staff in Shandong, China. Therefore, it is of utmost importance to conduct the long-term surveillance of AIV in pig population and the pig farm staff.

Detection of Wuchereria bancrofti DNA in paired serum and urine samples using polymerase chain reaction-based systems.

PubMed

Ximenes, Camila; Brandão, Eduardo; Oliveira, Paula; Rocha, Abraham; Rego, Tamisa; Medeiros, Rafael; Aguiar-Santos, Ana; Ferraz, João; Reis, Christian; Araujo, Paulo; Carvalho, Luiz; Melo, Fabio L

2014-12-01

The Global Program for the Elimination of Lymphatic Filariasis (GPELF) aims to eliminate this disease by the year 2020. However, the development of more specific and sensitive tests is important for the success of the GPELF. The present study aimed to standardise polymerase chain reaction (PCR)-based systems for the diagnosis of filariasis in serum and urine. Twenty paired biological urine and serum samples from individuals already known to be positive for Wuchereria bancrofti were collected during the day. Conventional PCR and semi-nested PCR assays were optimised. The detection limit of the technique for purified W. bancrofti DNA extracted from adult worms was 10 fg for the internal systems (WbF/Wb2) and 0.1 fg by using semi-nested PCR. The specificity of the primers was confirmed experimentally by amplification of 1 ng of purified genomic DNA from other species of parasites. Evaluation of the paired urine and serum samples by the semi-nested PCR technique indicated only two of the 20 tested individuals were positive, whereas the simple internal PCR system (WbF/Wb2), which has highly promising performance, revealed that all the patients were positive using both samples. This study successfully demonstrated the possibility of using the PCR technique on urine for the diagnosis of W. bancrofti infection.

[Cultivation of a permanent fish cell line in serum-free media special experiences with a cytotoxicity test for waste water samples

PubMed

Kohlpoth, Martin; Rusche, Brigitte

1997-01-01

The use of fetal calf serum (FCS) as standard medium additive for the cell cultivation must be regarded critically from the point of view of animal welfare as well as for scientific reasons and makes it necessary to look for alternatives. In the last years an in vitro cytotoxicity assay for the testing of industrial waste waters with the permanent fish cell line RTG-2 was established and pre-validated as an alternative to the fish test with the golden orfe. The application of FCS is also a special problem with regard to the testing of waste waters in a cytotoxicity test so that FCS-alternatives were tested. The RTG-2 cells were successfully adapted to the two solvents Basal Medium Supplement (BMS) and Ultroser-G (U-G) that are used to replace serum. The characterisation of these adapted cell lines showed no significant differences in growth rate, adhesion rate, viability and sensitivity to chemicals in comparison to the original RTG-2 cells. On the determination of the cytotoxicity of industrial waste waters the RTG-2 cells adapted to the BMS medium indicated a clearly higher toxicity of the waste water samples than the original RTG-2 cells. This result confirms the thesis that serum components react with waste water elements and thus change the bio-availability of toxic compounds.

Evaluation of two 4th generation point-of-care assays for the detection of Human Immunodeficiency Virus infection.

PubMed

Stafylis, Chrysovalantis; Klausner, Jeffrey D

2017-01-01

Fourth generation assays detect simultaneously antibodies for HIV and the p24 antigen, identifying HIV infection earlier than previous generation tests. Previous studies have shown that the Alere Determine HIV-1/2 Combo has lower than anticipated performance in detecting antibodies for HIV and the p24 antigen. Furthermore, there are currently very few studies evaluating the performance of Standard Diagnostics BIOLINE HIV Ag/Ab Combo. To evaluate the performance of the Alere Determine HIV-1/2 Combo and the Standard Diagnostics BIOLINE HIV Ag/Ab Combo in a panel of frozen serum samples. The testing panel included 133 previously frozen serum specimens from the UCLA Clinical Microbiology & Immunoserology laboratory. Reference testing included testing for HIV antibodies by a 3rd generation enzyme immunoassay followed by HIV RNA detection. Antibody negative and RNA positive sera were also tested by a laboratory 4th generation HIV Ab/Ag enzyme immunoassay. Reference testing yielded 97 positives for HIV infection and 36 negative samples. Sensitivity of the Alere test was 95% (88-98%), while the SD Bioline sensitivity was 91% (83-96%). Both assays showed 100% (90-100%) specificity. No indeterminate or invalid results were recorded. Among 13 samples with acute infection (HIV RNA positive, HIV antibody negative), 12 were found positive by the first assay and 8 by the second. The antigen component of the Alere assay detected 10 acute samples, while the SD Bioline assay detected only one. Both rapid assays showed very good overall performance in detecting HIV infection in frozen serum samples, but further improvements are required to improve the performance in acute infection.

An improved reference measurement procedure for triglycerides and total glycerides in human serum by isotope dilution gas chromatography-mass spectrometry.

PubMed

Chen, Yizhao; Liu, Qinde; Yong, Sharon; Teo, Hui Ling; Lee, Tong Kooi

2014-01-20

Triglycerides are widely tested in clinical laboratories using enzymatic methods for lipid profiling. As enzymatic methods can be affected by interferences from biological samples, this together with the non-specific nature of triglycerides measurement makes it necessary to verify the accuracy of the test results with a reference measurement procedure. Several such measurement procedures had been published. These procedures generally involved lengthy and laborious sample preparation steps. In this paper, an improved reference measurement procedure for triglycerides and total glycerides was reported which simplifies the sample preparation steps and greatly shortens the time taken. The procedure was based on isotope dilution gas chromatography-mass spectrometry (IDGC-MS)with tripalmitin as the calibration standard. Serum samples were first spiked with isotope-labeled tripalmitin. For the measurement of triglycerides, the serum samples were subjected to lipid extraction followed by separation of triglycerides from diglycerides and monoglycerides. Triglycerides were then hydrolyzed to glycerol, derivatized and injected into the GC–MS for quantification. For the measurement of total glycerides, the serum samples were hydrolyzed directly and derivatized before injection into the GC-MS for quantification. All measurement results showed good precision with CV <1%. A certified reference material (CRM) of lipids in frozen human serum was used to verify the accuracy of the measurement. The obtained values for both triglycerides and total glycerides were well within the certified ranges of the CRM, with deviation <0.4% from the certified values. The relative expanded uncertainties were also comparable with the uncertainties associated with the certified values of the CRM. The validated procedure was used in an External Quality Assessment (EQA) Program organized by our laboratory to establish the assigned values for triglycerides and total glycerides.

Multicenter Comparison of Serum and Whole-Blood Specimens for Detection of Aspergillus DNA in High-Risk Hematological Patients

PubMed Central

Springer, Jan; Morton, C. O.; Perry, Michael; Heinz, Werner J.; Paholcsek, Melinda; Alzheimer, Mona; Rogers, T. R.; Barnes, Rosemary A.; Einsele, Hermann; White, P. Lewis

2013-01-01

Samples from patients at high risk for invasive aspergillosis (IA) were prospectively collected and analyzed for the presence of molecular markers of fungal infection. Serum specimens were screened for galactomannan and Aspergillus DNA, and whole-blood specimens were screened only for Aspergillus DNA. Fungal infections were categorized according to the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group, National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) criteria. Forty-seven cases (proven and probable IA) and 31 controls (no evidence of IA) were selected retrospectively for this case-control study, comprising 803 samples, in order to determine the performance of whole-blood PCR, serum PCR, and serum galactomannan testing. Although no single assay was able to detect every case of IA, a combination of different assays provided the best performance. There was no significant difference between the use of whole-blood and serum specimens for PCR-based diagnosis of IA, but there was a trend for whole blood to be more sensitive (85% versus 79%) and to yield an earlier positive result (36 days versus 15 days) than for serum. However, DNA extraction from serum specimens is easier and faster than that from whole-blood specimens, and it allows the same specimen to be used for both galactomannan and PCR assays. In conclusion, the appropriate sample type for DNA extraction should be determined by the local requirements and the technical platforms available at each individual center. A combination of biomarker tests offered the best diagnostic utility for detecting IA. PMID:23426930

Sensitivity of mouse bioassay in clinical wound botulism.

PubMed

Wheeler, Charlotte; Inami, Gregory; Mohle-Boetani, Janet; Vugia, Duc

2009-06-15

California has an ongoing epidemic of wound botulism (WB) among injection drug users (IDUs). We retrospectively studied a cohort of patients with WB and determined the sensitivity of the mouse bioassay-the gold standard laboratory test for confirmation of botulism-in verifying WB. We defined a clinical case of WB as an acute, bilateral, descending, flaccid paralysis starting with 1 cranial nerve palsies in an IDU with no other explainable diagnosis. We calculated the sensitivity of the mouse bioassay as the proportion of clinical WB cases that had positive serum toxin test results by mouse bioassay. We compared serum toxin-positive with serum toxin-negative patients. Of 73 patients with WB, 50 tested serum toxin positive, yielding a sensitivity of 68%. Serum toxin-positive patients did not differ significantly from serum toxin-negative patients with respect to demographic characteristics or injection drug use practices or in days from patient symptom onset to collection of specimens for testing. Patients did not differ significantly by clinical characteristics, except that serum toxin-positive patients were more likely than serum toxin-negative patients to have required mechanical ventilation during their hospital courses (74% vs. 43%; P = .01). In this study, the mouse bioassay failed to detect botulinum toxin in the serum samples of nearly one-third of IDUs with characteristic WB. Such patients should be considered to have probable WB. Physicians should be aware of the test's limitations and base their final diagnosis of suspected WB on clinical criteria when the mouse bioassay produces negative results.

[Sterility in drained serums; before their use in emergency wards and in non-hospital emergency use].

PubMed

Carrasco, Juan Antonio Péculo; Escorza, Juana Gómez; Sánchez, María Del Mar Casal; Bouza, Mónica Rodríguez; Sanchez, José Luis Avecilla; Ruiz, Hugo José Rodríguez

2008-01-01

The authors detail the characteristics for the sterility of drained serums placed in a mobile emergency service unit and they determine how much time they can maintain optimum conditions for the use of these serums by means of a controlled, random, triple blind clinical test in which the units studied were serums prepared by an infusion system and a three-way valve. Data were gathered during a 12 month study period. The authors designed six operating groups based on their exposure time and where they were kept. These samples totaled 672 units. The authors carried out 12 microbiological cultural analyses for each unit; these analyses were tested for bacteria and fungi, both aerobic and anaerobic. The authors contrasted their hypothesis with a multivariable logistic regression model used as the variable for sterility A value of p < 0.05 was considered to be significant. Results were obtained on 669 serums, 98.82% of samples. Colonization was found in 1.7% of the 8028 cultures analyzed; in only 2 cases was there a register greater than 10 UFC/ml. This colonization corresponded to 103 serums which were then classified as non-sterile. For sterility the logistic regression model did not show any significant association with the various operating groups, which were controlled by means of two possible variable methods. The authors did not find evidence that recently drained serums were different, from the point of view of sterility, than the serums exposed during 24, 48 or 72 hours but, as well as for the groups, the incidences of these serums tripled the risk of colonization. The authors concluded they could use serums prepared previously and these serums sterility were guaranteed to the same degree as recently drained serums.

Oxidative stress, d-ROMs test, and ceruloplasmin.

PubMed

Colombini, Francesco; Carratelli, Mauro; Alberti, Angelo

2016-01-01

Human serum samples were evaluated for oxidative stress with the d-ROMs test. The ceruloplasmin (CP) and copper contents of the samples was independently measured and compared to those calculated on the basis of the d-ROMs test results for pure CP solutions. The d-ROMs readings did not show any significant correlation with either the CP or copper contents of the samples. Critical interference of CP on the d-ROMs test was therefore excluded and the usefulness of the test in the evaluation of global oxidative status of a biological sample could be reassessed.

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

PubMed

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

2016-12-01

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

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

PubMed Central

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

2016-01-01

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

Serological diagnosis of bovine brucellosis using B. melitensis strain B115.

PubMed

Corrente, Marialaura; Desario, Costantina; Parisi, Antonio; Grandolfo, Erika; Scaltrito, Domenico; Vesco, Gesualdo; Colao, Valeriana; Buonavoglia, Domenico

2015-12-01

Bovine brucellosis is diagnosed by official tests, such as Rose Bengal plate test (RBPT) and Complement Fixation test (CFT). Both tests detect antibodies directed against the lipolysaccharide (LPS) of Brucella cell wall. Despite their good sensitivity, those tests do not discriminate between true positive and false positive serological reactions (FPSR), the latter being generated by animals infected with other Gram negative microorganisms that share components of Brucella LPS. In this study, an antigenic extract from whole Brucella melitensis B115 strain was used to set up an ELISA assay for the serological diagnosis of bovine brucellosis. A total of 148 serum samples from five different groups of animals were tested: Group A: 28 samples from two calves experimentally infected with Yersinia enterocolitica O:9; Group B: 30 samples from bovines infected with Brucella abortus; Group C: 50 samples from brucellosis-free herds; Group D: 20 samples RBPT positive and CFT negative; Group E: 20 samples both RBPT and CFT positive. Group D and Group E serum samples were from brucellosis-free herds. Positive reactions were detected only by RBPT and CFT in calves immunized with Y. enterocolitica O:9. Sera from Group B animals tested positive also in the ELISA assay, whereas sera from the remaining groups were all negative. The results obtained encourage the use of the ELISA assay to implement the serological diagnosis of brucellosis. Copyright © 2015 Elsevier B.V. All rights reserved.

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

PubMed

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

2015-01-01

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

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

PubMed Central

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

2015-01-01

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

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

PubMed

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

2018-08-15

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

Quantitative assessment of anthrax vaccine immunogenicity using the dried blood spot matrix.

PubMed

Schiffer, Jarad M; Maniatis, Panagiotis; Garza, Ilana; Steward-Clark, Evelene; Korman, Lawrence T; Pittman, Phillip R; Mei, Joanne V; Quinn, Conrad P

2013-03-01

The collection, processing and transportation to a testing laboratory of large numbers of clinical samples during an emergency response situation present significant cost and logistical issues. Blood and serum are common clinical samples for diagnosis of disease. Serum preparation requires significant on-site equipment and facilities for immediate processing and cold storage, and significant costs for cold-chain transport to testing facilities. The dried blood spot (DBS) matrix offers an alternative to serum for rapid and efficient sample collection with fewer on-site equipment requirements and considerably lower storage and transport costs. We have developed and validated assay methods for using DBS in the quantitative anti-protective antigen IgG enzyme-linked immunosorbent assay (ELISA), one of the primary assays for assessing immunogenicity of anthrax vaccine and for confirmatory diagnosis of Bacillus anthracis infection in humans. We have also developed and validated high-throughput data analysis software to facilitate data handling for large clinical trials and emergency response. Published by Elsevier Ltd.

Dengue-yellow fever sera cross-reactivity; challenges for diagnosis.

PubMed

Houghton-Triviño, Natalia; Montaña, Diana; Castellanos, Jaime

2008-01-01

The Flavivirus genera share epitopes inducing cross-reactive antibodies leading to great difficulty in differentially diagnosing flaviviral infections. This work was aimed at evaluating the complexity of dengue and yellow fever serological differential diagnosis. Dengue antibody capture ELISA and a yellow fever neutralisation test were carried out on 13 serum samples obtained from yellow fever patients, 20 acute serum samples from dengue patients and 19 voluntary serum samples pre- and post-vaccination with YF vaccine. Dengue ELISA revealed IgM reactivity in 46,2 % of yellow fever patients and 42 % of vaccinees. Sixteen out of 20 dengue patients (80 %) had high YF virus neutralisation titres. Such very high cross-reactivity data challenged differential laboratory diagnosis of dengue and yellow fever in areas where both flaviviruses co-circulate. New laboratory strategies are thus needed for improving the tests and providing a specific laboratory diagnosis. Cross-reactivity between Flaviviruses represents a great difficulty for epidemiological surveillance and preventing dengue, both of which demand urgent attention.

Silver Nanoscale Hexagonal Column Chips for Detecting Cell-free DNA and Circulating Nucleosomes in Cancer Patients.

PubMed

Ito, Hiroaki; Hasegawa, Katsuyuki; Hasegawa, Yuuki; Nishimaki, Tadashi; Hosomichi, Kazuyoshi; Kimura, Satoshi; Ohba, Motoi; Yao, Hiroshi; Onimaru, Manabu; Inoue, Ituro; Inoue, Haruhiro

2015-05-21

Blood tests, which are commonly used for cancer screening, generally have low sensitivity. Here, we developed a novel rapid and simple method to generate silver nanoscale hexagonal columns (NHCs) for use in surface-enhanced Raman scattering (SERS). We reported that the intensity of SERS spectra of clinical serum samples obtained from gastrointestinal cancer patients is was significantly higher than that of SERS spectra of clinical serum samples obtained from non-cancer patients. We estimated the combined constituents on silver NHCs by using a field emission-type scanning electron microscope, Raman microscopes, and a 3D laser scanning confocal microscope. We obtained the Raman scattering spectra of samples of physically fractured cells and clinical serum. No spectra were obtained for chemically lysed cultured cells and DNA, RNA, and protein extracted from cultured cells. We believe that our method, which uses SERS with silver NHCs to detect circulating nucleosomes bound by methylated cell-free DNA, may be successfully implemented in blood tests for cancer screening.

Automated measurement of carbohydrate-deficient transferrin using the Bio-Rad %CDT by the HPLC test on a Variant HPLC system: evaluation and comparison with other routine procedures.

PubMed

Schellenberg, François; Mennetrey, Louise; Girre, Catherine; Nalpas, Bertrand; Pagès, Jean Christophe

2008-01-01

In this study, we evaluated the new %CDT by the HPLC method (Bio-Rad, Germany) on a Varianttrade mark HPLC system (Bio-Rad), checked the correlation with well-known methods and calculated the diagnostic value of the test. Intra-run and day-to-day precision values were calculated for samples with extreme serum transferrin concentrations, high trisialotransferrin and interfering conditions (haemolysed, lactescent and icteric samples). The method was compared with two routine procedures, the %CDT TIA (Bio-Rad, Hercules, CA, USA) and the Capillarystrade mark CDT (Sebia, France). A total of 350 clinical sera samples were used for a case-control study. Precision values were better in high CDT and medium CDT pools than in low CDT pools. The serum transferrin concentration had no effect on CDT measurement, except in samples with serum transferrin <1 g/L. Haemolysis was the only interfering situation. The method showed high correlation (r(2) > 0.95) with the two other methods (%CDT TIA and CZE %CDT). The global predictive value of the test was >0.90 at 1.9% cut-off. These results demonstrate that the %CDT by the HPLC test is suitable for CDT routine measurement; the results from the high-throughput Varianttrade mark system are well correlated with other methods and are of high diagnostic value.

Comparison of serum Concentration of Se, Pb, Mg, Cu, Zn, between MS patients and healthy controls

PubMed Central

Alizadeh, Anahita; Mehrpour, Omid; Nikkhah, Karim; Bayat, Golnaz; Espandani, Mahsa; Golzari, Alireza; Jarahi, Lida; Foroughipour, Mohsen

2016-01-01

Introduction Multiple Sclerosis (MS) is defined as one of the inflammatory autoimmune disorders and is common. Its exact etiology is unclear. There are some evidences on the role of environmental factors in susceptible genetics. The aim of this study is to evaluate the possible role of Selenium, Zinc, Copper, Lead and Magnesium metals in Multiple Sclerosis patients. Methods In the present analytical cross-sectional study, 56 individuals including 26 patients and 30 healthy controls were enrolled in the evaluation. The serum level of Se, Zn, Cu, Pb were quantified in graphite furnace conditions and flame conditions by utilizing an atomic absorption Perkin Elmer spectrophotometer 3030. The serum levels of Mg were measured by auto analyzer 1500 BT. The mean level of minerals (Zn, Pb, Cu, Mg, Se) in serum samples were compared in both cases and controls. The mean level of minerals (Zn, Pb, Cu, Mg, Se) in serum samples were compared in both cases and controls by using independent-samples t-test for normal distribution and Mann-Whitney U test as a non-parametric test. All statistical analyses were carried out using SPSS 11.0. Results As well as the Zn, Cu, and Se, there was no significant difference between MS patients and healthy individuals in Pb concentrations (p-value = 0.11, 0.14, 0.32, 0.20 respectively) but the level of Mg was significantly different (p= 0.001). Conclusion All serum concentrations of Zn, Pb, Se, Cu in both groups were in normal ranges and there was no difference in MS patients compared with the healthy group who were matched in genetics. Blood level of Mg was significantly lower in MS patients. But it should be noted that even with the low level of serum magnesium in MS patients, this value is still in the normal range. PMID:27757186

Evaluation of YB-1 levels in patients with endometriosis.

PubMed

Ahrens, Thorben; Silveira, Cassia G T; Banz-Jansen, Constanze; Rody, Achim; Hornung, Daniela

2015-08-01

The objective of this study is the evaluation of serum YB-1 levels in the diagnosis of endometriosis. Serum samples of 12 patients with histologically confirmed endometriosis and of 10 control patients were collected. Western blot analysis was used to assess serum YB-1 levels. Groups were compared with Student's t-test or, if not normally distributed, with the Mann-Whitney test. Sensitivity and specificity for the potential diagnostic performance of serum YB-1 were assessed by receiver operating characteristic (ROC) curves. Serum YB-1 levels were significantly higher in patients with endometriosis (=0.004). The area under the curve was 0.867 (95% confidence interval 0.714-1.019) with sensitivity and specificity of 83.3% and 70% respectively. Serum YB-1 levels in patients with endometriosis are significantly higher compared to control patients and may be used as a potential diagnostic biomarker for endometriosis. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

9 CFR 147.7 - Standard test procedures for mycoplasma. 5

Code of Federal Regulations, 2011 CFR

2011-01-01

... plate agglutination test, the tube agglutination test, and the enzyme-linked immunosorbent assay (ELISA... accurate however, and are useful in evaluating serum samples that react with the ELISA, plate, and/or tube...

9 CFR 147.7 - Standard test procedures for mycoplasma. 5

Code of Federal Regulations, 2012 CFR

2012-01-01

... plate agglutination test, the tube agglutination test, and the enzyme-linked immunosorbent assay (ELISA... accurate however, and are useful in evaluating serum samples that react with the ELISA, plate, and/or tube...

9 CFR 147.7 - Standard test procedures for mycoplasma. 5

Code of Federal Regulations, 2010 CFR

2010-01-01

... plate agglutination test, the tube agglutination test, and the enzyme-linked immunosorbent assay (ELISA... accurate however, and are useful in evaluating serum samples that react with the ELISA, plate, and/or tube...

9 CFR 147.7 - Standard test procedures for mycoplasma. 5

Code of Federal Regulations, 2014 CFR

2014-01-01

... plate agglutination test, the tube agglutination test, and the enzyme-linked immunosorbent assay (ELISA... accurate however, and are useful in evaluating serum samples that react with the ELISA, plate, and/or tube...

9 CFR 147.7 - Standard test procedures for mycoplasma. 5

Code of Federal Regulations, 2013 CFR

2013-01-01

... plate agglutination test, the tube agglutination test, and the enzyme-linked immunosorbent assay (ELISA... accurate however, and are useful in evaluating serum samples that react with the ELISA, plate, and/or tube...

Metabolic Footprinting of Fermented Milk Consumption in Serum of Healthy Men

PubMed Central

Pimentel, Grégory; Burton, Kathryn J; von Ah, Ueli; Bütikofer, Ueli; Pralong, François P; Vionnet, Nathalie; Portmann, Reto; Vergères, Guy

2018-01-01

Abstract Background Fermentation is a widely used method of natural food preservation that has consequences on the nutritional value of the transformed food. Fermented dairy products are increasingly investigated in view of their ability to exert health benefits beyond their nutritional qualities. Objective To explore the mechanisms underpinning the health benefits of fermented dairy intake, the present study followed the effects of milk fermentation, from changes in the product metabolome to consequences on the human serum metabolome after its ingestion. Methods A randomized crossover study design was conducted in 14 healthy men [mean age: 24.6 y; mean body mass index (in kg/m2): 21.8]. At the beginning of each test phase, serum samples were taken 6 h postprandially after the ingestion of 800 g of a nonfermented milk or a probiotic yogurt. During the 2-wk test phases, subjects consumed 400 g of the assigned test product daily (200 g, 2 times/d). Serum samples were taken from fasting participants at the end of each test phase. The serum metabolome was assessed through the use of LC-MS–based untargeted metabolomics. Results Postprandial serum metabolomes after milk or yogurt intake could be differentiated [orthogonal projections to latent structures discriminant analysis (OPLS-DA) Q2 = 0.74]. Yogurt intake was characterized by higher concentrations of 7 free amino acids (including proline, P = 0.03), reduced concentrations of 5 bile acids (including glycocholic acid, P = 0.04), and modulation of 4 indole derivative compounds (including indole lactic acid, P = 0.01). Fasting serum samples after 2 wk of daily intake of milk or yogurt could also be differentiated based on their metabolic profiles (OPLS-DA Q2 = 0.56) and were discussed in light of the postprandial results. Conclusion Metabolic pathways related to amino acids, indole derivatives, and bile acids were modulated in healthy men by the intake of yogurt. Further investigation to explore novel health effects of fermented dairy products is warranted.This trial was registered at clinicaltrials.gov as NCT02230345. PMID:29788433

[Applying competitive polymerase chain reaction to the detection of hepatitis B virus DNA].

PubMed

Wang, Ling; Yang, Peng; Li, Shuang-qing; Xu, Shu-hui; Cao, Gui-qun; Zhang, Fa-qiang; Zhang, Mei-xia; Chen, Qing-ying; Xia, Qing-jie; Liu, Kai; Tang, Fang; Zhang, Yuan-zheng

2004-11-01

To reduce the rate of accidental false negative result in the HBV DNA PCR test on clinical serum samples. A competitive polymerase chain reaction (C-PCR) was used to decrease the false negative ratio. In the C-PCR, a constructed inner control DNA was added for co-amplification with the HBV target DNA. In a 20 microl C-PCR system, about 60 to 200 copies of inner control DNA could give apparent co-amplification signal band after electrophoresis on a 2% agarose gel. Five of 120 samples of clinical serum (4.2%) could not be amplified. C-PCR has the advantage of yielding information on false negative in the HBV DNA PCR assay of clinical serum samples.

Serological and Molecular Biological Studies of Parvovirus B19, Coxsackie B Viruses, and Adenoviruses as Potential Cardiotropic Viruses in Bulgaria.

PubMed

Ivanova, Stefka Kr; Angelova, Svetla G; Stoyanova, Asya P; Georgieva, Irina L; Nikolaeva-Glomb, Lubomira K; Mihneva, Zafira G; Korsun, Neli St

2016-12-01

Inflammatory diseases of the heart (myocarditis, pericarditis) are commonly caused by viruses. Among the human cardiotropic viruses, parvovirus B19, Coxsackie B viruses, and adenoviruses play a leading role. The aim of the present study was to determine the presumptive causative role of parvovirus B19, Coxsackie B viruses, and adenoviruses in the development of myocarditis, pericarditis and dilated cardiomyopathy by demonstrating the presence of specific antiviral antibodies or viral DNA in patients' serum samples. We tested serum samples collected between 2010 and 2014 from 235 patients with myocarditis (n=108), pericarditis (n=79), myopericarditis (n=19), dilated cardiomyopathy (n=7), and fever of unknown origin accompanied by cardiac complaints (n=22). The mean age of patients with the standard deviation was 33 ± 18 years. Serological and molecular methods (ELISA for specific IgM/IgG antibodies to parvovirus B19 and IgM antibodies to Coxsackie B viruses and adenoviruses, and PCR for detection of parvovirus B19 in serum samples, respectively) were used in the study. Of all tested 235 serum samples, in 60 (25.5%) positive results for at least one of the three tested viruses were detected. Forty out of these 235 serum samples (17%) were Coxsackie B virus IgM positive. They were found in 17% (18/108) of the patients with myocarditis, in 15% (12/79) of those with pericarditis, in 16% (3/19) of those with myopericarditis and in 32% (7/22) in those with fever of unknown origin. The 63 Coxsackie B virus IgM negative patient's serum samples were tested by ELISA for presence of adenovirus IgM antibodies. Such were found in 4 patients with pericarditis and in 2 patients with fever of unknown origin. Every IgM negative sample (n=189) for Coxsackie B and adenovirus was further tested by ELISA for parvovirus B19 IgM/IgG antibodies. B19-IgM antibodies were detected in 14 patients (7.4%). The percentages for B19-IgM antibodies was 8% (7/90), 5% (3/63) and 31% (4/13) in the patients affected with myocarditis, pericarditis, and fever of unknown origin, respectively. Protective B19-IgG antibodies were found in 108 (57%) of the samples. A B19-PCR signal was detected in all the patients who were B19-IgM positive, and in only 1 patient with positive B19-IgG result, the latter presenting with dilated cardiomyopathy. The present study shows the involvement of Coxsackie B, parvovirus B19 and adenoviruses in the development of inflammatory diseases of the heart (myocarditis and pericarditis). It is the first ever study in the country that simultaneously analyzes the prevalence of the three major human cardiotropic viruses.

Human West Nile virus infection in Bosnia and Herzegovina.

PubMed

Ahmetagić, Sead; Petković, Jovan; Hukić, Mirsada; Smriko-Nuhanović, Arnela; Piljić, Dilista

2015-02-01

To describe the first two cases of West Nile virus (WNV) neuroinvasive infections in Bosnia and Herzegovina. At the Clinic for Infectious Diseases of the University Clinical Centre Tuzla, Bosnia and Herzegovina (BiH), specific screening for WNV infection was performed on patients with neuroinvasive diseases from 1 August to 31 October 2013. Serum samples were tested for the presence of WNV IgM and IgG antibodies using enzyme-linked immunosorbent assay (ELISA); positive serum samples were further analyzed by detection of WNV nucleic acid of two distinct lineages (lineage 1 and lineage 2) in sera by RT-PCR. Three (out of nine) patients met clinical criteria, and two of them had high serum titre of WNV specific IgM antibodies (3.5 and 5.2). Serum RT-PCR testing was negative. Conformation by neutralization testing was not performed. Both cases represented with encephalitis. None of these cases had recent travel history in WNW endemic areas, or history of blood transfusion and organ transplantation, so they represented autochthonous cases. Although there were no previous reports of flavivirus infections in BiH, described cases had high titre of WNV specific antibodies in serum, and negative flavivirus-vaccination history, they were defined as probable cases because recommended testing for case confirmation was not performed. The West Nile virus should be considered a possible causative pathogen in this area, probably in patients with mild influenza-like disease of unknown origin and those with neuroinvasive disease during late summer and early autumn.

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

PubMed

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

2017-10-01

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

Serologic survey for Toxoplasma gondii in wild animals in Florida.

PubMed

Burridge, M J; Bigler, W J; Forrester, D J; Hennemann, J M

1979-11-01

Blood samples were collected for serum separation from 114 species of wild animals (25 species of mammals, 82 species of birds, and 7 species of reptiles) in Florida. Each of the 3,471 samples was tested for antibodies to Toxoplasma gondii, using the indirect hemagglutination test. The highest prevalences of T gondii antibodies were 19% in armadillos (Dasypus novemcinctus), 18% in raccoons (Procyon lotor), 13% in black rats (Rattus rattus), and 11% in opossums (Didelphis marsupialis). Antibody prevalences were significantly higher in male than in female raccoons (P less than 0.05) and in adult than in nonadult raccoons and opossums (P less than 0.005). A high proportion of seropositive animals was found in three other mammalian species: 4 of 4 black bears (Ursus americanus), 2 of 3 bobcats (Lynx rufus), and 2 of 8 Norway rats (Rattus norvegicus) tested. Antibodies were found in 8 of the 1,279 avian serums; they were not found in any of the 13 reptilian serums tested. There were no significant geographic variations in antibody prevalence in any species.

Serum galactomannan in cystic fibrosis patients colonized with Aspergillus species.

PubMed

Warren, Thomas A; Yau, Yvonne; Ratjen, Felix; Tullis, Elizabeth; Waters, Valerie

2012-08-01

Cystic fibrosis patients are often colonized by Aspergillus species. Sera from 138 pediatric and adult cystic fibrosis patients were tested for the presence of galactomannan. All serum samples were negative for galactomannan and there was no difference among patients who were chronically, intermittently, and never colonized with Aspergillus.

Mass screening for Trypanosoma cruzi infections using the immunofluorescence, ELISA and haemagglutination tests on serum samples and on blood eluates from filter-paper.

PubMed Central

Zicker, F.; Smith, P. G.; Luquetti, A. O.; Oliveira, O. S.

1990-01-01

Methods used to diagnose Trypanosoma cruzi infection differ in their ability to discriminate between sera from infected and uninfected individuals. We compared the results of an immunofluorescence (IF) test, a haemagglutination (HA) test, and an enzyme-linked immunosorbent assay (ELISA) for the diagnosis of T. cruzi infections in a large population-based survey in central Brazil using blood eluates from filter-paper and venous blood samples. The sensitivities of the tests on eluates, compared with results on serum samples, were low: ELISA (78.1%), IF (69.2%) and HA (64.6%). The level of agreement between the tests on eluates was very poor, with the best co-positivity for IF and ELISA. Both the positive and negative predictive values of the three tests on eluates were similar (around 96%) to those for sera. Higher co-positivity values were obtained for the three tests on sera. The implications of these results are discussed in relation to blood screening, routine medical practice, sero-epidemiological surveys, and the follow-up of patients admitted to therapeutic trials. PMID:2119903

Mass screening for Trypanosoma cruzi infections using the immunofluorescence, ELISA and haemagglutination tests on serum samples and on blood eluates from filter-paper.

PubMed

Zicker, F; Smith, P G; Luquetti, A O; Oliveira, O S

1990-01-01

Methods used to diagnose Trypanosoma cruzi infection differ in their ability to discriminate between sera from infected and uninfected individuals. We compared the results of an immunofluorescence (IF) test, a haemagglutination (HA) test, and an enzyme-linked immunosorbent assay (ELISA) for the diagnosis of T. cruzi infections in a large population-based survey in central Brazil using blood eluates from filter-paper and venous blood samples. The sensitivities of the tests on eluates, compared with results on serum samples, were low: ELISA (78.1%), IF (69.2%) and HA (64.6%). The level of agreement between the tests on eluates was very poor, with the best co-positivity for IF and ELISA. Both the positive and negative predictive values of the three tests on eluates were similar (around 96%) to those for sera. Higher co-positivity values were obtained for the three tests on sera. The implications of these results are discussed in relation to blood screening, routine medical practice, sero-epidemiological surveys, and the follow-up of patients admitted to therapeutic trials.

Severe reactions from roe without concomitant fish allergy.

PubMed

Mäkinen-Kiljunen, Soili; Kiistala, Raija; Varjonen, Elina

2003-10-01

Although fish allergy is common, no studies have been published on allergy to fish roe. To describe 2 cases of IgE-mediated allergy to 2 roe species. Two patients, one with local symptoms and the other with anaphylaxis following ingestion of roe, underwent skin prick testing (SPT) with 2 roe species, whitefish roe (WFR) and rainbow trout roe (RTR). Serum samples were taken for IgE measurement and immunoblotting to identify roe allergens. Inhibition studies were performed to scrutinize the cross-reactivity between the roes and to fish. The results of the SPTs with the roes were clearly positive in both patients but negative in control persons. The results of SPTs to all other foods were negative. Roe-specific IgE levels were elevated in the serum samples of both patients. Immunoblotting revealed different IgE-binding patterns of the extracts and different inhibition profiles with the serum samples. In WFR blotting, both serum samples detected a heavy IgE-binding band at approximately 20 kDa, which was not inhibited with fish. Cross-reactivity between the roes was demonstrated in the patient with local symptoms from RTR but not in the patient with anaphylaxis from WFR. The first serum sample also detected several IgE-binding bands in the RTR blot, the most intensive at 21 to 23 kDa and 30 kDa, which were partially inhibited by WFR and more completely with fish. The anaphylaxis patient did not detect allergens in the RTR blot. After the investigation, the patients have remained symptom free and able to consume all kinds of fish without problems. IgE-mediated allergy to roe is possible without concomitant fish allergy. Roe allergy should be explored in patients who test negative to fish but are suspected of having seafood-related allergy.

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

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

Liu, Tao; Hossain, Mahmud; Schepmoes, Athena A.

2012-08-03

Sandwich immunoassay is the standard technique used in clinical labs for quantifying protein biomarkers for disease detection, monitoring and therapeutic intervention. Albeit highly sensitive, the development of a specific immunoassay is rather time-consuming and associated with extremely high cost due to the requirement for paired immunoaffinity reagents of high specificity. Recently, mass spectrometry-based methods, specifically selected reaction monitoring mass spectrometry (SRM-MS), have been increasingly applied to measure low abundance biomarker candidates in tissue and biofluids, owing to high sensitivity and specificity, simplicity of assay configuration, and great multiplexing capability. In this study, we report for the first time the developmentmore » of immunoaffinity depletion-based workflows and SRM-MS assays that enable sensitive and accurate quantification of total and free prostate-specific antigen (PSA) in serum without the requirement for specific PSA antibodies. With stable isotope dilution and external calibration, low ng/mL level detection of both total and free PSA was consistently achieved in both PSA-spiked female serum samples and actual patient serum samples. Moreover, comparison of the results obtained when SRM PSA assays and conventional immunoassays were applied to the same samples showed very good correlation (R2 values ranging from 0.90 to 0.99) in several independent clinical serum sample sets, including a set of 33 samples assayed in a blinded test. These results demonstrate that the workflows and SRM assays developed here provide an attractive alternative for reliably measuring total and free PSA in human blood. Furthermore, simultaneous measurement of free and total PSA and many other biomarkers can be performed in a single analysis using high-resolution liquid chromatographic separation coupled with SRM-MS.« less

[Serological diagnosis of dengue and yellow fever infections in suspected cases from Pará State, Brazil, 1999].

PubMed

de Araújo, Tais Pinheiro; Rodrigues, Sueli Guerreiro; Costa, Maria Irene Weyl de A; Vasconcelos, Pedro Fernando da Costa; da Rosa, Amélia P A Travassos

2002-01-01

From June to December 1999, 785 serum samples were obtained from patients clinically suspected of having dengue or yellow fever. The patients were referred by public health centers distributed within the six mesoregions of Par State, Brazil. Serum samples were tested for Flavivirus antibodies by hemagglutination inhibition test and for dengue and yellow fever viruses by enzyme-linked immunosorbent assay for IgM detection. Of the sera collected, 563 (71.7%) were positive by HI test and out of these 150 (26.6%) were positive by ELISA-IgM. Dengue virus was responsible for most of the recent infections in all regions; yellow fever cases detected in the current study were restricted to the Maraj and Southeast regions.

HCV-RNA quantification in liver bioptic samples and extrahepatic compartments, using the abbott RealTime HCV assay.

PubMed

Antonucci, FrancescoPaolo; Cento, Valeria; Sorbo, Maria Chiara; Manuelli, Matteo Ciancio; Lenci, Ilaria; Sforza, Daniele; Di Carlo, Domenico; Milana, Martina; Manzia, Tommaso Maria; Angelico, Mario; Tisone, Giuseppe; Perno, Carlo Federico; Ceccherini-Silberstein, Francesca

2017-08-01

We evaluated the performance of a rapid method to quantify HCV-RNA in the hepatic and extrahepatic compartments, by using for the first time the Abbott RealTime HCV-assay. Non-tumoral (NT), tumoral (TT) liver samples, lymph nodes and ascitic fluid from patients undergoing orthotopic-liver-transplantation (N=18) or liver resection (N=4) were used for the HCV-RNA quantification; 5/22 patients were tested after or during direct acting antivirals (DAA) treatment. Total RNA and DNA quantification from tissue-biopsies allowed normalization of HCV-RNA concentrations in IU/μg of total RNA and IU/10 6 liver-cells, respectively. HCV-RNA was successfully quantified with high reliability in liver biopsies, lymph nodes and ascitic fluid samples. Among the 17 untreated patients, a positive and significant HCV-RNA correlation between serum and NT liver-samples was observed (Pearson: rho=0.544, p=0.024). Three DAA-treated patients were HCV-RNA "undetectable" in serum, but still "detectable" in all tested liver-tissues. Differently, only one DAA-treated patient, tested after sustained-virological-response, showed HCV-RNA "undetectability" in liver-tissue. HCV-RNA was successfully quantified with high reliability in liver bioptic samples and extrahepatic compartments, even when HCV-RNA was "undetectable" in serum. Abbott RealTime HCV-assay is a good diagnostic tool for HCV quantification in intra- and extra-hepatic compartments, whenever a bioptic sample is available. Copyright © 2017 Elsevier B.V. All rights reserved.

[Seroprevalance of brucellosis, listeriosis and toxoplasmosis in cattle in Adana province of Turkey].

PubMed

Yağcı Yücel, Sükran; Yaman, Mehmet; Kurt, Cemal; Babür, Cahit; Celebi, Bekir; Kılıç, Selçuk; Ozen, Doğukan

2014-06-01

This study was conducted to identify the seroprevalance of diseases which are zoonotic and responsible from abortion such as toxoplasmosis, listeriosis and brucellosis in Holstein crossbred cattle of different age and sex in Adana province, between 2008 April-September. For this purpose, blood serum samples were collected from 132 cattle and analyzed for Toxoplasma gondii, Listeria monocytogenes and Brucella abortus antibodies. T. gondii, L. monocytogenes and Brucella abortus antibodies were determined by the standard Sabin- Feldman Dye Test (SFDT), Osebold method and Microtube Agglutination Test (MAT) respectively, from the blood serum samples. 132 serum tested 56.06% samples of T. gondii, 40.9% and 3.03% of L. monocytogenes and defined the B.abortus antibodies were found to be seropositive terms. There were no statistically significant difference between seropositive T. gondii, L. monocytogenes and B.abortus antibodies among age groups (p>0.05). In this study, for the first time in cattle in the region of Adana serological methods revealed the presence L. monocytogenes, toxoplasmosis and listeriosis were higher than brucellosis seropositivity. Moreover, the prevalence of these diseases in the same animal at the highest rate was determined for T. gondii and L. monocytogenes.

Cross-neutralization between three mumps viruses & mapping of haemagglutinin-neuraminidase (HN) epitopes

PubMed Central

Vaidya, Sunil R.; Dvivedi, Garima M.; Jadhav, Santoshkumar M.

2016-01-01

Background & objectives: The reports from the countries where mumps vaccine is given as routine immunization suggest differences in mumps virus neutralizing antibody titres when tested with vaccine and wild type viruses. Such reports are unavailable from countries like India where mumps vaccine is not included in routine immunization. We, therefore, undertook this study to understand the cross-neutralization activity of Indian mumps viruses. Methods: By using commercial mumps IgG enzyme immunoassay (EIA) and a rapid focus reduction neutralization test (FRNT), a panel of serum samples was tested. The panel consisted of 14 acute and 14 convalescent serum samples collected during a mumps outbreak and 18 archived serum samples. Two wild types (genotypes C and G) and Leningrad-Zagreb vaccine strain (genotype N) were used for the challenge experiments and FRNT titres were determined and further compared. The HN protein sequence of three mumps viruses was analyzed for the presence of key epitopes. Results: All serum samples effectively neutralized mumps virus wild types and a vaccine strain. However, significantly lower FRNT titres were noted to wild types than to vaccine strain (P<0.05). The comparison between EIA and FRNT results revealed 95.6 per cent agreement. No amino acid changes were seen in the epitopes in the Indian wild type strains. All potential N-linked glycosylation sites were observed in Indian strains. Interpretation & conclusions: Good cross-neutralization activity was observed for three mumps virus strains, however, higher level of FRNT titres was detected for mumps virus vaccine strain compared to Indian wild type isolates. PMID:26997012

Evaluation of rapid diagnostic test kits for feline leukemia virus infection using samples from naturally infected cats.

PubMed

Liu, Jiayou; O'Connor, Thomas; Beall, Melissa; Chandrashekar, Ramaswamy; Lappin, Michael

2016-01-01

Feline leukemia virus (FeLV) is a potentially life-threatening oncogenic retrovirus. The p27 viral core protein is produced by the virus in infected feline cells, is found in the cytoplasm in several blood cells and can be free in the serum and plasma. ELISA or particle-based immunoassay are commonly used to detect the presence of the p27 core protein in samples obtained from blood. The objective of this study was to compare the performance of several in-clinic tests: the SNAP Feline Triple Test (IDEXX Laboratories), the WITNESS FeLV-FIV Test (Zoetis) and the VetScan Feline FeLV/FIV Rapid Test (Abaxis). The sample population (100 positive, 105 negative samples) consisted of serum and plasma samples submitted to IDEXX's worldwide reference laboratory for feline retrovirus testing. Virus isolation and reverse transcriptase PCR results were not available and so samples were judged to be positive or negative based on the results of the ViraCHEK FeLV (Zoetis) microtiter plate assay. The percentage of samples positive and negative for FeLV p27 antigen using the three in-clinic tests compared with the ViraCHEK method were as follows: IDEXX Feline Triple (positive 98.0%, negative 100%); Zoetis WITNESS (positive 79.0%, negative 97.1%); Abaxis VetScan (positive 73.0%, negative 97.1%). The SNAP Feline Triple Test demonstrated a high level of agreement for FeLV-positive and FeLV-negative samples when assessed in this model. Results of FeLV assays can vary among tests.

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

PubMed

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

2004-04-01

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

Gender difference of serum angiotensin-converting enzyme (ACE) activity in DD genotype of ACE insertion/deletion polymorphism in elderly Chinese.

PubMed

Zhang, Ya-Feng; Cheng, Qiong; Tang, Nelson L S; Chu, Tanya T W; Tomlinson, Brian; Liu, Fan; Kwok, Timothy C Y

2014-12-01

In this study we investigated the gender difference of serum angiotensin-converting enzyme (ACE) activity in a population of Hong Kong-dwelling elderly Chinese. A total of 1767 (843 male, 924 female) Hong Kong-dwelling elderly Chinese were recruited. ACE I/D genotypes were identified by polymerase chain reaction amplification and serum ACE activity was determined using a commercially available kinetic kit. ACE I/D genotype distribution was compared by chi-square test, the correlation between ACE I/D polymorphism and serum ACE activity was analysed by ANOVA test and gender difference of serum ACE activity of different genotypes was compared by independent sample t-test. No statistically significant difference of genotype distribution between male and female subjects was found. Serum ACE activity was significantly correlated with ACE genotype. Overall, there was no gender difference of serum ACE activity; however, when sub-grouping the subjects by ACE I/D genotype, male subjects with DD genotype had higher serum ACE activity than female subjects with DD genotype. No significant gender difference of genotype distribution was found in elderly Chinese. Serum ACE activity was significantly correlated with ACE I/D polymorphism in elderly Chinese. Male subjects with DD genotype had higher serum ACE activity than female subjects with DD genotype. © The Author(s) 2013.

Comparative Diagnosis of Serum IgG1 and Coproantigen ELISA for Fasciolosis Detection of Goats in Mexico.

PubMed

Villa-Mancera, Abel; Molina-Mendoza, Pedro; Hernández-Guzmán, Karina; Olivares-Pérez, Jaime; Sarracent-Pérez, Jorge; Zumaquero-Ríos, José

2016-01-01

The objective of present study was to determine the prevalence of natural caprine fasciolosis in the Mixteca region of Mexico using coproantigen and serum IgG1 ELISA tests for comparative purposes. A total of 1070 serum and faecal samples were analyzed for IgG1 antibodies and coproantigens, using ELISA with E/S products as antigen and a monoclonal antibody-based sandwich ELISA. Prevalence of 73.46% was found using the serological ELISA and a percentage of 77.20 was found for coproantigen ELISA. The diagnostic sensitivity and specificity for serum ELISA were 86.7% and 96.4%, and for the coproantigen ELISA they were 93.1% and 97.8%, respectively. The seropositive samples were further categorized as low, medium, or high positivity. Results show a great proportion of low and medium positive goats when the serum ELISA test was used. Correlation coefficients between coproantigens and seropositivity were statistically significant (P < 0.01) for low seropositivity (r = 0.93) and medium seropositivity (r = 0.84). The accuracy of faecal antigen ELISA was higher compared to indirect ELISA serological test. Two ELISAs were shown to be useful for demonstrating the current status of F. hepatica infection in the endemic areas and can be employed in studies on epidemiology as well as anthelmintics treatment for preventing economic loss and the risk of transmission to humans.

Comparative Diagnosis of Serum IgG1 and Coproantigen ELISA for Fasciolosis Detection of Goats in Mexico

PubMed Central

Molina-Mendoza, Pedro; Hernández-Guzmán, Karina; Olivares-Pérez, Jaime; Sarracent-Pérez, Jorge; Zumaquero-Ríos, José

2016-01-01

The objective of present study was to determine the prevalence of natural caprine fasciolosis in the Mixteca region of Mexico using coproantigen and serum IgG1 ELISA tests for comparative purposes. A total of 1070 serum and faecal samples were analyzed for IgG1 antibodies and coproantigens, using ELISA with E/S products as antigen and a monoclonal antibody-based sandwich ELISA. Prevalence of 73.46% was found using the serological ELISA and a percentage of 77.20 was found for coproantigen ELISA. The diagnostic sensitivity and specificity for serum ELISA were 86.7% and 96.4%, and for the coproantigen ELISA they were 93.1% and 97.8%, respectively. The seropositive samples were further categorized as low, medium, or high positivity. Results show a great proportion of low and medium positive goats when the serum ELISA test was used. Correlation coefficients between coproantigens and seropositivity were statistically significant (P < 0.01) for low seropositivity (r = 0.93) and medium seropositivity (r = 0.84). The accuracy of faecal antigen ELISA was higher compared to indirect ELISA serological test. Two ELISAs were shown to be useful for demonstrating the current status of F. hepatica infection in the endemic areas and can be employed in studies on epidemiology as well as anthelmintics treatment for preventing economic loss and the risk of transmission to humans. PMID:27563665

Use of a recombinant burkholderia intracellular motility a protein for immunodiagnosis of glanders.

PubMed

Kumar, Subodh; Malik, Praveen; Verma, Shailendra Kumar; Pal, Vijai; Gautam, Vandana; Mukhopadhyay, Chiranjay; Rai, Ganga Prasad

2011-09-01

Glanders, caused by the Gram-negative, nonmotile bacterium Burkholderia mallei, is a contagious and highly fatal disease of equines. During the last decade, the number of glanders outbreaks has increased steadily. The disease also has high zoonotic significance and B. mallei is listed biological warfare agent. The complement fixation test (CFT) is a routinely used and internationally recognized test to screen equine sera for the glanders. However, discrepant results have been observed using the CFT. The low sensitivity and specificity of the CFT and enzyme-linked immunosorbent assay (ELISA) have been linked to the use of crude test antigens. We expressed a novel recombinant Burkholderia intracellular motility A (rBimA) protein in Escherichia coli for the diagnosis of equine glanders. Purified rBimA was used in an indirect ELISA format. All of the 21 true-positive serum samples used in the study tested positive, whereas only 17 of the 1,524 potentially negative sera tested positive by indirect ELISA, thus exhibiting 100% sensitivity and 98.88% specificity. Also, rBimA protein did not react with melioidosis patient and normal healthy human serum samples, showing its high specificity. The developed assay can be used as a simple and rapid tool for diagnosis of glanders in equine serum samples. An Indian patent (1328/DEL/2010) has been filed for the reagent.

Detection of reticuloendotheliosis virus as a contaminant of fowl pox vaccines.

PubMed

Awad, A M; Abd El-Hamid, H S; Abou Rawash, A A; Ibrahim, H H

2010-11-01

This study was designed to detect reticuloendotheliosis virus (REV) as a contaminant in fowl pox vaccines. A total of 30 fowl pox vaccine samples were examined for the presence of REV using both in vitro and in vivo methods. In in vitro testing, the fowl pox vaccine samples were inoculated into chicken embryo fibroblast cultures prepared from specific-pathogen-free embryonated chicken eggs, and the cultures were examined using PCR to detect REV. In in vivo testing, each fowl pox vaccine sample was inoculated into 5-d-old specific-pathogen-free chicks, which were kept under observation for up to 12 wk postinoculation; serum samples were collected at 15, 30, and 45 d postinoculation for the detection of REV-specific antibodies using ELISA. Tissue samples were collected at 8 and 12 wk postinoculation for histopathological examination. Of the tested vaccines, only one imported vaccine sample tested positive for REV using PCR. Serum samples collected from chicks infected with the PCR-positive vaccine batch also tested positive for REV-specific antibodies using ELISA. Histopathological examination of the liver, spleen, and bursa of Fabricius demonstrated the presence of tumor cells in these organs, confirming the results obtained using PCR and ELISA, and indicating that the sample was contaminated with REV. These data clearly indicate that the screening of all commercial poultry vaccines for viruses is an important factor in assuring the biosafety of animal vaccines.

Twenty-four cases of imported zika virus infections diagnosed by molecular methods.

PubMed

Alejo-Cancho, Izaskun; Torner, Nuria; Oliveira, Inés; Martínez, Ana; Muñoz, José; Jane, Mireia; Gascón, Joaquim; Requena-Méndez, Ana; Vilella, Anna; Marcos, M Ángeles; Pinazo, María Jesús; Gonzalo, Verónica; Rodriguez, Natalia; Martínez, Miguel J

2016-10-01

Zika virus is an emerging flavivirus widely spreading through Latin America. Molecular diagnosis of the infection can be performed using serum, urine and saliva samples, although a well-defined diagnostic algorithm is not yet established. We describe a series of 24 cases of imported zika virus infection into Catalonia (northeastern Spain). Based on our findings, testing of paired serum and urine samples is recommended. Copyright © 2016 Elsevier Inc. All rights reserved.

Bluetongue virus in Lebanon.

PubMed

El Hage, J; Lorusso, A; Carmine, I; Di Gennaro, A; Portanti, O; Olivieri, S; Casaccia, C; Pisciella, M; Teodori, L; Sghaier, S; Savini, G

2013-10-01

Since 2000, several incursions of bluetongue virus (BTV) occurred in the Mediterranean Basin involving European and surrounding Countries. The Middle East represents one of the most important gateways for the access of BTV in Europe. Limited data on the BTV situation in this area are available. In this perspective, an epidemiological survey on the presence of BTV in Lebanon was conducted. Of the 181 serum samples tested, 97 (mean = 53.6%; 95% CI: 46.3-60.7) resulted positive when tested for the presence of BTV antibodies by c-ELISA, of these 42 (mean = 42%; 95% CI: 32.8-51.8) serum samples were from sheep and 55 (mean = 67.9%; 95% CI: 57.1-77.1) serum samples were from goats. Fourteen blood samples (14/110; mean = 12.7%; 95% CI: 7.8-20.3), 6 (6/66; mean = 9.1%; 95% CI: 4.4-18.5) from sheep and 8 (8/44; mean = 18.2%; 95% CI: 9.6-32.0) from goats, were positive by qRT-PCR. The results with serum-neutralization assay and typing performed by RT-PCR confirmed that six BTV serotypes are currently circulating in Lebanon, and these serotypes are as follows: 1, 4, 6, 8, 16 and 24. This study is the first report that confirms the presence and circulation of BTV in Lebanon. © 2013 Blackwell Verlag GmbH.

Sub-tesla-field magnetization of vibrated magnetic nanoreagents for screening tumor markers

NASA Astrophysics Data System (ADS)

Chieh, Jen-Jie; Huang, Kai-Wen; Shi, Jin-Cheng

2015-02-01

Magnetic nanoreagents (MNRs), consisting of liquid solutions and magnetic nanoparticles (MNPs) coated with bioprobes, have been widely used in biomedical disciplines. For in vitro tests of serum biomarkers, numerous MNR-based magnetic immunoassay methods or schemes have been developed; however, their applications are limited. In this study, a vibrating sample magnetometer (VSM) was used for screening tumor biomarkers based on the same MNRs as those used in other immunoassay methods. The examination mechanism is that examined tumor biomarkers are typically conjugated to the bioprobes coated on MNPs to form magnetic clusters. Consequently, the sub-Tesla-field magnetization (Msub-T) of MNRs, including magnetic clusters, exceeds that of MNRs containing only separate MNPs. For human serum samples, proteins other than the targeted biomarkers induce the formation of magnetic clusters with increased Msub-T because of weak nonspecific binding. In this study, this interference problem was suppressed by the vibration condition in the VSM and analysis. Based on a referenced Msub-T,0 value defined by the average Msub-T value of a normal person's serum samples, including general proteins and few tumor biomarkers, the difference ΔMsub-T between the measured Msub-T and the reference Msub-T,0 determined the expression of only target tumor biomarkers in the tested serum samples. By using common MNRs with an alpha-fetoprotein-antibody coating, this study demonstrated that a current VSM can perform clinical screening of hepatocellular carcinoma.

Promoter methylation of MCAM, ERα and ERβ in serum of early stage prostate cancer patients.

PubMed

Brait, Mariana; Banerjee, Mithu; Maldonado, Leonel; Ooki, Akira; Loyo, Myriam; Guida, Elisa; Izumchenko, Evgeny; Mangold, Leslie; Humphreys, Elizabeth; Rosenbaum, Eli; Partin, Alan; Sidransky, David; Hoque, Mohammad Obaidul

2017-02-28

Prostate cancer (PC) is the second most common cancer among men worldwide. Currently, the most common non-invasive approach for screening and risk assessment of PC is measuring the level of serum prostate-specific antigen (PSA). However, the sensitivity of PSA is 42.8 % and specificity is 41.1%. As a result, the serum PSA test leads to numerous unneeded biopsies. Therefore, a rigorous search for biomarkers for early detection of PC is ongoing. In this study, we aim to assess a panel of epigenetic markers in an intend to develop an early detection test for PC. The sensitivity and specificity of hypermethylation of MCAM was 66% and 73% respectively which is an improvement from the sensitivity and specificity of PSA. Considering a combination marker panel of MCAM, ERα and ERβ increased the sensitivity to 75% and the specificity became 70% for the minimally invasive early detection test of PC. Sixteen primary matched tumor and serum were analyzed by quantitative methylation specific PCR (QMSP) to determine analytical and clinical sensitivity of the genes tested (SSBP2, MCAM, ERα, ERβ, APC, CCND2, MGMT, GSTP1, p16 and RARβ2). Additionally, serum samples from eighty four cases of PC, thirty controls and seven cases diagnosed as high grade Prostatic Intraepithelial Neoplasia (HGPIN) were analyzed. Promoter methylation of MCAM, ERα and ERβ have a potential to be utilized as biomarker for the early detection of prostate PC as their sensitivity and specificity seem to be better than serum PSA in our cohort of samples. After robust validation in a larger prospective cohort, our findings may reduce the numbers of unwarranted prostate biopsies.

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

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

Monroy, Rocio; Morrison, Katherine; Teo, Koon

2008-09-15

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

Evaluation of a Serum Lung Cancer Biomarker Panel.

PubMed

Mazzone, Peter J; Wang, Xiao-Feng; Han, Xiaozhen; Choi, Humberto; Seeley, Meredith; Scherer, Richard; Doseeva, Victoria

2018-01-01

A panel of 3 serum proteins and 1 autoantibody has been developed to assist with the detection of lung cancer. We aimed to validate the accuracy of the biomarker panel in an independent test set and explore the impact of adding a fourth serum protein to the panel, as well as the impact of combining molecular and clinical variables. The training set of serum samples was purchased from commercially available biorepositories. The testing set was from a biorepository at the Cleveland Clinic. All lung cancer and control subjects were >50 years old and had smoked a minimum of 20 pack-years. A panel of biomarkers including CEA (carcinoembryonic antigen), CYFRA21-1 (cytokeratin-19 fragment 21-1), CA125 (carbohydrate antigen 125), HGF (hepatocyte growth factor), and NY-ESO-1 (New York esophageal cancer-1 antibody) was measured using immunoassay techniques. The multiple of the median method, multivariate logistic regression, and random forest modeling was used to analyze the results. The training set consisted of 604 patient samples (268 with lung cancer and 336 controls) and the testing set of 400 patient samples (155 with lung cancer and 245 controls). With a threshold established from the training set, the sensitivity and specificity of both the 4- and 5-biomarker panels on the testing set was 49% and 96%, respectively. Models built on the testing set using only clinical variables had an area under the receiver operating characteristic curve of 0.68, using the biomarker panel 0.81 and by combining clinical and biomarker variables 0.86. This study validates the accuracy of a panel of proteins and an autoantibody in a population relevant to lung cancer detection and suggests a benefit to combining clinical features with the biomarker results.

Evaluation of a Serum Lung Cancer Biomarker Panel

PubMed Central

Mazzone, Peter J; Wang, Xiao-Feng; Han, Xiaozhen; Choi, Humberto; Seeley, Meredith; Scherer, Richard; Doseeva, Victoria

2018-01-01

Background: A panel of 3 serum proteins and 1 autoantibody has been developed to assist with the detection of lung cancer. We aimed to validate the accuracy of the biomarker panel in an independent test set and explore the impact of adding a fourth serum protein to the panel, as well as the impact of combining molecular and clinical variables. Methods: The training set of serum samples was purchased from commercially available biorepositories. The testing set was from a biorepository at the Cleveland Clinic. All lung cancer and control subjects were >50 years old and had smoked a minimum of 20 pack-years. A panel of biomarkers including CEA (carcinoembryonic antigen), CYFRA21-1 (cytokeratin-19 fragment 21-1), CA125 (carbohydrate antigen 125), HGF (hepatocyte growth factor), and NY-ESO-1 (New York esophageal cancer-1 antibody) was measured using immunoassay techniques. The multiple of the median method, multivariate logistic regression, and random forest modeling was used to analyze the results. Results: The training set consisted of 604 patient samples (268 with lung cancer and 336 controls) and the testing set of 400 patient samples (155 with lung cancer and 245 controls). With a threshold established from the training set, the sensitivity and specificity of both the 4- and 5-biomarker panels on the testing set was 49% and 96%, respectively. Models built on the testing set using only clinical variables had an area under the receiver operating characteristic curve of 0.68, using the biomarker panel 0.81 and by combining clinical and biomarker variables 0.86. Conclusions: This study validates the accuracy of a panel of proteins and an autoantibody in a population relevant to lung cancer detection and suggests a benefit to combining clinical features with the biomarker results. PMID:29371783

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

PubMed

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

2005-01-01

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

Biochemical correlates of neurosensory changes in weightlessness

NASA Technical Reports Server (NTRS)

Leach, Carolyn S.; Reschke, Millard F.

1989-01-01

The possible existence of a relationship between space motion sickness and chemical and biochemical variables measured in body fluids is studied. Clinical chemistry and endocrine measurements from blood and urine samples taken before and after Space Shuttle flights were analyzed along with the occurrence of SMS during flight and provocative testing before flight. Significant positive correlations were observed with serum chloride and significant negative correlations with serum phosphate, serum uric acid, and plasma thyroid stimulating hormone.

Spiking of serum specimens with exogenous reporter peptides for mass spectrometry based protease profiling as diagnostic tool.

PubMed

Findeisen, Peter; Peccerella, Teresa; Post, Stefan; Wenz, Frederik; Neumaier, Michael

2008-04-01

Serum is a difficult matrix for the identification of biomarkers by mass spectrometry (MS). This is due to high-abundance proteins and their complex processing by a multitude of endogenous proteases making rigorous standardisation difficult. Here, we have investigated the use of defined exogenous reporter peptides as substrates for disease-specific proteases with respect to improved standardisation and disease classification accuracy. A recombinant N-terminal fragment of the Adenomatous Polyposis Coli (APC) protein was digested with trypsin to yield a peptide mixture for subsequent Reporter Peptide Spiking (RPS) of serum. Different preanalytical handling of serum samples was simulated by storage of serum samples for up to 6 h at ambient temperature, followed by RPS, further incubation under standardised conditions and testing for stability of protease-generated MS profiles. To demonstrate the superior classification accuracy achieved by RPS, a pilot profiling experiment was performed using serum specimens from pancreatic cancer patients (n = 50) and healthy controls (n = 50). After RPS six different peak categories could be defined, two of which (categories C and D) are modulated by endogenous proteases. These latter are relevant for improved classification accuracy as shown by enhanced disease-specific classification from 78% to 87% in unspiked and spiked samples, respectively. Peaks of these categories presented with unchanged signal intensities regardless of preanalytical conditions. The use of RPS generally improved the signal intensities of protease-generated peptide peaks. RPS circumvents preanalytical variabilities and improves classification accuracies. Our approach will be helpful to introduce MS-based proteomic profiling into routine laboratory testing.

Preanalytical considerations in detection of colorectal cancer in blood serum using Raman molecular imaging (Conference Presentation)

NASA Astrophysics Data System (ADS)

Treado, Patrick J.; Stewart, Shona D.; Smith, Aaron; Kirschner, Heather; Post, Christopher; Overholt, Bergein F.

2016-03-01

Colorectal cancer (CRC) is the third most common cancer in men and women in the United States. Raman Molecular Imaging (RMI) is an effective technique to evaluate human tissue, cells and bodily fluids, including blood serum for disease diagnosis. ChemImage Corporation, in collaboration with clinicians, has been engaged in development of an in vitro diagnostic Raman assay focused on CRC detection. The Raman Assay for Colorectal Cancer (RACC) exploits the high specificity of Raman imaging to distinguish diseased from normal dried blood serum droplets without additional reagents. Pilot Study results from testing of hundreds of biobank patient samples have demonstrated that RACC detects CRC with high sensitivity and specificity. However, expanded clinical trials, which are ongoing, are revealing a host of important preanalytical considerations associated with sample collection, sample storage and stability, sample shipping, sample preparation and sample interferents, which impact detection performance. Results from recent clinical studies will be presented.

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

PubMed

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

2013-07-02

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

Bio-repository of post-clinical test samples at the national cancer center hospital (NCCH) in Tokyo.

PubMed

Furuta, Koh; Yokozawa, Karin; Takada, Takako; Kato, Hoichi

2009-08-01

We established the Bio-repository at the National Cancer Center Hospital in October 2002. The main purpose of this article is to show the importance and usefulness of a bio-repository of post-clinical test samples not only for translational cancer research but also for routine clinical oncology by introducing the experience of setting up such a facility. Our basic concept of a post-clinical test sample is not as left-over waste, but rather as frozen evidence of a patient's pathological condition at a particular point. We can decode, if not all, most of the laboratory data from a post-clinical test sample. As a result, the bio-repository is able to provide not only the samples, but potentially all related laboratory data upon request. The areas of sample coverage are the following: sera after routine blood tests; sera after cross-match tests for transfusion; serum or plasma submitted at a patient's clinically important time period by the physician; and samples collected by the individual investigator. The formats of stored samples are plasma or serum, dried blood spot (DBS) and buffy coat. So far, 150 218 plasmas or sera, 35 253 DBS and 536 buffy coats have been registered for our bio-repository system. We arranged to provide samples to various concerned parties under strict legal and ethical agreements. Although the number of the utilized samples was initially limited, the inquiries for sample utilization are now increasing steadily from both research and clinical sources. Further efforts to increase the benefits of the repository are intended.

[The importance of using a test to detect Chlamydia trachomatis infection in patients undergoing counseling and testing for sexually transmitted diseases at a public health center].

PubMed

Hatori, Toru; Nakamura, Tamiko; Tsukui, Satoshi

2013-01-01

Serum levels of antibody against Chlamydia trachomatis are used routinely to diagnose chlamydial infection among patients undergoing counseling and testing for sexually transmitted disease at public health centers in Japan, whereas tests for pathogen detection are conducted at most clinics. The objective of the present study was to evaluate the usefulness of a test for detecting C. trachomatis and to compare it with that of screening for serum chlamydia antibodies at a public health center. Serum chlamydia antibody titers were estimated using an ELISA, while C. trachomatis was detected in the urine using the nucleic acid amplification assay that included strand displacement amplification (SDA). We compared the results of 120 patients (64 men and 56 women), because the sensitivity of the method using urine samples from women has been reported to be as high as that using cervical samples. The serum antibody test results were considered to be positive when either one or both of the IgA and IgG antibodies were positive. The overall prevalence of positive results for the antibody test was 24.2% (14.1% for men and 35.7% for women), and that for the SDA method was 7.5% (3.1% for men and 12.5% for women). The concordance rate of the 2 tests was 81.7%, with a kappa statistic of 0.35 (95% CI, 0.10-0.59). Of the 9 subjects who were positive for C. trachomatis using the SDA assay, 1 had serum chlamydia IgA antibodies, 6 had IgG antibodies, 1 had both the antibodies, and 1 had no antibodies. On the other hand, of the 111 subjects who were negative for C. trachomatis, only 8 had chlamydia IgA antibodies, 5 had IgG antibodies, and 8 had both the antibodies. These results indicated there was no association between serum IgA antibodies and the presence of C. trachomatis. The presence of serum chlamydia antibodies was significantly higher in subjects with a past history of genital chlamydial infection than in those without a past history of infection (P<0.01). However, there was no difference in the results of the SDA assay between subjects with or without a previous infection. Thus, although serum chlamydia antibodies are generally measured at public health centers, tests for detecting C. trachomatis should also be used.

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

PubMed

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

2018-01-05

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

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

PubMed Central

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

2016-01-01

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

Incidence of anti-intermediate filament antibody in serum samples of students with suspected glandular fever.

PubMed

Kataaha, P K; Holborow, E J; Edwards, J M

1985-03-01

Serum samples from 40 students with suspected infectious mononucleosis were tested for the presence of antibodies to intermediate filaments (AIFA) of the cytoskeleton. Twenty had antibodies to the Epstein-Barr virus capsid antigen before their illness, and during it their sera remained negative by the Paul-Bunnell test. The other 20 patients did not have antibodies to the Epstein-Barr virus capsid antigen before their illness and seroconverted during the illness. These patients (true infectious mononucleosis group) developed positive Paul-Bunnell tests. Sera from normal subjects (blood donors) were also tested for AIFA. AIFA was present in titres greater than 1/10 in 80% of the infectious mononucleosis group (mean titre 1/40-1/80), 10% of the Paul-Bunnell negative glandular fever group, and 8.5% of the normal blood donors.

Fast and efficient detection of tuberculosis antigens using liposome encapsulated secretory proteins of Mycobacterium tuberculosis.

PubMed

Tiwari, Dileep; Haque, Shafiul; Tiwari, Ram P; Jawed, Arshad; Govender, Thavendran; Kruger, Hendrik G

2017-04-01

A rapid and efficient diagnostic test was developed for the detection of Mycobacterium tuberculosis antigens in serum samples of active tuberculosis (TB) and extrapulmonary TB patients via a liposomal agglutination-based method. A rapid card test has been developed to facilitate the recognition of high-affinity binding rabbit raised purified culture filtrate protein antibodies coupled on the surface of activated liposomal preparation. In the presence of TB antigens, the polyclonal antibodies bound to the liposomal particles demonstrate a visible agglutination reaction. The developed assay was simple, rapid, reliable, sensitive, and specific as a diagnostic test for the detection of antigens in serum samples of clinically confirmed cases of TB within 4-5 minutes' duration. The test was evaluated at different hospitals, medical colleges, and pathology centers, and involved 1483 participants. This investigation was conducted to detect the presence of these antigens during the period of active growth of the microorganism in serum samples for pulmonary TB and processed tissue biopsy for other extrapulmonary TB. Results obtained using this test were compared with acid-fast bacilli smear and culture results. Our study demonstrated that the newly developed liposome tuberculosis antigen card test detected antigens in our study population with approximately 97.48% sensitivity and 95.79% specificity. This is the first study to report the liposomal encapsulation of culture filtrate proteins from M. tuberculosis for diagnostic application. Copyright © 2015. Published by Elsevier B.V.

Comparative examination and validation of ELISA test systems for Salmonella typhimurium diagnosis of slaughtering pigs.

PubMed

Szabó, I; Scherer, K; Roesler, U; Appel, B; Nöckler, K; Hensel, A

2008-05-10

The most frequently isolated Salmonella serotype from pork in Germany is S. typhimurium, especially phagetype DT 104. The monitoring programs on Salmonella in swine are based on enzyme-linked immunoadsorbent assay (ELISA) detecting antibodies in serum or meat juice. These serological results are used to classify swine herds in three categories to assess the hygienic status of farm regarding Salmonella infection in pigs. The object of this study was the comparative evaluation of four indirect Salmonella ELISA tests approved in Germany to detect Salmonella typhimurium infection of swine. Three tests (A-C) are based on LPS-antigen and directed against specific IgG-antibodies. The fourth test (D) bases on a whole-cell-lysate antigen and discriminates between Salmonella specific IgA-, IgM- and IgG-antibodies. In a longitudinal study sixteen 6 weeks old weaning pigs were orally infected with S. typhimurium DT 104. During an observation period of 138d clinical and bacteriological parameters were monitored and serum samples obtained at regular intervals as well as meat juice samples taken at slaughter were examined by the respective ELISA systems. Study results reveal that all tested ELISA systems are able to detect S. typhimurium infection in pigs in both sample matrices, blood serum and meat juice whereas test D showed the highest sensitivity to detect Salmonella antibodies in pigs. The sensitivity to detect Salmonella antibodies varied between tests A and C according to the used cut-off (test specific cut-off vs. recommended surveillance cut-off) resulting in a change of seroprevalence and hence may influence the Salmonella status of the farm.

A large outbreak of hepatitis E among a displaced population in Darfur, Sudan, 2004: the role of water treatment methods.

PubMed

Guthmann, Jean-Paul; Klovstad, Hilde; Boccia, Delia; Hamid, Nuha; Pinoges, Loretxu; Nizou, Jacques-Yves; Tatay, Mercedes; Diaz, Francisco; Moren, Alain; Grais, Rebecca Freeman; Ciglenecki, Iza; Nicand, Elisabeth; Guerin, Philippe Jean

2006-06-15

The conflict in Darfur, Sudan, was responsible for the displacement of 1.8 million civilians. We investigated a large outbreak of hepatitis E virus (HEV) infection in Mornay camp (78,800 inhabitants) in western Darfur. To describe the outbreak, we used clinical and demographic information from cases recorded at the camp between 26 July and 31 December 2004. We conducted a case-cohort study and a retrospective cohort study to identify risk factors for clinical and asymptomatic hepatitis E, respectively. We collected stool and serum samples from animals and performed a bacteriological analysis of water samples. Human samples were tested for immunoglobulin G and immunoglobulin M antibody to HEV (for serum samples) and for amplification of the HEV genome (for serum and stool samples). In 6 months, 2621 hepatitis E cases were recorded (attack rate, 3.3%), with a case-fatality rate of 1.7% (45 deaths, 19 of which involved were pregnant women). Risk factors for clinical HEV infection included age of 15-45 years (odds ratio, 2.13; 95% confidence interval, 1.02-4.46) and drinking chlorinated surface water (odds ratio, 2.49; 95% confidence interval, 1.22-5.08). Both factors were also suggestive of increased risk for asymptomatic HEV infection, although this was not found to be statistically significant. HEV RNA was positively identified in serum samples obtained from 2 donkeys. No bacteria were identified from any sample of chlorinated water tested. Current recommendations to ensure a safe water supply may have been insufficient to inactivate HEV and control this epidemic. This research highlights the need to evaluate current water treatment methods and to identify alternative solutions adapted to complex emergencies.

Per- and polyfluoroalkyl substance (PFAS) exposure assessment in a community exposed to contaminated drinking water, New Hampshire, 2015.

PubMed

Daly, Elizabeth R; Chan, Benjamin P; Talbot, Elizabeth A; Nassif, Julianne; Bean, Christine; Cavallo, Steffany J; Metcalf, Erin; Simone, Karen; Woolf, Alan D

2018-04-01

Per- and polyfluoroalkyl substances (PFAS) are synthetic chemicals used in manufacturing that resist environmental degradation, can leach into drinking water, and bioaccumulate in tissues. Some studies have shown associations with negative health outcomes. In May 2014, a New Hampshire public drinking water supply was found to be contaminated with PFAS from a former U.S. Air Force base. We established a serum testing program to assess PFAS exposure in the affected community. Serum samples and demographic and exposure information were collected from consenting eligible participants. Samples were tested for PFAS at three analytical laboratories. Geometric means and 95% confidence intervals were calculated and analyzed by age and exposure variables. A total of 1578 individuals provided samples for PFAS testing; >94% were found to have perfluorooctane sulfonic acid (PFOS), perfluorooctanoic acid (PFOA), and perfluorohexane sulfonic acid (PFHxS) detectable in serum. Geometric mean serum concentrations of PFOS, PFOA, and PFHxS were 8.6 μg/L (95% CI:8.3-8.9), 3.1 μg/L (95% CI: 3.0-3.2), and 4.1 μg/L (95% CI: 3.9-4.3), respectively, which were statistically higher than the general U.S. Significant associations were observed between PFAS serum concentrations and age, time spent in the affected community, childcare attendance, and water consumption. PFOS, PFOA, and PFHxS were found in significantly higher levels in the affected population, consistent with PFAS drinking water contamination. Given increased recognition of PFAS contamination in the U.S, a coordinated national response is needed to improve access to biomonitoring and understand health impacts. Copyright © 2018 Elsevier GmbH. All rights reserved.

Evaluation of ID-PaGIA syphilis antibody test.

PubMed

Naaber, Paul; Makoid, Ene; Aus, Anneli; Loivukene, Krista; Poder, Airi

2009-01-01

Laboratory diagnosis of syphilis is usually accomplished by serology. There are currently a large number of different commercial treponemal tests available that vary in format, sensitivity and specificity. To evaluate the ID-PaGIA Syphilis Antibody Test as an alternative to other specific treponemal tests for primary screening or confirmation of diagnosis. Serum samples from healthy adults (n = 100) were used for detection of specificity of ID-PaGIA. To evaluate sensitivity of ID-PaGIA serum samples (n = 101) from patients with confirmed or suspected syphilis were tested for syphilis antibodies with FTA-Abs IgM, ID-PaGIA, ELISA IgM and TPHA tests. No false-positive results were found with ID-PaGIA. Sensitivity of various treponemal tests was the following: FTA-Abs IgM: 95.5%, ID-PaGIA and ELISA IgM: 94%, and TPHA 75%. The positive and negative predictive values of ID-PaGIA were 100 and 89.5%, respectively. Compared with other treponemal tests ID-PaGIA has excellent sensitivity and specificity.

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

PubMed Central

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

2013-01-01

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

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

PubMed

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

2008-08-20

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

Simple method for determining human serum 2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical scavenging activity - possible application in clinical studies on dietary antioxidants.

PubMed

Chrzczanowicz, Jacek; Gawron, Anna; Zwolinska, Anna; de Graft-Johnson, Jeffrey; Krajewski, Wojciech; Krol, Maciej; Markowski, Jaroslaw; Kostka, Tomasz; Nowak, Dariusz

2008-01-01

2,2-Diphenyl-1-picryl-hydrazyl (DPPH) radical decomposition in alcohol solution is widely used, characterizing plant antioxidants that can rise in serum after fruit and vegetable intake. However, this test failed reproducible results with serum due to protein precipitation. We describe the application of serum deproteinization with acetonitrile relating to the DPPH test. Assay sensitivity, linearity, repeatability and storage effect were determined in serum samples deproteinized with an equal volume of acetonitrile. Associations between the DPPH test and the ferric reducing ability of serum (FRAP) method, measuring total antioxidant potential, were evaluated in sera from 78 healthy non-smoking men. The effect of a single ingestion of 1 L of cloudy apple juice on the serum DPPH radical scavenging activity in healthy volunteers was also investigated. Assay linearity was within 5-25 microL (r=0.99, p<0.01). With 25 microL-deproteinized serum, coefficient of variation was 4.2% and detection limit was 0.5% of the initial amount of decomposed DPPH radical over 30 min incubation. There was no sera activity decrease over 14 days storage at -20 degrees C. Mean values of DPPH radical scavenging activity and FRAP obtained in human serum were 11.2+/-3.3% and 382.0+/-88.1 micromol/L, respectively. A positive significant linear correlation was observed between these two methods (r=0.42, p<0.01). Serum supplementation with 50 micromol/L of catechin, gallic acid, ascorbic acid or uric acid enhanced DPPH test results. One brisk serving of 1 L of apple juice caused a significant increment of serum DPPH radical scavenging activity (1.9+/-1.9%, p<0.01) in 12 healthy subjects 1 h after juice ingestion. Applicability of the DPPH test to deproteinized serum with acetonitrile revealed numerous advantages, validating its practicability, simplicity and cost effectiveness as a tool in the estimation of antioxidant status in humans.

Evaluation of a recombinant LigB protein of Leptospira interrogans serovar Canicola in an enzyme-linked immunosorbent assay for the serodiagnosis of bovine leptospirosis.

PubMed

Sankar, Surya; Harshan, Hiron M; Somarajan, S R; Srivastava, S K

2010-06-01

A recombinant leptospiral lipoprotein, LigB, was evaluated for use in the diagnosis of bovine leptospirosis by enzyme-linked immunosorbent assay (rLigB IgG ELISA). The standard reference test (Microscopic agglutination test, MAT) of 200 serum samples from cattle suspected of leptospirosis showed that 95 (47.5%) samples had positive agglutination titres, which ranged from 100 to 1600. In rLigB IgG ELISA, 49% of the samples were positive. Sensitivity of IgG ELISA for 95 bovine sera, which had MAT titres of greater than or equal to 100, were 100%. ELISA showed a specificity of 97.1% with 105 bovine sera, which were negative at a 1:50 dilution in MAT for Leptospira interrogans serovars. The results of ELISA and MAT correspond very good. When analytical specificity of IgG ELISA was evaluated using bovine serum samples from animals showing the serum antibodies to other pathogens, no cross-reaction was observed. Thus the recombinant LigB IgG ELISA can be used instead of the MAT as an aid to the diagnosis of bovine leptospirosis. Copyright 2009. Published by Elsevier India Pvt Ltd.

Analysis of anhydrous glucose and human serum assisted by Raman spectroscopy

NASA Astrophysics Data System (ADS)

González, Naara; Cerecedo-Núñez, H. H.; Padilla-Sosa, P.; Flores, Aaron; May, Manuel; Basurto, Miguel; Alcántara, Gabriela; Muñiz, Omar

2017-02-01

Raman spectroscopy has been considered like a potentially important clinical tool for real-time diagnosis of disease and evaluation of living tissue, whit the proposal to development noninvasive glucose measurements in a near future, with lower power than other reported studies, in this work are reported experimental tests made with a excitation source of semiconductor laser of 785 nm and 35 mW power. Measurements were made to different glucose concentrations, with variation from 50 mg/dL to 6000 mg/dL. For this, three intervals with different ranges of concentration were analyzed, these tests were put into plastic sampling cells, making incise the beam vertically on sample. In the same way measurements to serum human are reported, for healthy volunteers had 12 hours fasting and non-fasting conditions, with it's corresponding values of glucose taken through a conventional glucometer. Freeze-dried human serum was poured on object-holder, in the case of human serum reconstitute, it was used container in which were previously kept samples. Nine spectra per test were obtained and subsequently average was calculated, the spectra were studied in a range of 500 to 1700 cm-1. This work explores the intensity variation of the bands of glucose in 1065 cm-1 and 1127 cm-1 as a function of glucose concentration. In the obtained results, there observes a behavior with positive slope in both substances, interrelation being observed between the measurements, being promissory for non-invasive measurement.

Interference in diagnostic tests for brucellosis in cattle recently vaccinated against leptospirosis.

PubMed

Naves, Joao Helder Frederico de Faria; Rezende, Lais M; Ramos, Gabriel C; Soares, Pollyanna M; Tavares, Tatiane C F; França, Andre M S; Neves, Saira M N; Silva, Natascha A M; Lima-Ribeiro, Anna M C

2012-03-01

The aim of the current study was to verify if cattle vaccinated against leptospirosis may react in diagnostic tests for brucellosis. Sixty cows were divided into 5 groups, each comprising 12 animals. Four groups were given different vaccines against leptospirosis, while the control group received only saline. Two doses of vaccine were given, as recommended by the manufacturers. Serum samples were collected on the first day of immunization (day 0) and on postvaccination days 7, 14, 21, 28, 35, 42, 49, 56, 96, and 126. All the serum samples were tested for brucellosis and leptospirosis. Twenty animals were reactive at least once to the Rose Bengal test, but by day 96, no further reactions were elicited by this test. Twenty-six samples were reactive to the Rose Bengal test, but only 7 remained positive in confirmatory tests: 1 to the 2-mercaptoethanol test, 2 to the fluorescence polarization assay, and 6 to indirect enzyme-linked immunosorbent assays. None of the samples was reactive in the complement fixation test. None of the animals in the control group was reactive. A significant difference was found between the control group and the groups vaccinated against leptospirosis, according to Fisher exact test. However, the groups were found to respond independently of the vaccine brand. The results indicate that cattle vaccinated against leptospirosis may show reactivity on screening tests for brucellosis.

Serum adipokines and HIV viral replication in patients undergoing antiretroviral therapy

PubMed Central

Aramă, Victoria; Tilişcan, Cătălin; Ion, Daniela Adriana; Mihăilescu, Raluca; Munteanu, Daniela; Streinu-Cercel, Anca; Tudor, Ana Maria; Hristea, Adriana; Leoveanu, Viorica; Olaru, Ioana; Aramă, Ştefan Sorin

2012-01-01

Introduction Several studies have reported that cytokines secreted by adipose tissue (adipokines) may be linked to HIV replication. The aim of the study was to evaluate the relationship between HIV replication and serum levels of adipokines, in a Caucasian HIV-infected population of men and women undergoing complex antiretroviral therapy. Methods A cross-sectional study was conducted in an unselected sample of 77 HIV-1-positive patients. Serum adipokines levels were measured including circulating adiponectin, leptin, resistin, tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6). Patients were divided into two groups: Group 1 - with undetectable viral load and Group 2 - with persistent HIV viral replication. Differences between groups ? were tested using independent-sample t-test for Gaussian variables and Mann–Whitney–Wilcoxon test for non-parametric variables. Pearson's chi-squared test was used for correlation analysis. Results A total of 77 patients (age range: 17-65, mean: 32.5 years) including 44 men (57.1% men, age range: 17–63 years, mean: 34.1 years) and 33 women (42.9% women age range: 19–65 years, mean: 30.3 years) were included in the study. TNF-alpha had significantly higher serum levels in patients with detectable viral load (16.89 vs. 9.35 pg/mL), (p=0.043), but correlation analysis lacked statistical significance. Adiponectin had median serum levels of 9.22 ìg/mL in Group 1 vs. 16.50 ìg/mL in Group 2 but the results lacked statistical significance (p=0.059). Higher leptin, IL-6 and resistin serum levels were noted in patients with undetectable HIV viral load, without statistical significance. Conclusions The present study reported higher TNF-alpha serum levels in patients with persistent HIV viral load. We found no statistically significant correlations between adiponectin, leptin, resistin and IL-6 and HIV viral load in our Caucasian HIV-positive study population, undergoing antiretroviral therapy. PMID:24432258

Evaluation of a monoclonal antibody-based latex agglutination test for diagnosis of cryptococcosis: comparison with two tests using polyclonal antibodies.

PubMed Central

Temstet, A; Roux, P; Poirot, J L; Ronin, O; Dromer, F

1992-01-01

Cryptococcal antigen detection has become a routine biological test performed for patients with AIDS. The poor prognosis of cryptococcosis explains the need for reliable tests. We evaluated the performances of a newly commercialized agglutination test that uses a monoclonal antibody specific for cryptococcal capsular polysaccharide (Pastorex Cryptococcus; Sanofi-Diagnostics Pasteur, Marnes-la-Coquette, France) and compared them with those of tests that use polyclonal immune sera (Cryptococcal Antigen Latex Agglutination System, Meridian Diagnostics, Inc., Cincinnati, Ohio; and Crypto-LA, International Biological Labs Inc., Cranbury, N.J.). The sensitivities and specificities of the tests were compared by using purified polysaccharides and yeast suspensions. Clinical specimens (131 serum samples, 41 cerebrospinal fluid samples, 34 urine samples, and 19 bronchoalveolar lavage samples) from 87 human immunodeficiency virus-positive subjects with (40 patients) and without (47 patients) culture-proven cryptococcosis were retrospectively tested during a blinded study. The effect of pronase treatment of samples was assessed for Pastorex Cryptococcus and the Cryptococcal Antigen Latex Agglutination System, and the antigen titers were compared. Our results show that (i) during the screening, concordance among the three tests was 97%; (ii) the use of pronase enhanced both the sensitivities and specificities of the Pastorex Cryptococcus test; (iii) titers agreed for 67% of the cerebrospinal fluid samples and 60% of the serum samples; and (iv) cryptococcosis was detected equally well with Pastorex Cryptococcus and with the other tests, whatever the infecting serotype (A, B, or D). The meaning of in vitro sensitivity and the relationship between titers and sensitivity are discussed. The results show that Pastorex Cryptococcus is a rapid and reliable test for the detection of cryptococcal antigen in body fluids and suggest that kits cannot be used interchangeably to monitor antigen titers in patients. PMID:1400951

Long-lived immunity to canine core vaccine antigens in UK dogs as assessed by an in-practice test kit.

PubMed

Killey, R; Mynors, C; Pearce, R; Nell, A; Prentis, A; Day, M J

2018-01-01

To determine the utility of an in-practice test kit to detect protective serum antibody against canine distemper virus, canine adenovirus and canine parvovirus type 2 in a sample of the UK dog population. Serum samples from 486 dogs, last vaccinated between less than 1 month and 124 months previously, were tested with the VacciCheck™ test kit for protective antibodies against distemper, adenovirus and parvovirus type 2. A high proportion of the dogs tested (93·6%) had protective antibody against all three of the core vaccine antigens: 95·7% of the dogs were seropositive against canine distemper virus, 97·3% against canine adenovirus and 98·5% against canine parvovirus type 2. The small number of dogs that were seronegative for one or more of the antigens (n = 31) may have had waning of previous serum antibody or may have been rare genetic non-responders to that specific antigen. UK veterinarians can be reassured that triennial revaccination of adult dogs with core vaccines provides long-lived protective immunity. In-practice serological test kits are a valuable tool for informing decision-making about canine core revaccination. © 2017 British Small Animal Veterinary Association.

A fast and reproducible method for albumin isolation and depletion from serum and cerebrospinal fluid.

PubMed

Holewinski, Ronald J; Jin, Zhicheng; Powell, Matthew J; Maust, Matthew D; Van Eyk, Jennifer E

2013-03-01

Analysis of serum and plasma proteomes is a common approach for biomarker discovery, and the removal of high-abundant proteins, such as albumin and immunoglobins, is usually the first step in the analysis. However, albumin binds peptides and proteins, which raises concerns as to how the removal of albumin could impact the outcome of the biomarker study while ignoring the possibility that this could be a biomarker subproteome itself. The first goal of this study was to test a new commercially available affinity capture reagent from Protea Biosciences and to compare the efficiency and reproducibility to four other commercially available albumin depletion methods. The second goal of this study was to determine if there is a highly efficient albumin depletion/isolation system that minimizes sample handling and would be suitable for large numbers of samples. Two of the methods tested (Sigma and ProteaPrep) showed an albumin depletion efficiency of 97% or greater for both serum and cerebrospinal fluid (CSF). Isolated serum and CSF albuminomes from ProteaPrep spin columns were analyzed directly by LC-MS/MS, identifying 128 serum (45 not previously reported) and 94 CSF albuminome proteins (17 unique to the CSF albuminome). Serum albuminome was also isolated using Vivapure anti-HSA columns for comparison, identifying 105 proteins, 81 of which overlapped with the ProteaPrep method. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effects of mayonnaise on postprandial serum lutein/zeaxanthin and beta-carotene concentrations in humans.

PubMed

Takeda, Sayaka; Masuda, Yasunobu; Usuda, Mika; Marushima, Ranko; Ueji, Toshiyuki; Hasegawa, Mineo; Maruyama, Chizuko

2009-12-01

To clarify the effects of different physical forms of oil on postprandial serum lutein/zeaxanthin and beta-carotene concentrations, we performed a vegetable meal loading test. Eighteen healthy subjects participated in the test, which consisted of broccoli as a control (CON) meal, broccoli with oil (OIL), and broccoli with mayonnaise (MS), consumed in random order. After collection of fasting blood samples, subjects consumed one of the three test meals. Fasting and postprandial changes in serum carotenoids were assessed 2, 4, and 6 h after ingestion of each test meal. Serum lutein/zeaxanthin and beta-carotene concentrations were measured. Although no significant change was noted after the CON meal, the serum lutein/zeaxanthin concentration was higher at 4 h after consumption of the OIL meal, and at 2, 4 and 6 h after consumption of the MS meal, as compared with the fasting state. Serum beta-carotene concentrations did not change after ingestion of either the CON or the OIL meal but were elevated 2, 4, and 6 h after MS ingestion as compared with the fasting state. The incremental areas under the curves (IAUCs) of serum lutein/zeaxanthin and beta-carotene concentrations were higher after the MS meal than after the CON meal. IAUCs after the OIL meal exhibited no statistically significant differences from the CON and MS meals. We suggest that mayonnaise contributes to increase serum lutein/zeaxanthin and beta-carotene concentrations when consumed with vegetables rich in these carotenoids.

Antibody against infectious salmon anaemia virus among feral Atlantic salmon (Salmo salar)

USGS Publications Warehouse

Cipriano, R.C.

2009-01-01

Archived sera from Atlantic salmon (Salmo salar) that returned to the Penobscot River (Maine), Merrimack River (Massachusetts), and Connecticut River (in Massachusetts) from 1995 to 2002 were analysed for antibodies against infectious salmon anaemia virus (ISAV) using an enzyme-linked immunosorbent assay (ELISA). Up to 60 samples were archived per river system per year. In a given year, the number of fish sampled by ELISA for ISAV antibodies in the Penobscot River ranged from 2.9 to 11.2, and the range of salmon sampled in the Merrimack River and the Connecticut River was 31.3-100 and 20.0-67.5, respectively. Archived sera were not available for the 1995 and 2002 year classes from the Connecticut River. In all, 1141 samples were processed; 14 serum samples tested positive for antibodies to ISAV. In the Penobscot River, serum from one fish tested positive in each of the 1995 and 1999 year-class returns, and sera from two fish tested positive in the 1998 returns. In the Merrimack River, sera from four fish tested positive in each of the 1996 and 1997 returns, and sera from two fish were positive in the 2002 return. None of the archived sera from Atlantic salmon that returned to the Connecticut River tested positive. ?? 2009 United States Government, Department of the Interior.

Comparison of 2 electronic cowside tests to detect subclinical ketosis in dairy cows and the influence of the temperature and type of blood sample on the test results.

PubMed

Iwersen, M; Klein-Jöbstl, D; Pichler, M; Roland, L; Fidlschuster, B; Schwendenwein, I; Drillich, M

2013-01-01

The objective of this study was to determine the suitability of 2 electronic hand-held devices [FreeStyle Precision (FSP), Abbott GmbH & Co. KG, Wiesbaden, Germany and GlucoMen LX Plus (GLX), A. Menarini GmbH, Vienna, Austria] for measuring β-hydroxybutyrate (BHBA) in dairy cows. Three experiments were conducted to evaluate (1) the diagnostic performance of the devices, (2) the effect of the type of blood sample, and (3) the influence of the ambient temperature on the determined results. A total of 415 blood samples from lactating Holstein and Simmental cows were collected and analyzed with both devices (whole blood) and in a laboratory (serum). Correlation coefficients between whole-blood and serum BHBA concentrations were highly significant, with 94% for the FSP and 80% for the GLX device. Based on thresholds for subclinical ketosis of 1.2 and 1.4 mmol of BHBA/L, results obtained with the hand-held devices were evaluated by receiver operating characteristics analyses. This resulted in adjusted thresholds of 1.2 and 1.4 mmol/L for the FSP and 1.1 and 1.3 mmol/L for the GLX device. Applying these thresholds, sensitivities were 98 and 100% for the FSP and 80 and 86% for the GLX device, respectively. Corresponding specificities were 90 and 97% for the FSP and 87 and 96% for the GLX device, respectively. Additionally, concentrations of BHBA were tested with both devices in whole blood, EDTA-added whole blood, and in their resulting serum and plasma, collected from 65 animals. Determined BHBA concentrations were similar within each device for whole and EDTA-added blood, and in serum and plasma, but differed between whole blood and serum and between EDTA-added blood and plasma. Blood samples with low (0.4 mmol/L), medium (1.1 mmol/L), and high (1.6 mmol/L) BHBA concentrations were stored between +5 to +32°C and analyzed repeatedly at temperature levels differing by 4°C. Additionally, devices and test strips were stored at equal conditions and used for measurement procedures. Storage temperature of the devices and test strips did not influence the differences between the results of the laboratory and the devices, whereas the temperature of the blood samples caused significant differences. Although the level of agreement between the laboratory and the GLX device was lower than for the laboratory and the FSP device, both devices are useful tools for monitoring subclinical ketosis in dairy cows. Due to their effects on the determined results, the type and temperature of the tested sample should be considered. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Can serums be replaced by Mueller-Hinton agar in germ tube test?

PubMed

Atalay, M A; Koc, A N; Parkan, O M; Aydemir, G; Elmali, F; Sav, H

2017-01-01

The germ tube test (GTT) is inexpensive, easy, and well-defined test that differentiates Candida albicans (excluding Candida dubliniensis and Candida africana) from other species. The aim of this study was to evaluate various serums (i.e., human, rabbit, horse, and fetal bovine serum) used in the GTT and Mueller-Hinton agar (MHA). Fifty species isolated from various clinical samples that were defined as C. albicans by both conventional and DNA sequence analysis methods were included in the study. One to two colonies of C. albicans were mixed into 0.5-1 ml of fetal bovine serum, horse serum, rabbit serum, and human serum. Serums and MHA were incubated at 37°C for GTT. They were removed from the incubator and evaluated after 30 min, 1 h, 2 h, and 3 h of incubation. The GTT was accepted to be positive only if germ tube was 1/2 the width and 3 times the length of the parent yeast cell and with no constriction at the point of origin. When the use of serums and MHA for GTT was statistically evaluated, according to the positive scoring, the best results were obtained with MHA and with rabbit, horse, and fetal bovine serum, respectively. The best definition over time statistically was the third hour. It is suggested that inexpensive MHA is a fast, appropriate, and reliable medium for the probable diagnosis of GTT and C. albicans; however, additional studies are still needed to define other Candida species.

Performance of a commercial serum ELISA for the detection of antibodies to Neospora caninum in whole and skim milk samples.

PubMed

Byrem, T M; Bartlett, P C; Donohue, H; Voisinet, B D; Houseman, J T

2012-11-23

Control of Neospora caninum infection in cattle depends on specific, ante-mortem detection of infected animals and limiting their use as breeding stock or by culling. The objective of the present study was to determine appropriate cut-off values and diagnostic performance of a milk ELISA test using whole and skim milk in a commercial serum ELISA test (IDEXX Neospora Ab). Serum and milk samples were obtained from a total of 475 lactating cows from two herds with and two herds without a previous history of N. caninum-associated abortion. Overall seroprevalence determined by the ELISA was 18.3%. Compared to serum ELISA values, correlation and overall performance assessed by receiver operating characteristic analysis was higher when either whole or skim milk samples were diluted 1:2 compared to undiluted or 1:5 diluted samples. Diagnostic performance for analysis of whole and skim milk was compared at cut-off values that achieved a desired operating characteristic of at least 95% specificity. For whole milk diluted 1:2 and a cut-off of 0.14 (S/P ratio), sensitivity and kappa values were 74.7% (95% CI 64.3-83.4) and 0.70 (95% CI 0.61-0.78), respectively. For skim milk diluted 1:2 and a cut-off of 0.30, sensitivity and kappa values were 77.0% (95% CI 66.8-85.4) and 0.72 (95% CI 0.64-0.80), respectively. Using the selected cut-offs, the IDEXX Neospora Ab Test is equally suited for the analysis of whole and skim milk as a screening tool in neosporosis control programs. Copyright © 2012 Elsevier B.V. All rights reserved.

Prevalence of antibodies to Trypanosoma cruzi, Leishmania infantum, Encephalitozoon cuniculi, Sarcocystis neurona, and Neospora caninum in Capybara, Hydrochoerus hydrochaeris, from São Paulo State, Brazil.

PubMed

Valadas, Samantha; Gennari, Solange Maria; Yai, Lucia Eiko Oishi; Rosypal, Alexa C; Lindsay, David S

2010-06-01

Little is known about the importance of capybara, Hydrochoerus hydrochaeris, as reservoirs for parasites of zoonotic or veterinary importance. Sera from 63 capybaras, from 6 counties in the state of São Paulo, Brazil, were examined for antibodies to Trypanosoma cruzi, Leishmania infantum, Encephalitozoon cuniculi, Sarcocystis neurona, and Neospora caninum using an indirect immunofluorescent antibody test. Five (8%) of the 63 capybaras had antibodies to T. cruzi epimastigotes. None of the samples from capybara reacted positively with L. infantum promastigotes or with spores of E. cuniculi . Two (3%) of the serum samples were positive for antibodies to S. neurona merozoites, and 2 (3%) of the serum samples were positive for antibodies to N. caninum tachyzoites. A serum sample from 1 capybara was positive for antibodies to both T. cruzi and N. caninum. None of the remaining 62 samples reacted with more than 1 parasite.

TESTS TO ASSESS SENSITIZATION TO ASPERGILLUS FUMIGATUS IN CYSTIC FIBROSIS

PubMed Central

Aguiar, Simone Santana; Damaceno, Neiva; Forte, Wilma Carvalho Neves

2017-01-01

ABSTRACT Objective: To evaluate the results of the tests used to identify the IgE mediated sensitization to Aspergillus fumigatus in patients with cystic fibrosis. Methods: This is a cross-sectional descriptive study with a convenience sample of 86 patients diagnosed with cystic fibrosis in the Reference Service in Cystic Fibrosis at a tertiary teaching hospital. The following tests were performed to assess the sensitization to A. fumigatus in patients with cystic fibrosis: Total serum IgE, eosinophil count, fungus detection through oropharyngeal swab or sputum culture, serum-specific IgE, and immediate-type hypersensitivity (IgE) skin tests. We compared the results of the different tests performed. Results: In 33 (38.4%) patients with cystic fibrosis, with ages ranging from 1 to 33 years (median of 8 years), the IgE-mediated A. fumigatus sensitization test results were: in 16 patients, there was an increase in serum-specific IgE (>0.35 kU/L); in 23, skin tests were positive; and six had sensitization in both tests. We observed two patients with eosinophilia (>1,000 eosinophils/mm3) and seven with increasing total serum IgE (>1,000 IU/mL), all of whom obtained negative results in skin tests and had no IgE increase specific to A. fumigatus. A. fumigatus was not detected in oropharyngeal swabs and/or sputum culture of any patients. Conclusions: We conclude that, among the tests used to assess sensitization to A. fumigatus in cystic fibrosis patients, both serum-specific IgE and immediate-type hypersensitivity (IgE) skin tests are required. Serum eosinophilia and respiratory secretion culture were not essential in this study. PMID:28977288

Rapid antigen tests for dengue virus serotypes and Zika virus in patient serum.

PubMed

Bosch, Irene; de Puig, Helena; Hiley, Megan; Carré-Camps, Marc; Perdomo-Celis, Federico; Narváez, Carlos F; Salgado, Doris M; Senthoor, Dewahar; O'Grady, Madeline; Phillips, Elizabeth; Durbin, Ann; Fandos, Diana; Miyazaki, Hikaru; Yen, Chun-Wan; Gélvez-Ramírez, Margarita; Warke, Rajas V; Ribeiro, Lucas S; Teixeira, Mauro M; Almeida, Roque P; Muñóz-Medina, José E; Ludert, Juan E; Nogueira, Mauricio L; Colombo, Tatiana E; Terzian, Ana C B; Bozza, Patricia T; Calheiros, Andrea S; Vieira, Yasmine R; Barbosa-Lima, Giselle; Vizzoni, Alexandre; Cerbino-Neto, José; Bozza, Fernando A; Souza, Thiago M L; Trugilho, Monique R O; de Filippis, Ana M B; de Sequeira, Patricia C; Marques, Ernesto T A; Magalhaes, Tereza; Díaz, Francisco J; Restrepo, Berta N; Marín, Katerine; Mattar, Salim; Olson, Daniel; Asturias, Edwin J; Lucera, Mark; Singla, Mohit; Medigeshi, Guruprasad R; de Bosch, Norma; Tam, Justina; Gómez-Márquez, Jose; Clavet, Charles; Villar, Luis; Hamad-Schifferli, Kimberly; Gehrke, Lee

2017-09-27

The recent Zika virus (ZIKV) outbreak demonstrates that cost-effective clinical diagnostics are urgently needed to detect and distinguish viral infections to improve patient care. Unlike dengue virus (DENV), ZIKV infections during pregnancy correlate with severe birth defects, including microcephaly and neurological disorders. Because ZIKV and DENV are related flaviviruses, their homologous proteins and nucleic acids can cause cross-reactions and false-positive results in molecular, antigenic, and serologic diagnostics. We report the characterization of monoclonal antibody pairs that have been translated into rapid immunochromatography tests to specifically detect the viral nonstructural 1 (NS1) protein antigen and distinguish the four DENV serotypes (DENV1-4) and ZIKV without cross-reaction. To complement visual test analysis and remove user subjectivity in reading test results, we used image processing and data analysis for data capture and test result quantification. Using a 30-μl serum sample, the sensitivity and specificity values of the DENV1-4 tests and the pan-DENV test, which detects all four dengue serotypes, ranged from 0.76 to 1.00. Sensitivity/specificity for the ZIKV rapid test was 0.81/0.86, respectively, using a 150-μl serum input. Serum ZIKV NS1 protein concentrations were about 10-fold lower than corresponding DENV NS1 concentrations in infected patients; moreover, ZIKV NS1 protein was not detected in polymerase chain reaction-positive patient urine samples. Our rapid immunochromatography approach and reagents have immediate application in differential clinical diagnosis of acute ZIKV and DENV cases, and the platform can be applied toward developing rapid antigen diagnostics for emerging viruses. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Prevalence of Candidatus Mycoplasma haemolamae, bovine viral diarrhoea virus, and gastrointestinal parasitism in a sample of adult New Zealand alpaca (Vicugna pacos).

PubMed

Dittmer, K E; Hinkson, J A; Dwyer, C; Adlington, B; van Andel, M

2018-01-01

To determine the prevalence of infection with Candidatus Mycoplasma haemolamae (Mhl), antibodies to bovine viral diarrhoea virus (BVDV), and BVDV antigen, and the prevalence of animals with elevated faecal nematode egg counts (FEC) in a sample of adult New Zealand alpaca (Vicugna pacos). Blood samples were obtained from 175 alpaca, collected from 15 farms around New Zealand, and from 31 samples sent to a diagnostic laboratory for routine haematology. Blood smears (n=170) were examined microscopically for the presence of haemoplasma, and DNA was extracted from whole blood (n=206) for real-time PCR testing for Mhl. Packed cell volume (PCV) was determined for 193 samples. Serum samples (n=195) were tested for BVDV antibody using ELISA, and for BVDV antigen using a real-time PCR assay. Faecal samples were collected from 143 animals; FEC were measured, and samples pooled for larval culture. No haemoplasma organisms were present on blood smear examination. Of the 206 blood samples, two (from the same farm) were positive for Mhl by real-time PCR testing, giving a prevalence of infection with Mhl of 0.97%. Of the 195 serum samples tested, four (2.1%) were positive for antibodies to BVDV; animals with BVDV antibodies were from 3/15 (20%) farms, none of which farmed cattle. None of the serum samples were positive by PCR for BVDV antigen. The median FEC was 50 epg (min 0, max 4,700), with 55/143 (38.5%) samples having 0 epg, and 33/143 (23.1%) having ≥250 epg. Haemonchus spp. were the most common nematodes present in faecal larval cultures from the North Island. Log 10 FEC was negatively associated with PCV (p=0.02), and was higher in males than females (p<0.001), and in animals that were positive compared with negative for Mhl (p=0.022). The number of alpaca infected with Mhl was low, as was the seroprevalence of BVDV. Gastrointestinal parasitism was, however, a common finding in this sample of New Zealand alpaca.

Detection of porcine reproductive and respiratory syndrome virus (PRRSV) and influenza A virus (IAV) in oral fluid of pigs.

PubMed

Biernacka, Kinga; Karbowiak, Paweł; Wróbel, Paweł; Charęza, Tomasz; Czopowicz, Michał; Balka, Gyula; Goodell, Christa; Rauh, Rolf; Stadejek, Tomasz

2016-12-01

Recently oral fluid has become a novel sample type for pathogen nucleic acid and antibody detection, as it is easy to obtain with non-invasive procedures. The objective of the study was to analyze porcine reproductive and respiratory syndrome virus (PRRSV) and influenza A virus (IAV) circulation in growing pigs from three Polish production farms, using Real Time PCR and ELISA testing of oral fluid and serum. Oral fluids were collected every 2weeks, in the same 3-4 pens of pigs aged between 5 and 17weeks. Additionally, blood samples were collected every 4weeks from 4 pigs corresponding to the same pens as oral fluid and tested for the presence of PRRSV nucleic acid (pooled by 4) and antibodies. In farm A no PRRSV circulation was detected and only maternal antibodies were present. In farm B and farm C antibodies to PRRSV in serum and oral fluid were detected in most samples. In farm B PRRSV Type 1 was detected in 80.9% of oral fluid samples and in 58.3% of serum pools, and in farm C in 92.8% of oral fluid samples and 75% serum pools. Striking differences were observed between different pens in PRRSV detection patterns. In farms B and C ORF5 sequence analysis showed the presence of wild type strains which were about 84-85% identical to the modified live vaccine used. In all three farms two waves of IAV shedding with oral fluid were detected, in weaners and fatteners. Copyright © 2016 Elsevier Ltd. All rights reserved.

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

PubMed Central

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

2012-01-01

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

Serum allergen-specific immunoglobulin E in atopic and healthy cats: comparison of a rapid screening immunoassay and complete-panel analysis.

PubMed

Diesel, Alison; DeBoer, Douglas J

2011-02-01

Feline and canine atopic dermatitis are thought to have a similar immunopathogenesis. As with dogs, detection of allergen-specific IgE in cat serum merely supports a diagnosis of feline atopy based on compatible history, clinical signs and elimination of other pruritic dermatoses. In this study, a rapid screening immunoassay (Allercept(®) E-Screen 2nd Generation; Heska AG, Fribourg, Switzerland; ES2G) was compared with a complete-panel serum allergen-specific IgE assay (Allercept(®); Heska AG; CP) in healthy cats with no history of skin disease and in atopic cats. The latter had no diagnosis of external parasitism, infection, food hypersensitivity or other skin disease explaining their pruritus, and expressed cutaneous reaction patterns typically associated with feline allergic skin disease (head, neck or pinnal pruritus, miliary dermatitis, self-induced alopecia, eosinophilic granuloma complex). The proportion of cats positive on either the ES2G or the CP assays was not significantly different between the atopic and healthy cat groups. There was, however, strong agreement between the results of the ES2G and CP assay; overall, the two tests were in agreement for 43 of 49 (88%) serum samples. There was also strong agreement when individual allergen groups were evaluated (agreement noted: indoor, 41 of 49 samples; grasses/weeds, 37 of 49 samples; and trees, 41 of 49 samples). These results indicate that although neither test is diagnostic for feline atopic dermatitis, the screening assay is beneficial for predicting the results of a complete-panel serum allergen-specific IgE assay in cats. © 2010 The Authors. Journal compilation © 2010 ESVD and ACVD.

Determination of total creatine kinase activity in blood serum using an amperometric biosensor based on glucose oxidase and hexokinase.

PubMed

Kucherenko, I S; Soldatkin, O O; Lagarde, F; Jaffrezic-Renault, N; Dzyadevych, S V; Soldatkin, A P

2015-11-01

Creatine kinase (CK: adenosine-5-triphosphate-creatine phosphotransferase) is an important enzyme of muscle cells; the presence of a large amount of the enzyme in blood serum is a biomarker of muscular injuries, such as acute myocardial infarction. This work describes a bi-enzyme (glucose oxidase and hexokinase based) biosensor for rapid and convenient determination of CK activity by measuring the rate of ATP production by this enzyme. Simultaneously the biosensor determines glucose concentration in the sample. Platinum disk electrodes were used as amperometric transducers. Glucose oxidase and hexokinase were co-immobilized via cross-linking with BSA by glutaraldehyde and served as a biorecognition element of the biosensor. The biosensor work at different concentrations of CK substrates (ADP and creatine phosphate) was investigated; optimal concentration of ADP was 1mM, and creatine phosphate - 10 mM. The reproducibility of the biosensor responses to glucose, ATP and CK during a day was tested (relative standard deviation of 15 responses to glucose was 2%, to ATP - 6%, to CK - 7-18% depending on concentration of the CK). Total time of CK analysis was 10 min. The measurements of creatine kinase in blood serum samples were carried out (at 20-fold sample dilution). Twentyfold dilution of serum samples was chosen as optimal for CK determination. The biosensor could distinguish healthy and ill people and evaluate the level of CK increase. Thus, the biosensor can be used as a test-system for CK analysis in blood serum or serve as a component of multibiosensors for determination of important blood substances. Determination of activity of other kinases by the developed biosensor is also possible for research purposes. Copyright © 2015 Elsevier B.V. All rights reserved.

Coexistence of antibodies to tick-borne pathogens of babesiosis, ehrlichiosis, and Lyme borreliosis in human sera.

PubMed

Magnarelli, L A; Dumler, J S; Anderson, J F; Johnson, R C; Fikrig, E

1995-11-01

Serum specimens from persons with or without Lyme borreliosis were analyzed by indirect fluorescent antibody staining methods for total immunoglobulins to Babesia microti, Ehrlichia chaffeensis (Arkansas strain), and Ehrlichia equi (MRK strain). There was serologic evidence of human exposure to multiple tick-borne agents in 15 (6.6%) of 227 serum samples obtained in Connecticut and Minnesota. Of these, 10 serum samples were from Connecticut patients who had erythema migrans and antibodies to Borrelia burgdorferi (range, 1:160 to 1:40, 960). A maximal antibody titer of 1:640 was noted for a B. microti infection, whereas titration end points of 1:640 and 1:1,280 were recorded for E. chaffeensis and E. equi seropositives, respectively. In specificity tests, there was no cross-reactivity among the antisera and antigens tested for the four tick-borne pathogens. On the basis of serologic testing, a small group of persons who had Lyme borreliosis had been exposed to one or more other tick-borne agents, but there was no clinical diagnosis of babesiosis or ehrlichiosis. Therefore, if the clinical picture is unclear or multiple tick-associated illnesses are suspected, more extensive laboratory testing is suggested.

Effect of pronase on high-incidence blood group antigens and the prevalence of antibodies to pronase-treated erythrocytes.

PubMed

Reid, M E; Greeen, C A; Hoffer, J; Øyen, R

1996-01-01

Pronase is a useful and relatively nonspecific protease that cleaves many red blood cell (RBC) membrane proteins that carry blood group antigens. Unexpected findings in tests using pronase-treated RBCs during the investigation of a patient's blood sample led us to test which high-incidence blood group antigens were sensitive and which were resistant to pronase treatment, and to determine the prevalence of antipronase in the serum of blood donors. Our results show that antigens in the Cromer and Lutheran blood group systems and the JMH antigen were sensitive to pronase treatment of RBCs. Antigens in the Dombrock blood group system and Sc1 were either sensitive to or markedly weakened by pronase treatment of RBCs. The following high-incidence antigens were resistant to treatment of RBCs with pronase: AnWj, Ata, Coa, Co3, Dib, EnaFR, Era, Fy3, Jk3, Jra, k, Kpb, Jsb, K14, Lan, Oka, Rh17, U, Vel, and Wrb. Over half of the serum samples from normal blood donors contained antibodies to pronase-treated RBCs. When testing human serum against pronase-treated RBCs, it is essential either to use an autocontrol or to perform the testing with an eluate.

Serological, Bacteriological, and Molecular Diagnosis of Brucellosis in Domestic Animals in Croatia

PubMed Central

Špičić, Silvio; Zdelar-Tuk, Maja; Račić, Ivana; Duvnjak, Sanja; Cvetnić, Željko

2010-01-01

Aim To present the surveillance data on Brucella melitensis, B. suis, and B. ovis infection in cattle, sheep, goats, and swine in Croatia obtained in 2008 by serological, bacteriological, and molecular methods for diagnostics of brucellosis in domestic animals. Methods We serologically tested 42 785 cattle serums, 22 686 sheep and goat serums, and 28 520 swine serums using the Rose Bengal test, complement fixation test, and various immunosorbent assays. We also tested 10 173 ram blood samples for B. ovis infection using the complement fixation test. Bacteriological examination was conducted on 214 samples collected from 34 serologically positive animals. Different molecular methods were employed in the identification and typing of 20 isolates from the samples. Results B. melitensis biovar (bv.) 3 was confirmed with different identification methods in 2 flocks in 2 Croatian counties and B. suis bv. 2 in 3 herds in 3 counties. B. melitensis in cows was confirmed for the first time in Croatia. Infection with B. ovis was serologically confirmed in 202 rams in 12 counties. Conclusions In 2008, the size of the brucellosis-affected area in Croatia and the efficiency of detection and prevention of brucellosis in sheep, goats, and swine were satisfactory. Infection with B. melitensis in cattle was confirmed for the first time and possible links for infection in humans were detected. More efficient measures for suppression and control of ovine epididymitis are required and a new strategy may be necessary for complete eradication of this disease. PMID:20718085

Single-dilution enzyme-linked immunosorbent assay for quantification of antigen-specific salmonid antibody

USGS Publications Warehouse

Alcorn, S.W.; Pascho, R.J.

2000-01-01

An enzyme-linked immunosorbent assay (ELISA) was developed on the basis of testing a single dilution of serum to quantify the level of antibody to the p57 protein of Renibaclerium salmoninarum in sockeye salmon (Oncorhynchus nerka). The levels of antibody were interpolated from a standard curve constructed by relating the optical densities (OD) produced by several dilutions of a high-titer rainbow trout (O. mykiss) antiserum to the p57 protein. The ELISA OD values produced by as many as 36 test sera on each microplate were compared with the standard curve to calculate the antigen-specific antibody activity. Repeated measurements of 36 samples on 3 microplates on each of 6 assay dates indicated that the mean intraassay coefficient of variation (CV) was 6.68% (range, 0-23%) and the mean interassay CV was 8.29% (range, 4-16%). The antibody levels determined for the serum sample from 24 sockeye salmon vaccinated with a recombinant p57 protein generally were correlated with the levels determined by endpoint titration (r2 = 0.936) and with results from another ELISA that was based on extrapolation of antibody levels from a standard curve (r2 = 0.956). The single-dilution antibody ELISA described here increases the number of samples that can be tested on each microplate compared with immunoassays based on analysis of several dilutions of each test serum. It includes controls for interassay standardization and can be used to test fish weighing <3 g.

Detection of influenza A virus nucleoprotein antibodies in oral fluid specimens from pigs infected under experimental conditions using a blocking ELISA.

PubMed

Panyasing, Y; Goodell, C K; Wang, C; Kittawornrat, A; Prickett, J R; Schwartz, K J; Ballagi, A; Lizano, S; Zimmerman, J J

2014-04-01

In commercial swine populations, influenza is an important component of the porcine respiratory disease complex (PRDC) and a pathogen with major economic impact. Previously, a commercial blocking ELISA (FlockChek(™) Avian Influenza Virus MultiS-Screen(®) Antibody Test Kit, IDEXX Laboratories, Inc., Westbrook, ME, USA) designed to detect influenza A nucleoprotein (NP) antibodies in avian serum was shown to accurately detect NP antibodies in swine serum. The purpose of this study was to determine whether this assay could detect NP antibodies in swine oral fluid samples. Initially, the procedure for performing the NP-blocking ELISA on oral fluid was modified from the serum testing protocol by changing sample dilution, sample volume, incubation time and incubation temperature. The detection of NP antibody was then evaluated using pen-based oral fluid samples (n = 182) from pigs inoculated with either influenza A virus subtype H1N1 or H3N2 under experimental conditions and followed for 42 days post inoculation (DPI). NP antibodies in oral fluid were detected from DPI 7 to 42 in all inoculated groups, that is, the mean sample-to-negative (S/N) ratio of influenza-inoculated pigs was significantly different (P < 0.0001) from uninoculated controls (unvaccinated or vaccinated-uninoculated groups) through this period. Oral fluid versus serum S/N ratios from the same pen showed a correlation of 0.796 (Pearson's correlation coefficient, P < 0.0001). The results showed that oral fluid samples from influenza virus-infected pigs contained detectable levels of NP antibodies for ≥42 DPI. Future research will be required to determine whether this approach could be used to monitor the circulation of influenza virus in commercial pig populations. © 2012 Blackwell Verlag GmbH.

Analyte stability during the total testing process: studies of vitamins A, D and E by LC-MS/MS.

PubMed

Albahrani, Ali A; Rotarou, Victor; Roche, Peter J; Greaves, Ronda F

2016-10-01

There are limited evidence based studies demonstrating the stability of fat-soluble vitamins (FSV) measured in blood. This study aimed to examine the effects of light, temperature and time on vitamins A, D and E throughout the total testing process. Four experiments were conducted. Three investigated the sample matrix, of whole blood, serum and the extracted sample, against the variables of temperature and light; and the fourth experiment investigated the sample during the extraction process against the variable of light. All samples were analysed via our simultaneous FSV method using liquid chromatography-tandem mass spectrometry technology. The allowable clinical percentage change was calculated based on biological variation and desirable method imprecision for each analyte. The total change limit was ±7.3% for 25-OH-vitamin D3, ±11.8% for retinol and ±10.8% for α-tocopherol. Vitamins D and E were stable in the investigated conditions (concentration changes <4%) in the pre-analytical and analytical stages. Vitamin A showed photosensitivity in times >48 h with concentration changes of -6.8% (blood) and -6.5% (serum), both are within the allowable clinical percentage change. By contrast, the extracted retinol sample demonstrated a concentration change of -18.4% after 48 h of light exposure. However, vitamin A in the serum and extracted solution was stable for one month when stored at -20°C. Blood samples for vitamins D and E analyses can be processed in normal laboratory conditions of lighting and temperature. The required conditions for vitamin A analysis are similar when performed within 48 h. For longer-term storage, serum and vitamin A extracts should be stored at -20°C.

Identification of potential serum peptide biomarkers of biliary tract cancer using MALDI MS profiling

PubMed Central

2014-01-01

Background The aim of this discovery study was the identification of peptide serum biomarkers for detecting biliary tract cancer (BTC) using samples from healthy volunteers and benign cases of biliary disease as control groups. This work was based on the hypothesis that cancer-specific exopeptidases exist and that their activities in serum can generate cancer-predictive peptide fragments from circulating proteins during coagulation. Methods This case control study used a semi-automated platform incorporating polypeptide extraction linked to matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) to profile 92 patient serum samples. Predictive models were generated to test a validation serum set from BTC cases and healthy volunteers. Results Several peptide peaks were found that could significantly differentiate BTC patients from healthy controls and benign biliary disease. A predictive model resulted in a sensitivity of 100% and a specificity of 93.8% in detecting BTC in the validation set, whilst another model gave a sensitivity of 79.5% and a specificity of 83.9% in discriminating BTC from benign biliary disease samples in the training set. Discriminatory peaks were identified by tandem MS as fragments of abundant clotting proteins. Conclusions Serum MALDI MS peptide signatures can accurately discriminate patients with BTC from healthy volunteers. PMID:24495412

Prevalence of Coxiella burnetii Infection in Humans Occupationally Exposed to Animals in Poland

PubMed Central

Galińska, Elżbieta Monika; Niemczuk, Krzysztof; Knap, Józef Piotr

2015-01-01

Abstract Coxiella burnetii is the etiological agent of Q fever, and outbreaks of Q fever have been reported in different parts of Europe both in animals and humans. Human infections are mostly associated with infections in ruminants, e.g., sheep, goats, and cows. Various professional groups are occupationally exposed to infection with C. burnetii. The aim of this study was investigate the prevalence of C. burnetii in farm workers. Serum samples were collected from 151 persons from six different regions of Poland. The serum samples were tested using three serological methods—complement fixation test (CFT), enzyme-linked immunosorbent assay (ELISA), and indirect fluorescent antibody (IFA). A total of 71 samples of blood were also tested by real-time PCR. The results showed that antibodies against C. burnetii were present in the tested sera. Average percentages of seropositive samples in IFA, ELISA, and CFT were 31.12%, 39.07%, and 15.23%, respectively. Positive results were noted in each testing center. Of the three test types, IFA results were considered the most sensitive. Real-time PCR confirmed the presence of DNA specific for C. burnetii in 10 patients. The farming workforce constitutes an occupational risk group with an increased risk for C. burnetii infection, presumably because of their contact with infected livestock. PMID:25897813

Feasibility of Using Convalescent Plasma Immunotherapy for MERS-CoV Infection, Saudi Arabia.

PubMed

Arabi, Yaseen M; Hajeer, Ali H; Luke, Thomas; Raviprakash, Kanakatte; Balkhy, Hanan; Johani, Sameera; Al-Dawood, Abdulaziz; Al-Qahtani, Saad; Al-Omari, Awad; Al-Hameed, Fahad; Hayden, Frederick G; Fowler, Robert; Bouchama, Abderrezak; Shindo, Nahoko; Al-Khairy, Khalid; Carson, Gail; Taha, Yusri; Sadat, Musharaf; Alahmadi, Mashail

2016-09-01

We explored the feasibility of collecting convalescent plasma for passive immunotherapy of Middle East respiratory syndrome coronavirus (MERS-CoV) infection by using ELISA to screen serum samples from 443 potential plasma donors: 196 patients with suspected or laboratory-confirmed MERS-CoV infection, 230 healthcare workers, and 17 household contacts exposed to MERS-CoV. ELISA-reactive samples were further tested by indirect fluorescent antibody and microneutralization assays. Of the 443 tested samples, 12 (2.7%) had a reactive ELISA result, and 9 of the 12 had reactive indirect fluorescent antibody and microneutralization assay titers. Undertaking clinical trials of convalescent plasma for passive immunotherapy of MERS-CoV infection may be feasible, but such trials would be challenging because of the small pool of potential donors with sufficiently high antibody titers. Alternative strategies to identify convalescent plasma donors with adequate antibody titers should be explored, including the sampling of serum from patients with more severe disease and sampling at earlier points during illness.

Serodiagnosis of whooping cough in Belgium: results of the National Reference Centre for Bordetella pertussis anno 2013.

PubMed

Duterme, Sophie; Vanhoof, Raymond; Vanderpas, Jean; Pierard, Denis; Huygen, Kris

2016-04-01

Report on the pitfalls of serodiagnosis of pertussis in Belgium for 2013 by the NRC Bordetella. Determine cases of acute infection using an anti-pertussis toxin (PT) IgG antibody ELISA. A total of 2471 serum samples were received. Clinical information on the duration of cough (at moment of blood sampling) is essential for a reliable interpretation of the results. In order to avoid false negative results, 213 samples for which this information was lacking were not tested. For a total of 2179 patients tested, 520 (23.9%) had antibody levels indicative of an acute infection, 261 (12%) samples were diagnosed as positive (indicative of a pertussis infection or vaccination during the last year), 143 (6.7%) samples were classified as doubtful and 752 (34,5%) (35.5%) were diagnosed as negative. The serodiagnosis of pertussis has limited value for the early diagnosis of the disease and PCR analysis on nasopharyngeal swabs is the method of choice during the first 2 weeks and always for young children <1 year old. For sera collected during the first 2 weeks with anti-PT levels below the threshold for acute infection, a second sample collected 2-3 weeks later is needed a definitive diagnosis. For 503 (23.0%) early samples, a second serum sample was requested but not provided. For 85 patients, for whom a second sample was received, 12.9% were eventually diagnosed as having an acute infection. In order to generate reliable serodiagnostic results for pertussis, serum samples should preferentially be collected 3 weeks after onset of symptoms.

Large-scale analysis of tumor necrosis factor α levels in systemic lupus erythematosus.

PubMed

Weckerle, Corinna E; Mangale, Dorothy; Franek, Beverly S; Kelly, Jennifer A; Kumabe, Marissa; James, Judith A; Moser, Kathy L; Harley, John B; Niewold, Timothy B

2012-09-01

Systemic lupus erythematosus (SLE) disease manifestations are highly variable among patients, and the prevalence of individual clinical features differs significantly by ancestry. Serum tumor necrosis factor α (TNFα) levels are elevated in some SLE patients and may play a role in disease pathogenesis. The aim of this study was to look for associations between serum TNFα levels, clinical manifestations of SLE, autoantibodies, and serum interferon-α (IFNα) levels in a large multiancestral SLE cohort. We studied serum TNFα levels in 653 SLE patients (214 African Americans, 298 European Americans, and 141 Hispanic Americans). TNFα was measured using an enzyme-linked immunosorbent assay, and IFNα was measured with a functional reporter cell assay. Stratified and multivariate analyses were used to detect associations in each ancestral background separately, with meta-analysis when appropriate. Serum TNFα levels were significantly higher in SLE patients than in non-autoimmune disease controls (P < 5.0 × 10(-3) for each ancestral background). High serum TNFα levels were positively correlated with high serum IFNα levels when tested in the same sample across all ancestral backgrounds (odds ratio range 1.76-1.86, P = 4.8 × 10(-3) by Fisher's combined probability test). While serum TNFα levels alone did not differ significantly among SLE patients of different ancestral backgrounds, the proportion of patients with concurrently high levels of TNFα and IFNα was highest in African Americans and lowest in European Americans (P = 5.0 × 10(-3) ). Serum TNFα levels were not associated with autoantibodies, clinical criteria for the diagnosis of SLE, or age at the time of sampling. Serum TNFα levels are high in many SLE patients, and we observed a positive correlation between serum TNFα and IFNα levels. These data support a role for TNFα in the pathogenesis of SLE across all ancestral backgrounds and suggest important cytokine subgroups within the disease. Copyright © 2012 by the American College of Rheumatology.

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

PubMed

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

2017-11-27

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

Evaluation of the Aptima HBV Quant assay vs. the COBAS TaqMan HBV test using the high pure system for the quantitation of HBV DNA in plasma and serum samples.

PubMed

Schalasta, Gunnar; Börner, Anna; Speicher, Andrea; Enders, Martin

2018-03-28

Proper management of patients with chronic hepatitis B virus (HBV) infection requires monitoring of plasma or serum HBV DNA levels using a highly sensitive nucleic acid amplification test. Because commercially available assays differ in performance, we compared herein the performance of the Hologic Aptima HBV Quant assay (Aptima) to that of the Roche Cobas TaqMan HBV test for use with the high pure system (HPS/CTM). Assay performance was assessed using HBV reference panels as well as plasma and serum samples from chronically HBV-infected patients. Method correlation, analytical sensitivity, precision/reproducibility, linearity, bias and influence of genotype were evaluated. Data analysis was performed using linear regression, Deming correlation analysis and Bland-Altman analysis. Agreement between the assays for the two reference panels was good, with a difference in assay values vs. target <0.5 log. Qualitative assay results for 159 clinical samples showed good concordance (88.1%; κ=0.75; 95% confidence interval: 0.651-0.845). For the 106 samples quantitated by both assays, viral load results were highly correlated (R=0.92) and differed on average by 0.09 log, with 95.3% of the samples being within the 95% limit of agreement of the assays. Linearity for viral loads 1-7 log was excellent for both assays (R2>0.98). The two assays had similar bias and precision across the different genotypes tested at low viral loads (25-1000 IU/mL). Aptima has a performance comparable with that of HPS/CTM, making it suitable for use for HBV infection monitoring. Aptima runs on a fully automated platform (the Panther system) and therefore offers a significantly improved workflow compared with HPS/CTM.

Measles & rubella outbreaks in Maharashtra State, India.

PubMed

Vaidya, Sunil R; Kamble, Madhukar B; Chowdhury, Deepika T; Kumbhar, Neelakshi S

2016-02-01

Under the outbreak-based measles surveillance in Maharashtra State the National Institute of Virology at Pune receives 3-5 serum samples from each outbreak and samples from the local hospitals in Pune for laboratory diagnosis. This report describes one year data on the measles and rubella serology, virus isolation and genotyping. Maharashtra State Health Agencies investigated 98 suspected outbreaks between January-December 2013 in the 20 districts. Altogether, 491 serum samples were received from 20 districts and 126 suspected cases from local hospitals. Samples were tested for the measles and rubella IgM antibodies by commercial enzyme immunoassay (EIA). To understand the diagnostic utility, a subset of serum samples (n=53) was tested by measles focus reduction neutralization test (FRNT). Further, 37 throat swabs and 32 urine specimens were tested by measles reverse transcription (RT)-PCR and positive products were sequenced. Virus isolation was performed in Vero hSLAM cells. Of the 98 suspected measles outbreaks, 61 were confirmed as measles, 12 as rubella and 21 confirmed as the mixed outbreaks. Four outbreaks remained unconfirmed. Of the 126 cases from the local hospitals, 91 were confirmed for measles and three for rubella. Overall, 93.6 per cent (383/409) confirmed measles cases were in the age group of 0-15 yr. Measles virus was detected in 18 of 38 specimens obtained from the suspected cases. Sequencing of PCR products revealed circulation of D4 (n=9) and D8 (n=9) strains. Four measles viruses (three D4 & one D8) were isolated. Altogether, 94 measles and rubella outbreaks were confirmed in 2013 in the State of Maharasthra indicating the necessity to increase measles vaccine coverage in the State.

Comparison of the Sensitivity of Laboratory Diagnostic Methods from a Well-Characterized Outbreak of Mumps in New York City in 2009

PubMed Central

Rosen, Jennifer B.; Doll, Margaret K.; McNall, Rebecca J.; McGrew, Marcia; Williams, Nobia; Lopareva, Elena N.; Barskey, Albert E.; Punsalang, Amado; Rota, Paul A.; Oleszko, William R.; Hickman, Carole J.; Zimmerman, Christopher M.; Bellini, William J.

2013-01-01

A mumps outbreak in upstate New York in 2009 at a summer camp for Orthodox Jewish boys spread into Orthodox Jewish communities in the Northeast, including New York City. The availability of epidemiologic information, including vaccination records and parotitis onset dates, allowed an enhanced analysis of laboratory methods for mumps testing. Serum and buccal swab samples were collected from 296 confirmed cases with onsets from September through December 2009. All samples were tested using the Centers for Disease Control and Prevention (CDC) capture IgM enzyme immunoassay (EIA) and a real-time reverse transcription-PCR (rRT-PCR) that targets the short hydrophobic gene. A subset of the samples (n = 205) was used to evaluate 3 commercial mumps IgM assays and to assess the sensitivity of using an alternative target gene (nucleoprotein) in the rRT-PCR protocol. Among 115 cases of mumps with 2 documented doses of measles, mumps, and rubella (MMR) vaccine, the CDC capture IgM EIA detected IgM in 51% of serum samples compared to 9% to 24% using three commercial IgM assays. The rRT-PCR that targeted the nucleoprotein gene increased RNA detection by 14% compared to that obtained with the original protocol. The ability to detect IgM improved when serum was collected 3 days or more after symptom onset, whereas sensitivity of RNA detection by rRT-PCR declined when buccal swabs were collected later than 2 days after onset. Selection of testing methods and timing of sample collection are important factors in the ability to confirm infection among vaccinated persons. These results reinforce the need to use virus detection assays in addition to serologic tests. PMID:23324519

Measles & rubella outbreaks in Maharashtra State, India

PubMed Central

Vaidya, Sunil R.; Kamble, Madhukar B.; Chowdhury, Deepika T.; Kumbhar, Neelakshi S.

2016-01-01

Background & objectives: Under the outbreak-based measles surveillance in Maharashtra State the National Institute of Virology at Pune receives 3-5 serum samples from each outbreak and samples from the local hospitals in Pune for laboratory diagnosis. This report describes one year data on the measles and rubella serology, virus isolation and genotyping. Methods: Maharashtra State Health Agencies investigated 98 suspected outbreaks between January-December 2013 in the 20 districts. Altogether, 491 serum samples were received from 20 districts and 126 suspected cases from local hospitals. Samples were tested for the measles and rubella IgM antibodies by commercial enzyme immunoassay (EIA). To understand the diagnostic utility, a subset of serum samples (n=53) was tested by measles focus reduction neutralization test (FRNT). Further, 37 throat swabs and 32 urine specimens were tested by measles reverse transcription (RT)-PCR and positive products were sequenced. Virus isolation was performed in Vero hSLAM cells. Results: Of the 98 suspected measles outbreaks, 61 were confirmed as measles, 12 as rubella and 21 confirmed as the mixed outbreaks. Four outbreaks remained unconfirmed. Of the 126 cases from the local hospitals, 91 were confirmed for measles and three for rubella. Overall, 93.6 per cent (383/409) confirmed measles cases were in the age group of 0-15 yr. Measles virus was detected in 18 of 38 specimens obtained from the suspected cases. Sequencing of PCR products revealed circulation of D4 (n=9) and D8 (n=9) strains. Four measles viruses (three D4 & one D8) were isolated. Interpretation & conclusions: Altogether, 94 measles and rubella outbreaks were confirmed in 2013 in the State of Maharasthra indicating the necessity to increase measles vaccine coverage in the State. PMID:27121521

Stability of 35 biochemical and immunological routine tests after 10 hours storage and transport of human whole blood at 21°C.

PubMed

Henriksen, Linda O; Faber, Nina R; Moller, Mette F; Nexo, Ebba; Hansen, Annebirthe B

2014-10-01

Suitable procedures for transport of blood samples from general practitioners to hospital laboratories are requested. Here we explore routine testing on samples stored and transported as whole blood in lithium-heparin or serum tubes. Blood samples were collected from 106 hospitalized patients, and analyzed on Architect c8000 or Advia Centaur XP for 35 analytes at base line, and after storage and transport of whole blood in lithium-heparin or serum tubes at 21 ± 1°C for 10 h. Bias and imprecision (representing variation from analysis and storage) were calculated from values at baseline and after storage, and differences tested by paired t-tests. Results were compared to goals set by the laboratory. We observed no statistically significant bias and results within the goal for imprecision between baseline samples and 10-h samples for albumin, alkaline phosphatase, antitrypsin, bilirubin, creatinine, free triiodothyronine, γ-glutamyl transferase, haptoglobin, immunoglobulin G, lactate dehydrogenase, prostate specific antigen, total carbon dioxide, and urea. Alanine aminotransferase, amylase, C-reactive protein, calcium, cholesterol, creatine kinase, ferritin, free thyroxine, immunoglobulin A, immunoglobulin M, orosomucoid, sodium, transferrin, and triglycerides met goals for imprecision, though they showed a minor, but statistically significant bias in results after storage. Cobalamin, folate, HDL-cholesterol, iron, phosphate, potassium, thyroid stimulating hormone and urate warranted concern, but only folate and phosphate showed deviations of clinical importance. We conclude that whole blood in lithium-heparin or serum tubes stored for 10 h at 21 ± 1°C, may be used for routine analysis without restrictions for all investigated analytes but folate and phosphate.

Inositol and hepatic lipidosis. I. Effect of inositol supplementation and time from parturition on liver and serum lipids in dairy cattle.

PubMed

Gerloff, B J; Herdt, T H; Wells, W W; Liesman, J S; Emery, R S

1986-06-01

Percutaneous liver biopsies and blood samples were obtained from 80 multiparous dairy cows in nine Michigan herds. Biopsies and samples were obtained serially over the peripartum period. Thirty-nine cows received 17 g of supplemental myoinositol in the diet to test its use as a possible lipotropic substance and 41 received a placebo. Liver biopsies were assayed for triglyceride (TG) and total myoinositol content. Serum was assayed for dextran precipitable cholesterol and non-esterified fatty acids (NEFA). Inositol supplementation had no effect on any of the lipid variables. There was a significant herd effect on liver inositol, serum dextran precipitable cholesterol and NEFA concentrations. Serum NEFA and liver TG concentrations increased in the immediate postpartum period, while dextran precipitable cholesterol decreased. A significant herd X period interaction existed for liver TG and serum dextran precipitable cholesterol concentrations. Liver TG and serum NEFA concentrations were positively correlated. Excessive infiltration of bovine liver with lipid at calving appears to be an exaggerated manifestation of normal metabolic changes.

[Clinical evaluation of a novel HBsAg quantitative assay].

PubMed

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

2007-07-01

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

Vitamin B12 absorption judged by measurement of holotranscobalamin, active vitamin B12: evaluation of a commercially available EIA kit.

PubMed

Greibe, Eva; Nexo, Ebba

2011-11-01

Active vitamin B12 absorption is followed by an increase in holotranscobalamin (holoTC) upon loading with a high physiological dose of the vitamin (the CobaSorb test). This study evaluates the use of a newly launched EIA kit for measurement of holoTC (active B12) in relation to the CobaSorb test. Intra-assay imprecision and linearity of the EIA kit was examined, employing serum pools of increasing holoTC concentrations. For the CobaSorb test, holoTC was measured before and after loading with 3-times 9 μg of vitamin B12 employing both the in-house ELISA and the EIA kit (n=25). The EIA kit showed an intra-assay CV between 2.2% and 5.8% for holoTC values ranging from 21 to 80 pmol/L. Employing diluted serum samples resulted in spurious high values of holoTC. The EIA kit performed well in relation to the CobaSorb test and classified the patients studied as capable of absorbing vitamin B12 (n=10) or not (n=15), as did the in-house ELISA. The Active B12 (holoTC) EIA kit proved suitable for use with the CobaSorb test, but not for analysis of diluted serum samples.

Seroprevalence of Brucella antibodies in camels in Katsina State, Nigeria.

PubMed

Salisu, U S; Kudi, C A; Bale, J O O; Babashani, M; Kaltungo, B Y; Saidu, S N A; Asambe, A; Baba, A Y

2017-06-01

A cross-sectional study was carried out to determine the status of Brucella infection in one-humped (Dromedary) camels in the North and Central senatorial districts of Katsina State, Nigeria. Nine hundred and eighty serum samples from live and slaughtered camels were tested. Modified Rose Bengal plate test (RBPT) and serum agglutination test (SAT) with ethylenediaminetetraacetic acid, (EDTA) were used as screening and standard tests, respectively. The prevalence of Brucella antibodies were 110 (11.2%) and 103 (10.5%) for RBPT and SAT, respectively. Of the 472 and 508 serum samples tested from the herds and abattoirs, respectively, 63 (13.3%) and 47 (9.3%) were positive by RBPT while 62 (13.1%) and 41 (8.1%) were positive by SAT, respectively. Based on the results, it was concluded that Brucella antibodies were present in camels in the study area. Poor management practices and mixing of camels with other species of livestock as well as unrestricted movement of camels were proposed to be the reasons for the prevalence of the disease in the study area. In view of the public health importance of the disease, it is recommended that there is the need to develop a strategic plan to decrease spread of brucellosis in the study area.

Zika Virus Testing Considerations: Lessons Learned from the First 80 Real-Time Reverse Transcription-PCR-Positive Cases Diagnosed in New York State.

PubMed

St George, Kirsten; Sohi, Inderbir S; Dufort, Elizabeth M; Dean, Amy B; White, Jennifer L; Limberger, Ronald; Sommer, Jamie N; Ostrowski, Stephanie; Wong, Susan J; Backenson, P Bryon; Kuhles, Daniel; Blog, Debra; Taylor, Jill; Hutton, Brad; Zucker, Howard A

2017-02-01

The performance and interpretation of laboratory tests for Zika virus (ZKV) continue to be evaluated. Serology is cross-reactive, laborious, and frequently difficult to interpret, and serum was initially solely recommended for molecular diagnosis. ZKV testing was initiated in January 2016 in New York State for symptomatic patients, pregnant women, their infants, and patients with Guillain-Barré syndrome who had traveled to areas with ZKV transmission. Subsequently, eligibility was expanded to pregnant women with sexual partners with similar travel histories. Serum and urine collected within 4 weeks of symptom onset or within 6 weeks of travel were tested with real-time reverse transcription-PCR (RT-PCR) assays targeting the ZKV envelope and NS2B genes. In this review of lessons learned from the first 80 positive cases in NYS, ZKV RNA was detected in urine only in 50 patients, in serum only in 19 patients, and in both samples concurrently in 11 patients, with average viral loads in urine a log higher than those in serum. Among 93 positive samples from the 80 patients, 41 were positive on both gene assays, 52 were positive on the envelope only, and none were positive on the NS2B only. Of the 80 infected patients, test results for 74 (93%) would have defined their infection status as not detected or equivocal if the requirement for positive results from two assay targets (two-target-positive requirement) in the initial federal guidance to public health laboratories was enforced, if urine was not tested, or if the extended eligibility time for molecular testing was not implemented. These changes facilitated more extensive molecular diagnosis of ZKV, reducing reliance on time-consuming and potentially inconclusive serology. Copyright © 2017 American Society for Microbiology.

Seroprevalence of human T cell leukaemia/lymphoma virus type I (HTLV-I) in pregnant women, patients attending venereological outpatient services and intravenous drug users from Slovenia.

PubMed

Poljak, M; Bednarik, J; Rednak, K; Seme, K; Kristancic, L; Celan-Lucu, B

1998-01-01

To establish current seroprevalence of human T cell leukaemia/lymphoma virus type I (HTLV-I) infection in some low- and high-risk populations from Slovenia, 10,369 and 869 serum samples collected during Slovenian 1994 unlinked surveys of human immunodeficiency viruses seroprevalence in pregnant women and patients attending venereological outpatient services, respectively, and 219 serum samples collected from Slovenian intravenous drug abusers during 1995 and 1996, were screened for the presence of anti-HTLV-I antibodies using commercial particle agglutination test Serodia HTLV-I (Fujirebio, Tokyo, Japan). Only one sample obtained from a pregnant woman was found repeatedly positive in the screening test. Presence of anti-HTLV-I antibodies in the reactive sample was undoubtedly confirmed with supplemental Western blot test. The prevalence of antibodies to HTLV-I in the Slovenian population might be somewhere between one in 10,000 (0.01%) and one in 15,000 (0.0066%), which is similar or even higher to prevalence rates in other European countries.

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

PubMed

Mozaffari, A A; Samadieh, H

2013-09-01

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

Detection of Multiple Pathogens in Serum Using Silica-Encapsulated Nanotags in a Surface-Enhanced Raman Scattering-Based Immunoassay.

PubMed

Neng, Jing; Li, Yina; Driscoll, Ashley J; Wilson, William C; Johnson, Patrick A

2018-06-06

A robust immunoassay based on surface-enhanced Raman scattering (SERS) has been developed to simultaneously detect trace quantities of multiple pathogenic antigens from West Nile virus, Rift Valley fever virus, and Yersinia pestis in fetal bovine serum. Antigens were detected by capture with silica-encapsulated nanotags and magnetic nanoparticles conjugated with polyclonal antibodies. The magnetic pull-down resulted in aggregation of the immune complexes, and the silica-encapsulated nanotags provided distinct spectra corresponding to each antigen captured. The limit of detection was ∼10 pg/mL in 20% fetal bovine serum, a significant improvement over previous studies in terms of sensitivity, level of multiplexing, and medium complexity. This highly sensitive multiplex immunoassay platform provides a promising method to detect various antigens directly in crude serum samples without the tedious process of sample preparation, which is desirable for on-site diagnostic testing and real-time disease monitoring.

Relationship of glomerular filtration rate based on serum iodixanol clearance to IRIS staging in cats with chronic kidney disease.

PubMed

Iwama, Ryosuke; Sato, Tsubasa; Katayama, Masaaki; Shimamura, Shunsuke; Satoh, Hiroshi; Ichijo, Toshihiro; Furuhama, Kazuhisa

2015-08-01

We examined the correlation between the glomerular filtration rate (GFR) estimated from an equation based on the serum iodixanol clearance technique and International Renal Interest Society (IRIS) stages of chronic kidney disease (CKD) in cats. The equation included the injection dose, sampling time, serum concentration and estimated volume of distribution (Vd) of the isotonic, nonionic, contrast medium iodixanol as a test tracer. The percent changes in the median basal GFR values calculated from the equation in CKD cats resembled those of IRIS stages 1-3. These data validate the association between the GFR derived from the simplified equation and IRIS stages based on the serum creatinine concentration in cats with CKD. They describe the GFR ranges determined using single-sample iodixanol clearance for healthy cats and cats with various IRIS stages of CKD.

Heterogeneity of the IgE response to allergenic determinants of cefaclor in serum samples from patients with cefaclor-induced anaphylaxis.

PubMed

Kim, Sang-Hoon; Choi, Jeong-Hee; Park, Hae-Sim

2005-06-01

Beta-lactam antibiotics, such as cefaclor, may cause IgE-mediated anaphylactic reactions. However, the clinically available serologic test has not been widely accepted, and the antigenic determinants of these drugs are unclear. To describe 4 cases of anaphylaxis caused by cefaclor in which a specific IgE response to cefaclor was demonstrated. Four patients with anaphylaxis to cefaclor and 35 nonatopic controls never exposed to cefaclor were studied. Skin tests and oral challenges with this drug were performed. The specific IgE response to the antigenic determinant of cefaclor-human serum albumin (HSA) conjugate was compared in each patient. The serum specific IgE to cefaclor-HSA conjugate was detected using enzyme-linked immunosorbent assay (ELISA). Also, ELISA inhibition studies using various concentrations of cefaclor-HSA, HSA alone, and free cefaclor were performed, as were hapten inhibition studies using cefaclor, cephalexin, cefadroxil, ampicillin, ceftriaxone, and cefotaxime. Three patients showed high levels of serum specific IgE to cefaclor-HSA and marked inhibition patterns to free cefaclor and cefaclor-HSA conjugate on ELISA inhibition testing. Hapten inhibition testing in 3 individual serum samples showed 2 different patterns. In patient 3, significant dose-dependent inhibitions (up to 92%) were noted with additions of free cefaclor and cefaclor-HSA conjugate, and lesser inhibitions (up to 74%) were noted with cephalexin, which shares the aminobenzyl side chain. In patients 1 and 2, marked dose-dependent inhibitions were noted only with additions of cefaclor-HSA conjugate and free cefaclor, whereas minimal inhibitions were noted with the other 5 compounds. The specific IgE response to cefaclor-HSA conjugate in patients with cefaclor anaphylaxis occurs against the hapten, in which heterogeneity of the antigenic determinant was noted to depend on the individual.

Doxycycline assay hair samples for testing long-term compliance treatment.

PubMed

Angelakis, Emmanouil; Armstrong, Nicholas; Nappez, Claude; Richez, Magalie; Chabriere, Eric; Raoult, Didier

2015-11-01

Many patients undergoing long-term doxycycline treatment do not regularly take their treatment because of photosensitivity. Our objective was to create an assay for determining doxycycline levels and to use hair samples for monitoring the compliance over a longer period of time. We tested sera and hair samples from patients treated with doxycycline by a suitable ultra-high performance liquid chromatography (UHPLC) based assay. We estimated that the speed of hair growth is roughly 1.25 cm per month and we were able to determine doxycycline levels over a 6-month period. We tested 14 patients treated with doxycycline and we found similar levels of doxycycline in the serum and the hair samples representing the last 4 months. Linear regression analysis revealed that the level of doxycycline in the serum remained stable over time (p = 0.7) but the level of doxycycline in the hair decreased significantly over time (p = 0.03) indicating a degradation of this molecule in the hair. We detected two patients who did not have antibiotic in the hair, indicating a lack of compliance that was also confirmed by interview. Hair samples can be used to test long-term compliance in patients to explain failures or relapses. Copyright © 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Salivary alterations in insulin-dependent diabetes mellitus.

PubMed

Belazi, M A; Galli-Tsinopoulou, A; Drakoulakos, D; Fleva, A; Papanayiotou, P H

1998-03-01

To examine the flow rate and composition of unstimulated whole saliva and of serum in children with newly diagnosed insulin-dependent diabetes mellitus (IDDM) and to compare these with values for a group of healthy controls. Cross-sectional. Diabetic Department of a University Hospital in Thessaloniki, Greece. The test group was made up of 10 recently diagnosed child patients with IDDM, aged 4-15 years and free of other systemic disease. Ten healthy children aged 5-17 years served as controls. Children were clinically examined before unstimulated saliva was collected over a 5 minute period and blood samples taken. Saliva and serum measurements included glucose, total proteins, albumin and immunoglobulins (IgA, IgG). No significant difference was seen in salivary flow rate between the two groups. Significantly greater concentrations of glucose were seen in saliva and serum in children with IDDM. Neither total protein nor albumin differed significantly between the two groups. Salivary IgA concentration was higher in the test group as was serum IgG. Findings in this sample suggest that changes in salivary composition may, together with estimated glucose levels, play a helpful diagnostic role in the early stages of IDDM in some children.

Quantitation of antibodies to Haemophilus influenzae type b in humans by enzyme-linked immunosorbent assay.

PubMed Central

Dahlberg, T

1981-01-01

The enzyme-linked immunosorbent assay was adapted to detect serum immunoglobulin G, immunoglobulin M, immunoglobulin A, and secretory immunoglobulin A antibodies to Haemophilus influenzae type b capsular polysaccharide in humans. I studied serum samples from 92 healthy children of various ages, 50 healthy adults, 24 patients with various H. influenzae type b infections, and 16 patients with clinical signs of epiglottis and cellulitis suspected to be caused by H. influenzae type b. The mean antibody titers of the sera from healthy children increased with age and reached adult levels in children more than 6 years old. A significant antibody response to capsular polysaccharide was observed in serum samples from the majority of patients with infections due to H. influenzae type b and in 4 of 16 patients with clinical signs of epiglottis and cellulitis. In addition to the enzyme-linked immunosorbent assay, the antibody responses of patients were tested by a bactericidal assay. When the two methods were compared, there was no evident correlation (r, about 0.22). The enzyme-linked immunosorbent assay was further adapted to test secretory immunoglobulin A antibodies specific to capsular polysaccharide in nasopharynx secretions and in milk samples from lactating women. Antibodies were detected in 12 of 24 secretions and 9 of 11 milk samples. PMID:7019237

[Demonstration of intraocular leptospira in 4 horses suffering from equine recurrent uveitis (ERU)].

PubMed

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

1998-01-01

Vitreous samples from 43 horses which underwent vitrectomy because of equine recurrent uveitis (ERU) were cultured for leptospires. Out of 4 vitreous samples (4/43 = 9%), leptospires could be isolated. In 3 cases, serovar grippotyphosa, and in one case, a serovar out of the serogroup Australis were identified. So for the first time, in several horses with ERU in vivo cultures of vitreous material were positive for leptospires. A strong evidence of association between leptospiral infection and uveitis is discussed for many years. In this investigation the leptospiral etiology is confirmed. Vitreous material from 42 and serum samples from 40 horses were tested for specific antibodies to leptospira by microagglutination test (MAT). In 34 vitreous samples (34/42 = 81%), leptospiral antibody titers of 1:50 or higher were detected. In 33 horses (33/40 = 83%) leptospiral antibody titers of 1:50 or higher could also be detected in the serum. Altogether, leptospiral antibodies were detected by the MAT in the serum and in the vitreous material of 39 of 43 horses (= 91%) subjected to vitrectomy. These results indicate, that ERU is probably often a sequel to systemic Leptospira interrogans infection. The presence of intact leptospires and specific antibodies in eyes affected with ERU indicates a local antibody production to leptospira organisms and/or their antigens.

Improvement for identification of heterophile antibody interference and AFP hook effect in immunoassays with multiplex suspension bead array system.

PubMed

Wang, Yajie; Yu, Jinsheng; Ren, Yuan; Liu, Li; Li, Haowen; Guo, Anchen; Shi, Congning; Fang, Fang; Juehne, Twyla; Yao, Jianer; Yang, Enhuan; Zhou, Xuelei; Kang, Xixiong

2013-11-15

A variety of immunoassays including multiplex suspension bead array have been developed for tumor marker detections; however, these assays could be compromised in their sensitivity and specificity by well-known heterophile antibody interference and hook effect. Using Luminex® multiplex suspension bead arrays, we modified protocols with two newly-developed solutions that can identify heterophile antibody interference and AFP hook effect. Effectiveness of the two solutions was assessed in serum samples from patients. Concentrations of 9 tumor markers in heterophile antibody positive samples assayed with Solution A, containing murine monoclonal antibodies and mouse serum, were significantly reduced when compared with those false high signals assayed without Solution A (all p<0.01). With incorporation of Solution H (fluorescent beads linked with AFP antigen), a new strategy for identification of AFP hook effect was established, and with this strategy AFP hook effect was identified effectively in serum samples with very high levels of AFP. Two proprietary solutions improve the identification of heterophile antibody interference and AFP hook effect. With these solutions, multiplex suspension bead arrays provide more reliable testing results in tumor marker detection where complex clinical serum samples are used. © 2013.

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

PubMed Central

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

2016-01-01

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

[35 leptospira isolated from the vitreous body of 32 horses with recurrent uveitis (ERU)].

PubMed

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

1999-01-01

130 vitreous samples, systematically collected in 1998 from 117 horses during vitrectomy, were cultured for the presence of leptospires. All horses suffered from equine recurrent uveitis (ERU), also known as periodic ophthalmia or moon blindness, and were treated surgically to combat painful attacks, and to preserve vision. In 35 out of 130 vitreous samples (35/130 = 26.9%), leptospires could be isolated. These isolates belong to the grippotyphosa serogroup (n = 31) and to the australis serogroup (n = 4). So, for the first time, leptospires were recovered from eyes in vivo in a large number of horses with ERU. Vitreous samples and one serum sample from each horse were also tested for leptospiral antibodies using the microscopic agglutination test (MAT). In 92 vitreous samples (92/130 = 70.7%) and 96 serum samples (96/117 = 82.0%) leptospiral antibodies were detected at a dilution of > 1:100. The presence of intact leptospires and specific antibodies in eyes affected with ERU demonstrates a local antibody production to leptospiral antigen. These results indicate an important etiological role of leptospires in equine recurrent uveitis.

Identification of bovine serum albumin as an IgE-reactive beef component in a dog with food hypersensitivity against beef.

PubMed

Ohmori, Keitaro; Masuda, Kenichi; Kawarai, Shinpei; Yasuda, Nobutaka; Sakaguchi, Masahiro; Tsujimoto, Hajime

2007-08-01

IgE-reactive beef components were examined by an immunoblot analysis using a serum from a dog with food hypersensitivity against beef. The immunoblot analysis revealed a distinct band at approximately 66 kDa and a faint band at approximately 50 kDa. The immunoblot analysis for serum IgE reactivity to bovine serum albumin (BSA) also revealed a positive band at 66 kDa. Serum IgE reactivity to the 66-kDa protein of beef was diminished by pre-incubating the serum sample with BSA. Furthermore, a positive reaction to BSA was detected in intradermal testing in the dog. These results clearly indicated that BSA was an IgE-reactive beef component in the dog with food hypersensitivity against beef.

Detection of human parvovirus B19 in serum samples from children under 5 years of age with rash-fever illnesses in the Democratic Republic of the Congo.

PubMed

Wawina, Tony Bokalanga; Tshiani, Olivier Mbaya; Ahuka, Steve Mundeke; Pukuta, Elisabeth Simbu; Aloni, Michel Ntetani; Kasanga, Christopher Jacob; Muyembe, Jean-Jacques Tamfum

2017-12-01

It has been demonstrated that infection with human parvovirus B19 (B19V) is associated with rash-fever illnesses. The present study aimed to investigate B19V as an aetiological agent of rash-fever syndromes in Congolese children confirmed as measles and rubella IgM-negative. An ELISA IgM test and PCR were performed to screen for B19V. A total of 177 archived serum samples were randomly selected from the measles biobank of the National Institute for Biomedical Research (INRB). Samples were investigated for anti-B19V IgM and B19V DNA. These samples originated from children <5years of age with measles-like rashes, previously confirmed as negative for both measles and rubella IgM. Out of 177 serum samples tested by ELISA and 168 tested by PCR, 109 were positive for B19V IgM antibodies (61.6%) and 87 (51.8%) were positive for B19V DNA. Positive samples in both assays were from all provinces of DRC. B19V plays a role in rash-fever illnesses in children under 5 years of age suspected of having measles or rubella infections in DRC. As an aetiological cause of rash and fever syndromes, the present study demonstrates that B19V should also be considered during the laboratory investigation of rash-fever illnesses in DRC, particularly in the paediatric population. There is a need to conduct further studies in order to gain a better understanding of the spatiotemporal pattern of B19V and to define the genotype(s) of B19V circulating in DRC. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

9 CFR 113.309 - Bovine Parainfluenza3 Vaccine.

Code of Federal Regulations, 2011 CFR

2011-01-01

...-challenge for serum antibody studies. (6) Satisfactory Test Criteria: (i) All virus isolations attempts... develop antibody titers of 1:32 or greater by day 6 ±2 days post-challenge. (8) A sequential test... parainfluenza, susceptible calves shall be used as test animals (20 vaccinates and five controls). Blood samples...

9 CFR 113.309 - Bovine Parainfluenza3 Vaccine.

Code of Federal Regulations, 2014 CFR

2014-01-01

...-challenge for serum antibody studies. (6) Satisfactory Test Criteria: (i) All virus isolations attempts... develop antibody titers of 1:32 or greater by day 6 ±2 days post-challenge. (8) A sequential test... parainfluenza, susceptible calves shall be used as test animals (20 vaccinates and five controls). Blood samples...

9 CFR 113.309 - Bovine Parainfluenza3 Vaccine.

Code of Federal Regulations, 2013 CFR

2013-01-01

...-challenge for serum antibody studies. (6) Satisfactory Test Criteria: (i) All virus isolations attempts... develop antibody titers of 1:32 or greater by day 6 ±2 days post-challenge. (8) A sequential test... parainfluenza, susceptible calves shall be used as test animals (20 vaccinates and five controls). Blood samples...

Stability of Chronic Hepatitis-Related Parameters in Serum Samples After Long-Term Storage.

PubMed

Yu, Rentao; Dan, Yunjie; Xiang, Xiaomei; Zhou, Yi; Kuang, Xuemei; Yang, Ge; Tang, Yulan; Liu, Mingdong; Kong, Weilong; Tan, Wenting; Deng, Guohong

2017-06-01

Serum samples are widely used in clinical research, but a comprehensive research of the stability of parameters relevant to chronic hepatitis and the effect of a relatively long-term (up to 10 years) storage on the stability have rarely been studied. To investigate the stability of chronic hepatitis-related parameters in serum samples after long-term storage. The storage stability of common clinical parameters such as total bile acid (TBA), total bilirubin (TBIL), potassium, cholesterol, and protein parameters such as alanine aminotransferase (ALT), creatine kinase (CK), γ-glutamyltransferase (GGT), albumin, high-density lipoprotein (HDL) and also hepatitis B virus (HBV) DNA, hepatitis C virus (HCV) RNA, hepatitis B surface antigen (HBsAg), and chemokine (C-X-C motif) ligand 10 (CXCL10) were tested in serum samples after storing at -20°C or -70°C for 1, 2, 3, 7, 8, and 10 years. Levels of TBA, TBIL, and protein parameters such as ALT, CK, GGT, HDL, and HBsAg decreased significantly, but levels of potassium and cholesterol increased significantly after long-term storage, whereas blood glucose and triglycerides were stable during storage. HBV DNA remained stable at -70°C but changed at -20°C, whereas HCV RNA was stable after 1-, 2-, and 3-year storage. CXCL10 was still detectable after 8-year storage. Low temperatures (-70°C/80°C) are necessary for storage of serum samples in chronic hepatitis B research after long-term storage.

Diagnostic performance of serological tests to detect antibodies against acute scrub typhus infection in central India.

PubMed

Pote, Kiran; Narang, Rahul; Deshmukh, Pradeep

2018-01-01

Differentiating scrub typhus from other acute febrile illness is difficult due to non specificity of clinical symptoms and relative absence of eschar in Indian population. The diagnosis thus relies mainly on laboratory tests. Antibody based serological tests are mainstay of scrub typhus diagnosis. Here, we evaluated the diagnostic performance of IgM ELISA, IgM IFA and ICT to detect antibodies against O. tsutsugamushi in acute serum of febrile patients. The serum samples from 600 randomly selected patients suffering from acute undifferentiated fever were tested by all the three tests mentioned above. We used latent class analysis to generate unbiased results as all the tests for scrub typhus diagnosis are imperfect and none of them can be considered as reference standard. We found that IgM ELISA with cutoff titer 0.5 OD has high diagnostic accuracy (sensitivity 99.9% and specificity 99.15) than IgM IFA (sensitivity 96.8% and specificity 99.7%) for scrub typhus diagnosis. ICT used in our study had very high specificity 100% but low sensitivity (38%) which would limit its use for acute serum samples. ICT being a screening or point of care test, has to be more sensitive while some compromise with specificity is affordable. Hence, optimal cutoff for ICT should be evaluated under different settings. IgM ELISA being simple and affordable could be an alternative diagnostic test to IgM IFA which is subjective and costly.

Development of a one-step immunochromatographic strip test using gold nanoparticles for the rapid detection of Salmonella typhi in human serum.

PubMed

Preechakasedkit, Pattarachaya; Pinwattana, Kulwadee; Dungchai, Wijitar; Siangproh, Weena; Chaicumpa, Wanpen; Tongtawe, Pongsri; Chailapakul, Orawon

2012-01-15

An immunochromatographic strip test using gold nanoparticles was developed for the rapid detection of Salmonella typhi (S. typhi) in human serum. The strip test based on the principle of sandwich immunoassay by the specific binding of antigens from S. typhi O901 and antibody of S. typhi O901 on a nitrocellulose membrane. Antibody-gold nanoparticle conjugate was used as the label and was coated onto a glass fiber membrane, which was used as a conjugate pad. To create a test and control zone, antibody of S. typhi O901 and an anti-IgG were dotted on the nitrocellulose membrane, respectively. Positive samples were displayed as red dots at the test and control zones of the nitrocellulose membrane, while negative samples resulted in a red dot only in the control zone. The limit of detection (LOD) was found to be 1.14×10(5) cfu mL(-1), which could be visually detected by the naked eye within 15 min. This strip test provided a lower detection limit and analysis time than a dot blot immunoassay (8.88×10(6) cfu mL(-1) for LOD and 110 min for reaction time). In addition, our immunochromatographic strip test was employed to detect S. typhi in human serum effectively, with high accuracy. This strip test offers great promise for a rapid, simple and low-cost analysis of S. typhi. Copyright © 2011 Elsevier B.V. All rights reserved.

Bias due to Preanalytical Dilution of Rodent Serum for Biochemical Analysis on the Siemens Dimension Xpand Plus

PubMed Central

Johns, Jennifer L.; Moorhead, Kaitlin A.; Hu, Jing; Moorhead, Roberta C.

2018-01-01

Clinical pathology testing of rodents is often challenging due to insufficient sample volume. One solution in clinical veterinary and exploratory research environments is dilution of samples prior to analysis. However, published information on the impact of preanalytical sample dilution on rodent biochemical data is incomplete. The objective of this study was to evaluate the effects of preanalytical sample dilution on biochemical analysis of mouse and rat serum samples utilizing the Siemens Dimension Xpand Plus. Rats were obtained from end of study research projects. Mice were obtained from sentinel testing programs. For both, whole blood was collected via terminal cardiocentesis into empty tubes and serum was harvested. Biochemical parameters were measured on fresh and thawed frozen samples run straight and at dilution factors 2–10. Dilutions were performed manually, utilizing either ultrapure water or enzyme diluent per manufacturer recommendations. All diluted samples were generated directly from the undiluted sample. Preanalytical dilution caused clinically unacceptable bias in most analytes at dilution factors four and above. Dilution-induced bias in total calcium, creatinine, total bilirubin, and uric acid was considered unacceptable with any degree of dilution, based on the more conservative of two definitions of acceptability. Dilution often caused electrolyte values to fall below assay range precluding evaluation of bias. Dilution-induced bias occurred in most biochemical parameters to varying degrees and may render dilution unacceptable in the exploratory research and clinical veterinary environments. Additionally, differences between results obtained at different dilution factors may confound statistical comparisons in research settings. Comparison of data obtained at a single dilution factor is highly recommended. PMID:29497614

Schoolchildren with Learning Difficulties Have Low Iron Status and High Anemia Prevalence

PubMed Central

Arcanjo, C. P. C.; Santos, P. R.

2016-01-01

Background. In developing countries there is high prevalence of iron deficiency anemia, which reduces cognitive performance, work performance, and endurance; it also causes learning difficulties and negative impact on development for infant population. Methods. The study concerns a case-control study; data was collected from an appropriate sample consisting of schoolchildren aged 8 years. The sample was divided into two subgroups: those with deficient initial reading skills (DIRS) (case) and those without (control). Blood samples were taken to analyze hemoglobin and serum ferritin levels. These results were then used to compare the two groups with Student's t-test. Association between DIRS and anemia was analyzed using odds ratio (OR). Results. Hemoglobin and serum ferritin levels of schoolchildren with DIRS were statistically lower when compared to those without, hemoglobin p = 0.02 and serum ferritin p = 0.04. DIRS was statistically associated with a risk of anemia with a weighted OR of 1.62. Conclusions. In this study, schoolchildren with DIRS had lower hemoglobin and serum ferritin levels when compared to those without. PMID:27703806

Schoolchildren with Learning Difficulties Have Low Iron Status and High Anemia Prevalence.

PubMed

Arcanjo, F P N; Arcanjo, C P C; Santos, P R

2016-01-01

Background . In developing countries there is high prevalence of iron deficiency anemia, which reduces cognitive performance, work performance, and endurance; it also causes learning difficulties and negative impact on development for infant population. Methods . The study concerns a case-control study; data was collected from an appropriate sample consisting of schoolchildren aged 8 years. The sample was divided into two subgroups: those with deficient initial reading skills (DIRS) (case) and those without (control). Blood samples were taken to analyze hemoglobin and serum ferritin levels. These results were then used to compare the two groups with Student's t -test. Association between DIRS and anemia was analyzed using odds ratio (OR). Results . Hemoglobin and serum ferritin levels of schoolchildren with DIRS were statistically lower when compared to those without, hemoglobin p = 0.02 and serum ferritin p = 0.04. DIRS was statistically associated with a risk of anemia with a weighted OR of 1.62. Conclusions . In this study, schoolchildren with DIRS had lower hemoglobin and serum ferritin levels when compared to those without.

Correlation of total serum immunoglobulin E level, sputum, and peripheral eosinophil count in assessing the clinical severity in bronchial asthma.

PubMed

Kumar, Roshan M; Pajanivel, R; Koteeswaran, G; Menon, Surendra K; Charles, Pravin Mv

2017-01-01

Asthma is a chronic inflammatory disorder of the airway with involvement of various cellular populations and release of many inflammatory mediators. Eosinophils and serum immunoglobulin E (IgE) are considered a good marker of airway inflammation in asthma. The correlation of clinical assessment with various markers of airway inflammation in asthma is not well established in the Indian population. This study aims to study the correlation of serum IgE, sputum eosinophil count, and peripheral eosinophil count with clinical severity of Asthma. This is a cross-sectional study involving 76 stable asthmatic patients of 18-60 years of age attending the pulmonary medicine OPD. Spirometry measured at baseline. Participants were categorized according to the GINA criteria based on clinical symptoms and pulmonary function test. Blood samples were collected for peripheral eosinophil count, serum IgE levels, and sputum samples for eosinophil count. All three parameters were compared with severity of asthma. The correlation of sputum eosinophil count, peripheral eosinophil count, and serum IgE with severity of asthma was analyzed by Pearson's Chi-square test, Fisher's exact test, and the correlation coefficient was reported together with standard error of the estimate. The mean age of patients in our study was 37.42 years and 56.6% were male. There was a significant inverse correlation between serum IgE levels and predicted forced expiratory volume 1 s (FEV1). Sputum eosinophilia was significantly seen in severe persistent asthma patients (19.7%). There was a significant inverse correlation between sputum eosinophil count and predicted FEV1and forced vital capacity. We also found there was a significant association between peripheral eosinophil count, sputum eosinophil count, and elevated serum IgE (g100 IU/mL) with severe persistent asthma. The assessment of sputum eosinophil count is simple, inexpensive, noninvasive, and direct measurement of airway inflammation. It could be the preferred method in monitoring airway inflammation and guided management in day-to-day practice.

Application of Fourier-transform infrared (FT-IR) spectroscopy for simple and easy determination of chylomicron-triglyceride and very low density lipoprotein-triglyceride.

PubMed

Sato, Kenichi; Seimiya, Masanori; Kodera, Yoshio; Kitamura, Akihide; Nomura, Fumio

2010-02-01

Fourier-transform infrared (FT-IR) spectroscopy is a simple and reagent-free physicochemical analysis method, and is a potential alternative to more time-consuming and labor-intensive procedures. In this study, we aimed to use FT-IR spectroscopy to determine serum concentrations of chylomicron-triglyceride (TG) and very low density lipoprotein (VLDL)-TG. We analyzed a chylomicron fraction and VLDL fraction, which had been obtained by ultracentrifugation, to search for wavelengths to designate to each fraction. Then, partial least square (PLS) calibrations were developed using a training set of samples, for which TG concentrations had been determined by conventional procedures. Validation was conducted with another set of samples using the PLS model to predict serum TG concentrations on the basis of the samples' IR spectra. We analyzed a total of 150 samples. Serum concentrations of chylomicron-TG and VLDL-TG estimated by FT-IR spectroscopy agreed well with those obtained by the reference method (r=0.97 for both lipoprotein fractions). FT-IR spectrometric analysis required 15mul of serum and was completed within 1min. Serum chylomicron-TG and VLDL-TG concentrations can be determined with FT-IR spectroscopy. This rapid and simple test may have a great impact on the management of patients with dyslipidemia. Copyright 2009. Published by Elsevier B.V.

Longitudinal Study of Hepatitis A Infection by Saliva Sampling: The Kinetics of HAV Markers in Saliva Revealed the Application of Saliva Tests for Hepatitis A Study.

PubMed

Amado Leon, Luciane Almeida; de Almeida, Adilson José; de Paula, Vanessa Salete; Tourinho, Renata Santos; Villela, Daniel Antunes Maciel; Gaspar, Ana Maria Coimbra; Lewis-Ximenez, Lia Laura; Pinto, Marcelo Alves

2015-01-01

Despite the increasing numbers of studies investigating hepatitis A diagnostic through saliva, the frequency and the pattern of hepatitis A virus (HAV) markers in this fluid still remains unknown. To address this issue, we carried on a longitudinal study to examine the kinetics of HAV markers in saliva, in comparison with serum samples. The present study followed-up ten patients with acute hepatitis A infection during 180 days post diagnosis (dpd). Total anti-HAV was detected in paired serum and saliva samples until the end of the follow-up, showing a peak titer at 90th. However, total anti-HAV level was higher in serum than in saliva samples. This HAV marker showed a probability of 100% to be detected in both serum and saliva during 180 dpd. The IgM anti-HAV could be detected in saliva up to 150 dpd, showing the highest frequency at 30th, when it was detected in all individuals. During the first month of HAV infection, this acute HAV marker showed a detection probability of 100% in paired samples. The detection of IgM anti-HAV in saliva was not dependent on its level in serum, HAV-RNA detection and/or viral load, since no association was found between IgM anti-HAV positivity in saliva and any of these parameter (p>0.05). Most of the patients (80%) were found to contain HAV-RNA in saliva, mainly at early acute phase (30th day). However, it was possible to demonstrate the HAV RNA presence in paired samples for more than 90 days, even after seroconversion. No significant relationship was observed between salivary HAV-RNA positivity and serum viral load, demonstrating that serum viral load is not predictive of HAV-RNA detection in saliva. Similar viral load was seen in paired samples (on average 104 copies/mL). These data demonstrate that the best diagnostic coverage can be achieved by salivary anti-HAV antibodies and HAV-RNA tests during 30-90 dpd. The long detection and high probability of specific-HAV antibodies positivity in saliva samples make the assessment of salivary antibodies a useful tool for diagnosis and epidemiological studies. The high frequency of HAV-RNA in saliva and the probability of detection of about 50%, during the first 30 dpd, demonstrate that saliva is also useful for molecular investigation of hepatitis A cases, mainly during the early course of infection. Therefore, the collection of saliva may provide a simple, cheap and non-invasive means of diagnosis, epidemiological surveys and monitoring of hepatitis A infection purposes.

Longitudinal Study of Hepatitis A Infection by Saliva Sampling: The Kinetics of HAV Markers in Saliva Revealed the Application of Saliva Tests for Hepatitis A Study

PubMed Central

Amado Leon, Luciane Almeida; de Almeida, Adilson José; de Paula, Vanessa Salete; Tourinho, Renata Santos; Villela, Daniel Antunes Maciel; Gaspar, Ana Maria Coimbra; Lewis-Ximenez, Lia Laura; Pinto, Marcelo Alves

2015-01-01

Despite the increasing numbers of studies investigating hepatitis A diagnostic through saliva, the frequency and the pattern of hepatitis A virus (HAV) markers in this fluid still remains unknown. To address this issue, we carried on a longitudinal study to examine the kinetics of HAV markers in saliva, in comparison with serum samples. The present study followed-up ten patients with acute hepatitis A infection during 180 days post diagnosis (dpd). Total anti-HAV was detected in paired serum and saliva samples until the end of the follow-up, showing a peak titer at 90th. However, total anti-HAV level was higher in serum than in saliva samples. This HAV marker showed a probability of 100% to be detected in both serum and saliva during 180 dpd. The IgM anti-HAV could be detected in saliva up to 150 dpd, showing the highest frequency at 30th, when it was detected in all individuals. During the first month of HAV infection, this acute HAV marker showed a detection probability of 100% in paired samples. The detection of IgM anti-HAV in saliva was not dependent on its level in serum, HAV-RNA detection and/or viral load, since no association was found between IgM anti-HAV positivity in saliva and any of these parameter (p>0.05). Most of the patients (80%) were found to contain HAV-RNA in saliva, mainly at early acute phase (30th day). However, it was possible to demonstrate the HAV RNA presence in paired samples for more than 90 days, even after seroconversion. No significant relationship was observed between salivary HAV-RNA positivity and serum viral load, demonstrating that serum viral load is not predictive of HAV-RNA detection in saliva. Similar viral load was seen in paired samples (on average 104 copies/mL). These data demonstrate that the best diagnostic coverage can be achieved by salivary anti-HAV antibodies and HAV-RNA tests during 30–90 dpd. The long detection and high probability of specific-HAV antibodies positivity in saliva samples make the assessment of salivary antibodies a useful tool for diagnosis and epidemiological studies. The high frequency of HAV-RNA in saliva and the probability of detection of about 50%, during the first 30 dpd, demonstrate that saliva is also useful for molecular investigation of hepatitis A cases, mainly during the early course of infection. Therefore, the collection of saliva may provide a simple, cheap and non-invasive means of diagnosis, epidemiological surveys and monitoring of hepatitis A infection purposes. PMID:26690904

Persistent HyperCKemia in Athletes

PubMed Central

Brancaccio, Paola; Maffulli, Nicola; Politano, Luisa; Lippi, Giuseppe; Limongelli, Francesco Mario

2011-01-01

Summary We compared the effects of exercise on serum levels of creatin kinase (CK) in athletes with persistent hyperCKemia at rest (CK group) and in healthy athletes (control group). Prospective controlled study. Eighteen male Caucasian athletes with high serum CK levels at rest (CK between 80 and 150 U/L) and 25 male Caucasian athletes with normal serum CK levels at rest (CK between 10 and 80 U/L) Main Outcome Measures Blood samples were collected at rest, 30 minutes, 6 hours, 24 hours, 48 hours and 72 hours after a progressive cycloergometer test to exhaustion. The levels of serum CK and its isoenzymes were measured. In the control group, serum CK values at rest were normal (48.18 ± 14.14 U/L). After exercise, they increased slightly, though they always remained <80 U/L, decreasing to the rest level after 48 hours. The CK group had serum CK levels at rest higher than normal (116.56 ± 33.30 U/L). Serum CK levels were still outwith the normal range after 48 hours (130.11 ± 46.95 U/L) and 72 hours (116.55 ± 24.84 U/L). Serum CK levels were significantly different in both groups both before and after progressive cycloergometer test to exhaustion. In athletes with high serum CK levels at rest, serum CK levels remained elevated and had a different kinetics after exercise when compared with healthy athletes. PMID:23738242

Identification of novel serum peptide biomarkers for high-altitude adaptation: a comparative approach

NASA Astrophysics Data System (ADS)

Yang, Juan; Li, Wenhua; Liu, Siyuan; Yuan, Dongya; Guo, Yijiao; Jia, Cheng; Song, Tusheng; Huang, Chen

2016-05-01

We aimed to identify serum biomarkers for screening individuals who could adapt to high-altitude hypoxia at sea level. HHA (high-altitude hypoxia acclimated; n = 48) and HHI (high-altitude hypoxia illness; n = 48) groups were distinguished at high altitude, routine blood tests were performed for both groups at high altitude and at sea level. Serum biomarkers were identified by comparing serum peptidome profiling between HHI and HHA groups collected at sea level. Routine blood tests revealed the concentration of hemoglobin and red blood cells were significantly higher in HHI than in HHA at high altitude. Serum peptidome profiling showed that ten significantly differentially expressed peaks between HHA and HHI at sea level. Three potential serum peptide peaks (m/z values: 1061.91, 1088.33, 4057.63) were further sequence identified as regions of the inter-α trypsin inhibitor heavy chain H4 fragment (ITIH4 347-356), regions of the inter-α trypsin inhibitor heavy chain H1 fragment (ITIH1 205-214), and isoform 1 of fibrinogen α chain precursor (FGA 588-624). Expression of their full proteins was also tested by ELISA in HHA and HHI samples collected at sea level. Our study provided a novel approach for identifying potential biomarkers for screening people at sea level who can adapt to high altitudes.

Clinical evaluation of a Mucorales-specific real-time PCR assay in tissue and serum samples.

PubMed

Springer, Jan; Lackner, Michaela; Ensinger, Christian; Risslegger, Brigitte; Morton, Charles Oliver; Nachbaur, David; Lass-Flörl, Cornelia; Einsele, Hermann; Heinz, Werner J; Loeffler, Juergen

2016-12-01

Molecular diagnostic assays can accelerate the diagnosis of fungal infections and subsequently improve patient outcomes. In particular, the detection of infections due to Mucorales is still challenging for laboratories and physicians. The aim of this study was to evaluate a probe-based Mucorales-specific real-time PCR assay (Muc18S) using tissue and serum samples from patients suffering from invasive mucormycosis (IMM). This assay can detect a broad range of clinically relevant Mucorales species and can be used to complement existing diagnostic tests or to screen high-risk patients. An advantage of the Muc18S assay is that it exclusively detects Mucorales species allowing the diagnosis of Mucorales DNA without sequencing within a few hours. In paraffin-embedded tissue samples this PCR-based method allowed rapid identification of Mucorales in comparison with standard methods and showed 91 % sensitivity in the IMM tissue samples. We also evaluated serum samples, an easily accessible material, from patients at risk from IMM. Mucorales DNA was detected in all patients with probable/proven IMM (100 %) and in 29 % of the possible cases. Detection of IMM in serum could enable an earlier diagnosis (up to 21 days) than current methods including tissue samples, which were gained mainly post-mortem. A screening strategy for high-risk patients, which would enable targeted treatment to improve patient outcomes, is therefore possible.

Metabonomics identifies serum metabolite markers of colorectal cancer.

PubMed

Tan, Binbin; Qiu, Yunping; Zou, Xia; Chen, Tianlu; Xie, Guoxiang; Cheng, Yu; Dong, Taotao; Zhao, Linjing; Feng, Bo; Hu, Xiaofang; Xu, Lisa X; Zhao, Aihua; Zhang, Menghui; Cai, Guoxiang; Cai, Sanjun; Zhou, Zhanxiang; Zheng, Minhua; Zhang, Yan; Jia, Wei

2013-06-07

Recent studies suggest that biofluid-based metabonomics may identify metabolite markers promising for colorectal cancer (CRC) diagnosis. We report here a follow-up replication study, after a previous CRC metabonomics study, aiming to identify a distinct serum metabolic signature of CRC with diagnostic potential. Serum metabolites from newly diagnosed CRC patients (N = 101) and healthy subjects (N = 102) were profiled using gas chromatography time-of-flight mass spectrometry (GC-TOFMS) and ultraperformance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-QTOFMS). Differential metabolites were identified with statistical tests of orthogonal partial least-squares-discriminant analysis (VIP > 1) and the Mann-Whitney U test (p < 0.05). With a total of 249 annotated serum metabolites, we were able to differentiate CRC patients from the healthy controls using an orthogonal partial least-squares-discriminant analysis (OPLS-DA) in a learning sample set of 62 CRC patients and 62 matched healthy controls. This established model was able to correctly assign the rest of the samples to the CRC or control groups in a validation set of 39 CRC patients and 40 healthy controls. Consistent with our findings from the previous study, we observed a distinct metabolic signature in CRC patients including tricarboxylic acid (TCA) cycle, urea cycle, glutamine, fatty acids, and gut flora metabolism. Our results demonstrated that a panel of serum metabolite markers is of great potential as a noninvasive diagnostic method for the detection of CRC.

Exposure of Piglets to Enteroviruses Investigated by an Immunoassay Based on the EV-G1 VP4 Peptide.

PubMed

Benkahla, Mehdi A; Sane, Famara; Desailloud, Rachel; Hober, Didier

2016-01-01

The aim of this study was to investigate the exposure of piglets to enteroviruses-G (EV-G) through the presence of antibodies in their serum. Serum samples were obtained from the vena cava of 10 piglets at 9 weeks of age and again 39 days later (day 39). They were tested using an immunoassay based on the EV-G1 VP4 peptide, since VP4 is highly conserved among the four Enterovirus capsid proteins, and by using a seroneutralization assay. For each serum collected on day 39 the optical density was high compared to the value obtained in serum collected earlier (p = 0.002). However, the titers of anti-EV-G1 serum neutralizing activity were not different in paired samples (p > 0.999). The sequence alignment of the EV-G1 VP4 peptide, encompassing 50 amino acids, used in the immunoassay showed 88% homology with EV-G, suggesting that antibodies directed toward other EV-G than EV-G1 may be detected. An immunoassay based on EV-G1 VP4 can detect an increased level of EV-G antibodies in piglet serum samples. Further studies are needed to determine whether this immunoassay may be useful for diagnosis and/or epidemiological studies and to monitor EV-G infection in pigs to evaluate strategies aimed to prevent enterovirus infections. © 2016 S. Karger AG, Basel.

[Evaluation of serum total carnitine values in persons with severe motor and intellectual disabilities with enteral (tube) feeding].

PubMed

Ohtaki, Ushio; Ozawa, Hiroshi; Ishizuka, Takehiro; Kamiishi, Akiko; Sasaki, Kyoko; Nakajima, Suemi; Katayama, Ayako; Arimoto, Kiyoshi; Yagihashi, Tatsuhiko; Kimiya, Satoshi

2012-09-01

The nutritive evaluation and the serum carnitine values were measured for persons with severe motor and intellectual disabilities with enteral (tube) feeding. In Shimada Rehabilitation Center, twenty one people who had serum albumin levels of 3.4 g/dl or less, and were taking nutrition with enteral (tube) feeding, were tested. Body weight, blood samples, and serum carnitine levels were measured. The total carnitine value was less than the standard value in 19 patients. The total carnitine value decreased in the group taking valporate sodium (VPA), compared to the values from the group non-taking VPA. From our evaluation, we think that daily carnitine supplements is essential for persons with sever motor and intellectual disabilities taking VPA to maintain carnitine levels in the blood, and regular urine test should be done for earlier detection secondary lack complications from the secondary lack of carnitine.

Detection and zoonotic potential of Trichinella spp. from free-range pig farming in Greece.

PubMed

Papatsiros, V G; Boutsini, S; Ntousi, D; Stougiou, D; Mintza, D; Bisias, A

2012-06-01

Trichinellosis is a serious parasitic zoonosis, which is widely distributed around the world. Pork meat is still the predominant source of outbreaks of human trichinellosis in many countries. The aim of this study is to examine the impact of Trichinella spp. as an important risk factor on the free-range pig farming sector in Greece. In 2009, during routine testing for the detection of Trichinella larvae at slaughterhouses and the National Reference Laboratory for Parasites (NRL), a total of 826,426 pigs were tested with the magnetic stirrer method for Trichinella spp. at slaughterhouses, including 2,892 samples from free-range pigs. Two positive samples were detected: one positive for Trichinella britovi and one positive for Trichinella spp. (unspecified) in the samples from wild farmed free-range pigs. It is alarming that one of these cases was connected with clinical signs of trichinellosis in five persons of the same family in northeastern Greece, who consumed undercooked pork meat from a free-range pig farm. During 2010, a total number of 1,295,034 pigs were tested with same method, including 4,159 samples from free-range pig farms. Five positive samples for Trichinella spp. (unspecified) were detected from 4,159 free-range pigs tested by the Greek NRL. Moreover, 363 serum samples from free-range pigs were serologically tested with enzyme-linked immunosorbent assay (ELISA). Moreover, 363 serum samples from farmed free-range pigs were serologically tested with ELISA, and 15 samples were found positive. Finally, the present study is the first report of detection of T. britovi in Greece. In conclusion, based on the results of the present study, Trichinella spp. is a high-risk factor for the free-range pig farming in Greece.

Potential metabolite markers of schizophrenia.

PubMed

Yang, J; Chen, T; Sun, L; Zhao, Z; Qi, X; Zhou, K; Cao, Y; Wang, X; Qiu, Y; Su, M; Zhao, A; Wang, P; Yang, P; Wu, J; Feng, G; He, L; Jia, W; Wan, C

2013-01-01

Schizophrenia is a severe mental disorder that affects 0.5-1% of the population worldwide. Current diagnostic methods are based on psychiatric interviews, which are subjective in nature. The lack of disease biomarkers to support objective laboratory tests has been a long-standing bottleneck in the clinical diagnosis and evaluation of schizophrenia. Here we report a global metabolic profiling study involving 112 schizophrenic patients and 110 healthy subjects, who were divided into a training set and a test set, designed to identify metabolite markers. A panel of serum markers consisting of glycerate, eicosenoic acid, β-hydroxybutyrate, pyruvate and cystine was identified as an effective diagnostic tool, achieving an area under the receiver operating characteristic curve (AUC) of 0.945 in the training samples (62 patients and 62 controls) and 0.895 in the test samples (50 patients and 48 controls). Furthermore, a composite panel by the addition of urine β-hydroxybutyrate to the serum panel achieved a more satisfactory accuracy, which reached an AUC of 1 in both the training set and the test set. Multiple fatty acids and ketone bodies were found significantly (P<0.01) elevated in both the serum and urine of patients, suggesting an upregulated fatty acid catabolism, presumably resulting from an insufficiency of glucose supply in the brains of schizophrenia patients.

Evaluation of Selected Borrelia burgdorferi lp54 Plasmid-Encoded Gene Products Expressed during Mammalian Infection as Antigens To Improve Serodiagnostic Testing for Early Lyme Disease

PubMed Central

Weiner, Zachary P.; Crew, Rebecca M.; Brandt, Kevin S.; Ullmann, Amy J.; Schriefer, Martin E.; Molins, Claudia R.

2015-01-01

Laboratory testing for the diagnosis of Lyme disease is performed primarily by serologic assays and is accurate for detection beyond the acute stage of the infection. Serodiagnostic assays to detect the early stages of infection, however, are limited in their sensitivity, and improvement is warranted. We analyzed a series of Borrelia burgdorferi proteins known to be induced within feeding ticks and/or during mammalian infection for their utility as serodiagnostic markers against a comprehensive panel of Lyme disease patient serum samples. The antigens were assayed for IgM and IgG reactivity in line immunoblots and separately by enzyme-linked immunosorbent assay (ELISA), with a focus on reactivity against early Lyme disease with erythema migrans (EM), early disseminated Lyme neuroborreliosis, and early Lyme carditis patient serum samples. By IgM immunoblotting, we found that recombinant proteins BBA65, BBA70, and BBA73 reacted with early Lyme EM samples at levels comparable to those of the OspC antigen used in the current IgM blotting criteria. Additionally, these proteins reacted with serum samples from patients with early neuroborreliosis and early carditis, suggesting value in detecting early stages of this disease progression. We also found serological reactivity against recombinant proteins BBA69 and BBA73 with early-Lyme-disease samples using IgG immunoblotting and ELISA. Significantly, some samples that had been scored negative by the Centers for Disease Control and Prevention-recommended 2-tiered testing algorithm demonstrated positive reactivity to one or more of the antigens by IgM/IgG immunoblot and ELISA. These results suggest that incorporating additional in vivo-expressed antigens into the current IgM/IgG immunoblotting tier in a recombinant protein platform assay may improve the performance of early-Lyme-disease serologic testing. PMID:26376927

Evaluation of Selected Borrelia burgdorferi lp54 Plasmid-Encoded Gene Products Expressed during Mammalian Infection as Antigens To Improve Serodiagnostic Testing for Early Lyme Disease.

PubMed

Weiner, Zachary P; Crew, Rebecca M; Brandt, Kevin S; Ullmann, Amy J; Schriefer, Martin E; Molins, Claudia R; Gilmore, Robert D

2015-11-01

Laboratory testing for the diagnosis of Lyme disease is performed primarily by serologic assays and is accurate for detection beyond the acute stage of the infection. Serodiagnostic assays to detect the early stages of infection, however, are limited in their sensitivity, and improvement is warranted. We analyzed a series of Borrelia burgdorferi proteins known to be induced within feeding ticks and/or during mammalian infection for their utility as serodiagnostic markers against a comprehensive panel of Lyme disease patient serum samples. The antigens were assayed for IgM and IgG reactivity in line immunoblots and separately by enzyme-linked immunosorbent assay (ELISA), with a focus on reactivity against early Lyme disease with erythema migrans (EM), early disseminated Lyme neuroborreliosis, and early Lyme carditis patient serum samples. By IgM immunoblotting, we found that recombinant proteins BBA65, BBA70, and BBA73 reacted with early Lyme EM samples at levels comparable to those of the OspC antigen used in the current IgM blotting criteria. Additionally, these proteins reacted with serum samples from patients with early neuroborreliosis and early carditis, suggesting value in detecting early stages of this disease progression. We also found serological reactivity against recombinant proteins BBA69 and BBA73 with early-Lyme-disease samples using IgG immunoblotting and ELISA. Significantly, some samples that had been scored negative by the Centers for Disease Control and Prevention-recommended 2-tiered testing algorithm demonstrated positive reactivity to one or more of the antigens by IgM/IgG immunoblot and ELISA. These results suggest that incorporating additional in vivo-expressed antigens into the current IgM/IgG immunoblotting tier in a recombinant protein platform assay may improve the performance of early-Lyme-disease serologic testing. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Retrospective analysis of serum and nasal mucus from cattle in Northern Ireland for evidence of infection with influenza A virus.

PubMed

Graham, D A; Calvert, V; McLaren, E

2002-02-16

Eighty-four pairs of acute and convalescent serum samples collected in 1998 and 1999 from 17 outbreaks of respiratory disease, milk drop syndrome or diarrhoea in cattle were tested by haemagglutination inhibition against human influenza viruses A/Eng/333/80 (HIN1) and A/Eng/427/88 (H3N2). Antibodies to these viruses were present in the convalescent sera of 56.5 per cent and 58.8 per cent cattle tested, respectively, with 56 per cent of the animals seroconverting to one or both viruses. Titres were typically higher to A/Eng/427/88 (H3N2). Further testing of a subset of 21 of these serum pairs against the predominant H1N1 and H3N2 human and porcine strains circulating when the samples were collected revealed that the highest reactivity, in terms of both the magnitude of the recorded titres and the number of positive sera, was to human H3N2 strains. The titres to human H1N1 strains and to both porcine subtypes were low or absent. Attempts to isolate influenza A virus from nasal mucus or swab samples from 142 cattle from 46 cases of respiratory disease and/or milk drop syndrome by passage in embryonated specific pathogen-free eggs were unsuccessful.

Antenatal screening for Down syndrome using serum placental growth factor with the combined, quadruple, serum integrated and integrated tests.

PubMed

Wald, Nicholas J; Bestwick, Jonathan P; George, Lynne M; Huttly, Wayne J

2012-01-01

To estimate the value of first or second trimester placental growth factor (PlGF) as an additional antenatal screening marker for Down syndrome. Nested case-control study. Antenatal screening service. 532 Down syndrome pregnancies and 1,155 matched unaffected pregnancies. Stored maternal serum samples (-40°C) were assayed for PlGF. Monte Carlo simulation was used to estimate the screening performance of PlGF with the Combined, Quadruple, serum Integrated and Integrated tests. Median PlGF levels in affected and unaffected pregnancies and screening performance (detection rates [DR] for specified false-positive rates [FPR] and vice versa). First trimester median PlGF was 15%, 28% and 39% lower in Down syndrome than unaffected pregnancies at 11, 12 and 13 completed weeks' gestation respectively (all p<0.001). Second trimester median PlGF was 31% lower at 14 weeks (p<0.001), and the difference decreased (6% lower at 17 weeks). At a 90% DR with first trimester markers measured at 13 weeks, adding PlGF decreased the FPR from 11.1 to 5.1% using the Combined test, 9.3% to 4.5% using the serum Integrated test, and 3.4% to 1.5% using the Integrated test (or 1.5 to 1.4% with first trimester markers measured at 11 weeks). Adding PlGF to the Quadruple test (measured at 15 weeks) decreased the FPR from 10.0% to 9.6% at a 90% DR. First trimester PlGF measurements improve the performance of antenatal screening for Down syndrome using the Combined, serum Integrated and Integrated tests. Second trimester PlGF measurements are of limited value.

Association of Mood Disorders with Serum Zinc Concentrations in Adolescent Female Students.

PubMed

Tahmasebi, Kobra; Amani, Reza; Nazari, Zahra; Ahmadi, Kambiz; Moazzen, Sara; Mostafavi, Seyed-Ali

2017-08-01

Among various factors influencing mood disorders, the impact of micronutrient deficiencies has attracted a great attention. Zinc deficiency is considered to play a crucial role in the onset and progression of mood disorders in different stages of life. The main objective of this study was to assess the correlation between serum zinc levels and mood disorders in high school female students. This cross-sectional study was conducted on a random sample of 100 representative high school female students. The participants completed 24-h food recall questionnaires to assess the daily zinc intakes. Serum zinc status was assessed using flame atomic absorption spectrometry, and zinc deficiency was defined accordingly. Mood disorders were estimated by calculating the sum of two test scores including Beck's depression inventory (BDI) and hospital anxiety depression scale (HADS) tests. General linear model (GLM) and Pearson's regression test were applied to show the correlation of serum zinc levels and mood disorder scores and the correlation between zinc serum levels and BDI scores, respectively. Dietary zinc intake was higher in subjects with normal zinc concentrations than that of zinc-deficient group (p = 0.001). Serum zinc levels were inversely correlated with BDI and HADS scores (p < 0.05). Each 10 μg/dL increment in serum zinc levels led to 0.3 and 0.01 decrease in depression and anxiety scores, respectively (p < 0.05). Serum zinc levels were inversely correlated with mood disorders including depression and anxiety in adolescent female students. Increasing serum levels of zinc in female students could improve their mood disorders.

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

PubMed

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

2015-06-15

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

Sensitivity and Specificity of Human Immunodeficiency Virus Rapid Serologic Assays and Testing Algorithms in an Antenatal Clinic in Abidjan, Ivory Coast

PubMed Central

Koblavi-Dème, Stéphania; Maurice, Chantal; Yavo, Daniel; Sibailly, Toussaint S.; N′guessan, Kabran; Kamelan-Tano, Yvonne; Wiktor, Stefan Z.; Roels, Thierry H.; Chorba, Terence; Nkengasong, John N.

2001-01-01

To evaluate serologic testing algorithms for human immunodeficiency virus (HIV) based on a combination of rapid assays among persons with HIV-1 (non-B subtypes) infection, HIV-2 infection, and HIV-1–HIV-2 dual infections in Abidjan, Ivory Coast, a total of 1,216 sera with known HIV serologic status were used to evaluate the sensitivity and specificity of four rapid assays: Determine HIV-1/2, Capillus HIV-1/HIV-2, HIV-SPOT, and Genie II HIV-1/HIV-2. Two serum panels obtained from patients recently infected with HIV-1 subtypes B and non-B were also included. Based on sensitivity and specificity, three of the four rapid assays were evaluated prospectively in parallel (serum samples tested by two simultaneous rapid assays) and serial (serum samples tested by two consecutive rapid assays) testing algorithms. All assays were 100% sensitive, and specificities ranged from 99.4 to 100%. In the prospective evaluation, both the parallel and serial algorithms were 100% sensitive and specific. Our results suggest that rapid assays have high sensitivity and specificity and, when used in parallel or serial testing algorithms, yield results similar to those of enzyme-linked immunosorbent assay-based testing strategies. HIV serodiagnosis based on rapid assays may be a valuable alternative in implementing HIV prevention and surveillance programs in areas where sophisticated laboratories are difficult to establish. PMID:11325995

Comparison of the serological tests ICT and ELISA for the diagnosis of alveolar echinococcosis in France☆

PubMed Central

Knapp, Jenny; Sako, Yasuhito; Grenouillet, Frédéric; Bresson-Hadni, Solange; Richou, Carine; Gbaguidi-Haore, Houssein; Ito, Akira; Millon, Laurence

2014-01-01

Serological diagnosis of alveolar echinococcosis (AE) is a key element for efficient patient treatment management. A rapid immunochromatography test kit (ICT) using the recombinant Em18 antigen (rEm18) was recently developed. The aim of our study was to assess this test on a panel of sera from French patients with alveolar echinococcosis and control patients. In a blind test, a total of 112 serum samples were tested including samples of AE (n = 30), cystic echinococcosis [CE] (n = 15), and polycystic echinococcosis [PE] (n = 1). For the comparison, 66 sera from patients with hepatocarcinoma, fascioliasis, toxocariasis, Caroli’s disease, or autoimmune chronic active hepatitis were used. The diagnostic test sets we used were the rEm18-ICT and two validated ELISAs with rEm18 and Em2-Em18 antigens, respectively. For the ICT, 27/30 sera from AE patients, 4/15 sera from CE patients and the PE patient serum were positive. One serum from the control panel (toxocariasis) was positive for the ICT. The rEm18-ICT sensitivity (90.0%) and specificity (92.7%) for detection of Em18-specific antibodies confirmed it as a relevant tool for AE diagnosis. The rEm18-ELISA had a sensitivity of 86.7% and specificity of 91.5%, and the Em2-Em18-ELISA had a sensitivity of 96.7% and specificity of 87.8%. However, when AE patient sera are recorded as weak in intensity with the ICT, we recommend a double reading and use of a reference sample if the ICT is used for patient follow-up. PMID:25058754

Comparison of the serological tests ICT and ELISA for the diagnosis of alveolar echinococcosis in France.

PubMed

Knapp, Jenny; Sako, Yasuhito; Grenouillet, Frédéric; Bresson-Hadni, Solange; Richou, Carine; Gbaguidi-Haore, Houssein; Ito, Akira; Millon, Laurence

2014-01-01

Serological diagnosis of alveolar echinococcosis (AE) is a key element for efficient patient treatment management. A rapid immunochromatography test kit (ICT) using the recombinant Em18 antigen (rEm18) was recently developed. The aim of our study was to assess this test on a panel of sera from French patients with alveolar echinococcosis and control patients. In a blind test, a total of 112 serum samples were tested including samples of AE (n = 30), cystic echinococcosis [CE] (n = 15), and polycystic echinococcosis [PE] (n = 1). For the comparison, 66 sera from patients with hepatocarcinoma, fascioliasis, toxocariasis, Caroli's disease, or autoimmune chronic active hepatitis were used. The diagnostic test sets we used were the rEm18-ICT and two validated ELISAs with rEm18 and Em2-Em18 antigens, respectively. For the ICT, 27/30 sera from AE patients, 4/15 sera from CE patients and the PE patient serum were positive. One serum from the control panel (toxocariasis) was positive for the ICT. The rEm18-ICT sensitivity (90.0%) and specificity (92.7%) for detection of Em18-specific antibodies confirmed it as a relevant tool for AE diagnosis. The rEm18-ELISA had a sensitivity of 86.7% and specificity of 91.5%, and the Em2-Em18-ELISA had a sensitivity of 96.7% and specificity of 87.8%. However, when AE patient sera are recorded as weak in intensity with the ICT, we recommend a double reading and use of a reference sample if the ICT is used for patient follow-up. © J. Knapp et al., published by EDP Sciences, 2014.

Quality of different in-clinic test systems for feline immunodeficiency virus and feline leukaemia virus infection.

PubMed

Hartmann, Katrin; Griessmayr, Pascale; Schulz, Bianka; Greene, Craig E; Vidyashankar, Anand N; Jarrett, Os; Egberink, Herman F

2007-12-01

Many new diagnostic in-house tests for identification of feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) infection have been licensed for use in veterinary practice, and the question of the relative merits of these kits has prompted comparative studies. This study was designed to define the strengths and weaknesses of seven FIV and eight FeLV tests that are commercially available. In this study, 536 serum samples from randomly selected cats were tested. Those samples reacting FIV-positive in at least one of the tests were confirmed by Western blot, and those reacting FeLV-positive were confirmed by virus isolation. In addition, a random selection of samples testing negative in all test systems was re-tested by Western blot (100 samples) and by virus isolation (81 samples). Specificity, sensitivity, positive and negative predictive values of each test and the quality of the results were compared.

Influenza A Virus Surveillance Based on Pre-Weaning Piglet Oral Fluid Samples.

PubMed

Panyasing, Y; Goodell, C; Kittawornrat, A; Wang, C; Levis, I; Desfresne, L; Rauh, R; Gauger, P C; Zhang, J; Lin, X; Azeem, S; Ghorbani-Nezami, S; Yoon, K-J; Zimmerman, J

2016-10-01

Influenza A virus (IAV) surveillance using pre-weaning oral fluid samples from litters of piglets was evaluated in four ˜12 500 sow and IAV-vaccinated, breeding herds. Oral fluid samples were collected from 600 litters and serum samples from their dams at weaning. Litter oral fluid samples were tested for IAV by virus isolation, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), RT-PCR subtyping and sequencing. Commercial nucleoprotein (NP) enzyme-linked immunosorbent assay (ELISA) kits and NP isotype-specific assays (IgM, IgA and IgG) were used to characterize NP antibody in litter oral fluid and sow serum. All litter oral fluid specimens (n = 600) were negative by virus isolation. Twenty-five oral fluid samples (25/600 = 4.2%) were qRT-PCR positive based on screening (Laboratory 1) and confirmatory testing (Laboratory 2). No hemagglutinin (HA) and neuraminidase (NA) gene sequences were obtained, but matrix (M) gene sequences were obtained for all qRT-PCR-positive samples submitted for sequencing (n = 18). Genetic analysis revealed that all M genes sequences were identical (GenBank accession no. KF487544) and belonged to the triple reassortant influenza A virus M gene (TRIG M) previously identified in swine. The proportion of IgM- and IgA-positive samples was significantly higher in sow serum and litter oral fluid samples, respectively (P < 0.01). Consistent with the extensive use of IAV vaccine, no difference was detected in the proportion of IgG- and blocking ELISA-positive sow serum and litter oral fluids. This study supported the use of oral fluid sampling as a means of conducting IAV surveillance in pig populations and demonstrated the inapparent circulation of IAV in piglets. Future work on IAV oral fluid diagnostics should focus on improved procedures for virus isolation, subtyping and sequencing of HA and NA genes. The role of antibody in IAV surveillance remains to be elucidated, but longitudinal assessment of specific antibody has the potential to provide information regarding patterns of infection, vaccination status and herd immunity. © 2014 Blackwell Verlag GmbH.

Highly accurate detection of ovarian cancer using CA125 but limited improvement with serum matrix-assisted laser desorption/ionization time-of-flight mass spectrometry profiling.

PubMed

Tiss, Ali; Timms, John F; Smith, Celia; Devetyarov, Dmitry; Gentry-Maharaj, Aleksandra; Camuzeaux, Stephane; Burford, Brian; Nouretdinov, Ilia; Ford, Jeremy; Luo, Zhiyuan; Jacobs, Ian; Menon, Usha; Gammerman, Alex; Cramer, Rainer

2010-12-01

Our objective was to test the performance of CA125 in classifying serum samples from a cohort of malignant and benign ovarian cancers and age-matched healthy controls and to assess whether combining information from matrix-assisted laser desorption/ionization (MALDI) time-of-flight profiling could improve diagnostic performance. Serum samples from women with ovarian neoplasms and healthy volunteers were subjected to CA125 assay and MALDI time-of-flight mass spectrometry (MS) profiling. Models were built from training data sets using discriminatory MALDI MS peaks in combination with CA125 values and tested their ability to classify blinded test samples. These were compared with models using CA125 threshold levels from 193 patients with ovarian cancer, 290 with benign neoplasm, and 2236 postmenopausal healthy controls. Using a CA125 cutoff of 30 U/mL, an overall sensitivity of 94.8% (96.6% specificity) was obtained when comparing malignancies versus healthy postmenopausal controls, whereas a cutoff of 65 U/mL provided a sensitivity of 83.9% (99.6% specificity). High classification accuracies were obtained for early-stage cancers (93.5% sensitivity). Reasons for high accuracies include recruitment bias, restriction to postmenopausal women, and inclusion of only primary invasive epithelial ovarian cancer cases. The combination of MS profiling information with CA125 did not significantly improve the specificity/accuracy compared with classifications on the basis of CA125 alone. We report unexpectedly good performance of serum CA125 using threshold classification in discriminating healthy controls and women with benign masses from those with invasive ovarian cancer. This highlights the dependence of diagnostic tests on the characteristics of the study population and the crucial need for authors to provide sufficient relevant details to allow comparison. Our study also shows that MS profiling information adds little to diagnostic accuracy. This finding is in contrast with other reports and shows the limitations of serum MS profiling for biomarker discovery and as a diagnostic tool.

Identification of cattle, buffaloes and rodents as reservoir animals of Leptospira in the District of Gampaha, Sri Lanka.

PubMed

Denipitiya, D T H; Chandrasekharan, N V; Abeyewickreme, W; Hartskeerl, R A; Hapugoda, M D

2017-03-23

Leptospirosis is an important emerging infectious disease in Sri Lanka. Rats are the most important reservoir of Leptospira but domestic and wild mammals may also act as important maintenance or accidental hosts. In Sri Lanka, knowledge of reservoir animals of leptospires is poor. The objective of this study was to identify potential reservoir animals of Leptospira in the District of Gampaha, Sri Lanka. Blood and kidney samples were collected from 38 rodents and mid-stream urine samples were randomly collected from 45 cattle and five buffaloes in the District of Gampaha. Kidney and urine samples were tested by real-time polymerase chain reaction (PCR) and serum samples were tested by the microscopic agglutination test (MAT). Of the 38 rodent kidney samples, 11% (4/38) were positive by real-time PCR. The prevalence of leptospiral carriage was 11% (3/26) and 8% (1/12) in female and male rodents, respectively. Three rodent serum samples were positive by MAT. Of the 50 cattle/buffalo urine samples tested, 10% (5/50) were positive by real-time PCR. The prevalence of leptospiral carriage was 9% (4/45) and 20% (1/5) in cattle and buffaloes, respectively. Results of PCR and MAT showed that Leptospira were present in a significant proportion of the rodents and farm animals tested in this study and suggest that these (semi-) domestic animals form an infection reservoir for Leptospira. Therefore, there is a potential zoonotic risk to public health, most notably to farmers in this area.

Urine and serum fetuin-A levels in patients with urolithiasis.

PubMed

Arora, Rajat; Abrol, Nitin; Antonisamy, B; Vanitha, S; Chandrasingh, J; Kumar, Santosh; Kekre, Nitin; Devasia, Antony

2017-01-01

Fetuin-A is a glycoprotein secreted by liver and has been shown to inhibit extraosseous mineralization. Urolithiasis may be a manifestation in the urinary tract due to fetuin deficiency in urine. The objective of this study was to compare the 24-h urine and serum fetuin-A levels of patients with and without urolithiasis. Serum and 24-h urine fetuin-A levels were measured in 41 patients with bilateral, multiple, or recurrent urinary tract calculi (Group A) and 41 matched controls with no calculi (Group B). Fetuin levels were measured by enzyme linked immunosorbent assay. Serum and urine fetuin-A levels in the two groups were compared. The median (range) 24-h urine fetuin-A value in Group A was 11.9 (1.12-221) mg/day and in Group B was 37.7 (1.28-125) mg/day. This difference was statistically significant (Mann-Whitney test, P = 0.0169). The median (range) serum fetuin-A in Group A was 0.67 (0.05-2.68) g/L and in Group B was 0.99 (0.01-5.5) g/L. The difference between serum values in the two arms was not statistically significant (Mann-Whitney test, P = 0.1817). However, the serum creatinine-adjusted mean log serum fetuin and urine fetuin were significantly different in the two arms ( P = 0.003). The mean ± standard deviation (range) serum creatinine in Group A was 0.98 ± 0.25 (0.56-1.58) mg% and in Group B was 0.83 ± 0.16 (0.58-1.18) mg% (two sample t -test, P = 0.0031). Patients with urolithiasis have lower urine fetuin-A and creatinine-adjusted serum fetuin-A levels.

Assessment of cytologic evaluation of preputial epithelial cells as a diagnostic test for detection of adrenocortical disease in castrated ferrets.

PubMed

Protain, Holly J; Kutzler, Michelle A; Valentine, Beth A

2009-05-01

To determine whether results of cytologic evaluation of preputial epithelial cells correspond to results of a serum endocrine hormone assay and clinical signs associated with adrenocortical disease in castrated ferrets. 13 clinically normal ferrets and 8 ferrets with signs of adrenocortical disease. Blood and preputial lavage samples were collected from each ferret. Serum samples were submitted to the University of Tennessee Veterinary Diagnostic Laboratory for performance of an endocrine hormone assay. Differential epithelial cell counts were performed on preputial lavage samples to determine the percentage of cornified cells. Results of cytologic evaluation were compared with results of the endocrine hormone assay and clinical status of ferrets. The percentage of cornified preputial epithelial cells was not significantly correlated with serum 17B-estradiol or androstenedione concentration but was significantly correlated with serum 17-hydroxyprogesterone concentration (r = 0.60). The percentage of cornified preputial epithelial cells was higher in ferrets with clinical signs of adrenocortical disease (mean +/- SD, 71.3 +/- 16.9%) than in clinically normal ferrets (55.5 +/- 19.0%). Cornification of preputial epithelial cells was correlated with an increase in serum 17-hydroxyprogesterone concentration as well as clinical signs of adrenocortical disease in castrated ferrets. Additional investigation is needed to elucidate the mechanism of preputial epithelial cell cornification in castrated ferrets.

Roles of circulating WNT-signaling proteins and WNT-inhibitors in human adiposity, insulin resistance, insulin secretion, and inflammation.

PubMed

Almario, R U; Karakas, S E

2015-02-01

Wingless-type MMTV integration site family member (WNT) signaling and WNT-inhibitors have been implicated in regulation of adipogenesis, insulin resistance, pancreatic function, and inflammation. Our goal was to determine serum proteins involved in WNT signaling (WNT5 and WISP2) and WNT inhibition (SFRP4 and SFRP5) as they relate to obesity, serum adipokines, insulin resistance, insulin secretion, and inflammation in humans. Study population comprised 57 insulin resistant women with polycystic ovary syndrome (PCOS) and 27 reference women. In a cross-sectional study, blood samples were obtained at fasting, during oral, and frequently sampled intravenous glucose tolerance tests. Serum WNT5, WISP2, and SFRP4 concentrations did not differ between PCOS vs. reference women. Serum WNT5 correlated inversely with weight both in PCOS and reference women, and correlated directly with insulin response during oral glucose tolerance test in PCOS women. Serum WISP2 correlated directly with fatty acid binding protein 4. Serum SFRP5 did not differ between obese (n=32) vs. nonobese (n=25) PCOS women, but reference women had lower SFRP5 (p<5×10(-6) as compared to both PCOS groups). Serum SFRP5 correlated inversely with IL-1β, TNF-α, cholesterol, and apoprotein B. These findings demonstrated that WNT5 correlated inversely with adiposity and directly with insulin response, and the WNT-inhibitor SFRP5 may be anti-inflammatory. Better understanding of the role of WNT signaling in obesity, insulin resistance, insulin secretion, lipoprotein metabolism, and inflammation is important for prevention and treatment of metabolic syndrome, diabetes and cardiovascular disease. © Georg Thieme Verlag KG Stuttgart · New York.

Serum glycerophosphate levels are increased in Japanese men with type 2 diabetes.

PubMed

Daimon, Makoto; Soga, Tomoyoshi; Hozawa, Atsushi; Oizumi, Toshihide; Kaino, Wataru; Takase, Kaoru; Karasawa, Shigeru; Jimbu, Yumi; Wada, Kiriko; Kameda, Wataru; Susa, Shinji; Kayama, Takamasa; Saito, Kaori; Tomita, Masaru; Kato, Takeo

2012-01-01

To identify metabolites showing changes in serum levels among Japanese male with diabetes. We performed metabolite profiling by coupling capillary electrophoresis with electrospray ionization time-of-flight mass spectrometry using fasting serum samples from Japanese male subjects with diabetes (n=17), impaired glucose tolerance (IGT; n=5) and normal glucose tolerance (NGT; n=14). Other than the expected differences in characteristics related to abnormal glucose metabolism, the percent body fat was significantly different among subjects with diabetes, IGT and NGT (27.3±6.2, 22.2±4.5 and 19.2±6.0%, respectively, p=0.0022). Therefore, percent body fat was considered as a possible confounding factor in subsequent analyses. Of 560 metabolites detected using our platform, the levels of 74 metabolites were quantified in all of the serum samples. Significant differences between diabetes and NGT were observed for 24 metabolites. The top-ranked metabolite was glycerol-3-phophate (glycerophosphate), which was significantly higher in subjects with diabetes than in those with NGT, even after Bonferroni correction for multiple testing (11.7±3.6 vs. 6.4±1.9 µM, respectively; corrected p=0.0222). Stepwise multiple regression analyses revealed that serum glycerophosphate levels were significantly correlated with 2-h plasma glucose after a 75-g oral glucose tolerance test (r=0.553, p=0.0005), independently of other characteristics, including FPG and HbA1c. Serum glycerophosphate levels were found to be elevated in Japanese men with diabetes, and correlated with 2-h PG, independent of FPG and HbA1c. Namely, serum glycerophosphate level at fasting condition can be a marker for predicting glucose intolerance. These results warrant further studies to evaluate the relevance of glycerophosphate in the pathophysiology of diabetes.

Searching for the best real-time RT-PCRs to detect Zika virus infections: the importance of comparing several protocols.

PubMed

de Moraes, F M; Espósito, D L A; Klein, T M; da Fonseca, B A L

2018-01-01

Clinical manifestations of Zika, dengue, and chikungunya virus infections are very similar, making it difficult to reach a diagnosis based only on clinical grounds. In addition, there is an intense cross-reactivity between antibodies directed to Zika virus and other flaviviruses, and an accurate Zika diagnosis is best achieved by real-time RT-PCR. However, some real-time RT-PCR show better performance than others. To reach the best possible Zika diagnosis, the analytic sensitivity of some probe-based real-time RT-PCR amplifying Zika virus RNA was evaluated in spiked and clinical samples. We evaluated primers and probes to detect Zika virus, which had been published before, and tested sensitivity using serum spiked and patient samples by real-time RT-PCR. When tested against spiked samples, the previously described primers showed different sensitivity, with very similar results when samples from patients (serum and urine) were analyzed. Real-time RT-PCR designed to amplify Zika virus NS1 showed the best analytical sensitivity for all samples.

Proposal and validation of prognostic scoring systems for IgG and IgA monoclonal gammopathies of undetermined significance.

PubMed

Rossi, Francesca; Petrucci, Maria Teresa; Guffanti, Andrea; Marcheselli, Luigi; Rossi, Davide; Callea, Vincenzo; Vincenzo, Federico; De Muro, Marianna; Baraldi, Alessandra; Villani, Oreste; Musto, Pellegrino; Bacigalupo, Andrea; Gaidano, Gianluca; Avvisati, Giuseppe; Goldaniga, Maria; Depaoli, Lorenzo; Baldini, Luca

2009-07-01

The presenting clinico-hematologic features of 1,283 patients with IgG and IgA monoclonal gammopathies of undetermined significance (MGUS) were correlated with the frequency of evolution into multiple myeloma (MM). Two IgG MGUS populations were evaluated: a training sample (553 patients) and a test sample (378 patients); the IgA MGUS population consisted of 352 patients. Forty-seven of the 553 training group patients and 22 of 378 test group IgG patients developed MM after a median follow-up of 6.7 and 3.6 years, respectively. Multivariate analysis showed that serum monoclonal component (MC) levels of < or =1.5 g/dL, the absence of light-chain proteinuria and normal serum polyclonal immunoglobulin levels defined a prognostically favorable subset of patients, and could be used to stratify the patients into three groups at different 10-year risk of evolution (hazard ratio, 1.0, 5.04, 11.2; P < 0.001). This scoring system was validated in the test sample. Thirty of the 352 IgA patients developed MM after a median follow-up of 4.8 years, and multivariate analysis showed that hemoglobin levels of <12.5 g/dL and reduced serum polyclonal immunoglobulin correlated with progression. A pooled statistical analysis of all of the patients confirmed the validity of Mayo Clinic risk model showing that IgA class, serum MC levels, and light-chain proteinuria are the most important variables correlated with disease progression. Using simple variables, we validated a prognostic model for IgG MGUS. Among the IgA cases, the possible prognostic role of hemoglobin emerged in addition to a decrease in normal immunoglobulin levels.

Human serum miR-34a as an indicator of exposure to ionizing radiation.

PubMed

Halimi, Mohammad; Shahabi, Ahmad; Moslemi, Dariush; Parsian, Hadi; Asghari, S Mohsen; Sariri, Reyhaneh; Yeganeh, Farshid; Zabihi, Ebrahim

2016-11-01

Radiation exposure in industrial accidents or nuclear device attacks is a major public health concern. There is an urgent need for markers that rapidly identify people exposed to ionizing radiation (IR). Finding a blood-based marker is advantageous because of the ease of sample collection. This study was designed to test the hypothesis that serum miR-34a could serve as an indicator of exposure to IR. Therefore, 44 women with breast cancer, where radiotherapy was part of their therapeutic protocol, were investigated in this study. After demonstrating the appropriateness of our microRNA (miRNA) extraction efficiency and miRNA assay in human serum, we analyzed the miR-34a level in paired serum samples before and after radiotherapy. Fifty Gy X-ray irradiation in daily dose fractions of 2 Gy, 5 days per week, was used in this study. We demonstrated that IR significantly increased serum level of miR-34a. By measuring miR-34a in serum, we could distinguish irradiated patients with sensitivity of 65 % and specificity of 75 %. According to this study, serum miR-34a has the potential to be used as an indicator of radiation exposure.

A Serum Protein Profile Predictive of the Resistance to Neoadjuvant Chemotherapy in Advanced Breast Cancers*

PubMed Central

Hyung, Seok-Won; Lee, Min Young; Yu, Jong-Han; Shin, Byunghee; Jung, Hee-Jung; Park, Jong-Moon; Han, Wonshik; Lee, Kyung-Min; Moon, Hyeong-Gon; Zhang, Hui; Aebersold, Ruedi; Hwang, Daehee; Lee, Sang-Won; Yu, Myeong-Hee; Noh, Dong-Young

2011-01-01

Prediction of the responses to neoadjuvant chemotherapy (NACT) can improve the treatment of patients with advanced breast cancer. Genes and proteins predictive of chemoresistance have been extensively studied in breast cancer tissues. However, noninvasive serum biomarkers capable of such prediction have been rarely exploited. Here, we performed profiling of N-glycosylated proteins in serum from fifteen advanced breast cancer patients (ten patients sensitive to and five patients resistant to NACT) to discover serum biomarkers of chemoresistance using a label-free liquid chromatography-tandem MS method. By performing a series of statistical analyses of the proteomic data, we selected thirteen biomarker candidates and tested their differential serum levels by Western blotting in 13 independent samples (eight patients sensitive to and five patients resistant to NACT). Among the candidates, we then selected the final set of six potential serum biomarkers (AHSG, APOB, C3, C9, CP, and ORM1) whose differential expression was confirmed in the independent samples. Finally, we demonstrated that a multivariate classification model using the six proteins could predict responses to NACT and further predict relapse-free survival of patients. In summary, global N-glycoproteome profile in serum revealed a protein pattern predictive of the responses to NACT, which can be further validated in large clinical studies. PMID:21799047

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

PubMed

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

2015-01-01

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

Seroprevalence of Toxoplasma gondii infection in domestic sheep in Liaoning Province, northeastern China.

PubMed

Yang, Na; Li, Hongkui; He, Jianbin; Mu, Mingyang; Yang, Shuhua

2013-02-01

There is a lack of information regarding the prevalence of Toxoplasma gondii infection in sheep from northeastern China. In the present study, serum samples from 566 domestic sheep were collected from Liaoning Province, northeastern China, to investigate the prevalence of T. gondii antibodies using an indirect hemagglutination antibody test. Twenty-five of 566 samples (4.4%) were seropositive at the cutoff of 1:64 serum dilution. No difference was found among 6 geographic regions (P > 0.05). This is the first report of T. gondii infection in sheep in Liaoning Province.

Comparison of methods of assessing vitamin A status in children.

PubMed

Makdani, D; Sowell, A L; Nelson, J D; Apgar, J; Gunter, E W; Hegar, A; Potts, W; Rao, D; Wilcox, A; Smith, J C

1996-10-01

A study of children (2-8 years; n = 613) in Belize, Central America, was conducted to determine what proportion of the children might be at risk of vitamin A (vit A) deficiency. The data provide an opportunity to compare results of three methods of assessing vit A status in a population which was not severely malnourished. Serum retinyl ester concentrations were also determined; their relevance to one of the tests, the relative dose response (RDR) test, is discussed. The three methods of assessing vit A status were: RDR test, fasting serum retinol concentration, and conjunctival impression cytology (CIC). Retinol-binding protein (RBP), serum retinyl esters and serum zinc concentrations were also determined. Inadequate vit A status was indicated for 17% of subjects by the RDR test (14% cutoff), for 24% by fasting serum retinol concentration (< 0.87 mumol/L), and for 49% by "abnormal" CIC score. Retinyl esters constituted 24% of serum retinoids at the time (5 hours after a retinyl palmitate dose) at which the second blood sample is taken for the RDR test. Regression tree analyses (CART) indicated ethnicity was a predictor of RDR score; ethnicity, stunting and age were predictors of fasting serum retinol concentration; ethnicity and stunting were predictors of 0-hour retinyl ester concentration. The three indices of vit A status did not identify the same individuals nor indicate the same percentage of the population to be at risk for vit A deficiency. Increased concentrations of retinyl esters at 5 hours compared to those at 0 hours suggest that insufficient retinol may have been taken up by the liver at 5 hours to release all accumulated retinol-binding protein (RBP) in deficient individuals; prevalence of vit A deficiency might therefore be underestimated by the RDR test. The selection of ethnicity as a predictor of RDR score and of 0-hour retinol and retinyl ester concentrations suggests that factors other than vit A status affect vit A metabolism and may affect the RDR test.

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

PubMed

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

2017-05-01

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

Simultaneous LC-MS/MS determination of JWH-210, RCS-4, ∆(9)-tetrahydrocannabinol, and their main metabolites in pig and human serum, whole blood, and urine for comparing pharmacokinetic data.

PubMed

Schaefer, Nadine; Kettner, Mattias; Laschke, Matthias W; Schlote, Julia; Peters, Benjamin; Bregel, Dietmar; Menger, Michael D; Maurer, Hans H; Ewald, Andreas H; Schmidt, Peter H

2015-05-01

A series of new synthetic cannabinoids (SC) has been consumed without any toxicological testing. For example, pharmacokinetic data have to be collected from forensic toxicological case work and/or animal studies. To develop a corresponding model for assessing such data, samples of controlled pig studies with two selected SC (JWH-210, RCS-4) and, as reference, ∆(9)-tetrahydrocannabinol (THC) should be analyzed as well as those of human cases. Therefore, a method for determination of JWH-210, RCS-4, THC, and their main metabolites in pig and human serum, whole blood, and urine samples is presented. Specimens were analyzed by liquid-chromatography tandem mass spectrometry and multiple-reaction monitoring with three transitions per compound. Full validation was carried out for the pig specimens and cross-validation for the human specimens concerning precision and bias. For the pig studies, the limits of detection were between 0.05 and 0.50 ng/mL in serum and whole blood and between 0.05 and 1.0 ng/mL in urine, the lower limits of quantification between 0.25 and 1.0 ng/mL in serum and 0.50 and 2.0 ng/mL in whole blood and urine, and the intra- and interday precision values lower than 15% and bias values within ±15%. The applicability was tested with samples taken from a pharmacokinetic pilot study with pigs following intravenous administration of a mixture of 200 μg/kg body mass dose each of JWH-210, RCS-4, and THC. The cross-validation data for human serum, whole blood, and urine showed that this approach should also be suitable for human specimens, e.g., of clinical or forensic cases.

Physicochemical inactivation of Lassa, Ebola, and Marburg viruses and effect on clinical laboratory analyses

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

Mitchell, S.W.; McCormick, J.B.

1984-09-01

Clinical specimens from patients infected with Lassa, Ebola, or Marburg virus may present a serious biohazard to laboratory workers. The authors have examined the effects of heat, alteration of pH, and gamma radiation on these viruses in human blood and on the electrolytes, enzymes, and coagulation factors measured in laboratory tests that are important in the care of an infected patient. Heating serum at 60 degrees C for 1 h reduced high titers of these viruses to noninfectious levels without altering the serum levels of glucose, blood urea nitrogen, and electrolytes. Dilution of blood in 3% acetic acid, diluent formore » a leukocyte count, inactivated all of these viruses. All of the methods tested for viral inactivation markedly altered certain serum proteins, making these methods unsuitable for samples that are to be tested for certain enzyme levels and coagulation factors.« less

Effect of paratuberculosis on culling, milk production, and milk quality in dairy herds.

PubMed

Hendrick, Steven H; Kelton, David F; Leslie, Ken E; Lissemore, Kerry D; Archambault, Marie; Duffield, Todd F

2005-10-15

To determine the effect of paratuberculosis on culling, milk production, and milk quality in infected dairy herds. Cross-sectional study. 689 lactating dairy cows in 9 herds. Milk, blood, and fecal samples were obtained from all cows. Fecal samples were evaluated via mycobacterial culture. Serum samples were tested with a commercially available ELISA for antibodies against Mycobacterium avium subsp paratuberculosis, and preserved milk samples were tested with an ELISA for antibodies against M paratuberculosis. Mixed effect and proportional hazards models were used to determine the effect of paratuberculosis on 305-day milk, fat, and protein production; somatic cell count linear score; and the risk of culling. Cows with positive results of bacteriologic culture of feces and milk ELISA produced less milk, fat, and protein, compared with herdmates with negative results. No difference in 305-day milk or fat production was detected in cows with positive results of serum ELISA, compared with seronegative cows. The 3 survival analyses revealed that cows with positive results of each test were at higher risk of being culled than cows with negative results. Paratuberculosis status, as determined by use of all 3 diagnostic tests, was not associated with milk somatic cell count linear score. Results suggest that for the 9 herds in this study, paratuberculosis significantly decreased milk production and cow longevity.

Within-herd prevalence and clinical incidence distributions of Mycobacterium avium subspecies paratuberculosis infection on dairy herds in Chile.

PubMed

Verdugo, Cristobal; Valdes, Maria Francisca; Salgado, Miguel

2018-06-01

This study aimed to estimate the distributions of the within-herd true prevalence (TP) and the annual clinical incidence proportion (CIp) of Mycobacterium avium subsp. paratuberculosis (MAP) infection in dairy cattle herds in Chile. Forty two commercial herds with antecedents of MAP infection were randomly selected to participate in the study. In small herds (≤30 cows), serum samples were collected from all animals present. Whereas, in larger herds, milk or serum samples were collected from all milking cows with 2 or more parities. Samples were analysed using the Pourquier® ELISA PARATUBERCULOSIS (Insitute Pourquier, France) test. Moreover, a questionnaire gathering information on management practices and the frequency of clinical cases, compatible with paratuberculosis (in the previous 12 months), was applied on the sampling date. A Bayesian latent class analysis was used to obtain TP and clinical incidence posterior distributions. The model adjusts for uncertainty in test sensitivity (serum or milk) and specificity, and prior TP & CIp estimates. A total of 4963 animals were tested, with an average contribution of 124 samples per herd. A mean apparent prevalence of 6.3% (95% confidence interval: 4.0-8.0%) was observed. Model outputs indicated an overall TP posterior distribution, across herds, with a median of 13.1% (95% posterior probability interval (PPI); 3.2-38.1%). A high TP variability was observed between herds. CIp presented a posterior median of 1.1% (95% PPI; 0.2-4.6%). Model results complement information missing from previously conducted epidemiological studies in the sector, and they could be used for further assessment of the disease impact and planning of control programs. Copyright © 2018 Elsevier B.V. All rights reserved.

Suitability of selected free-gas and dissolved-gas sampling containers for carbon isotopic analysis.

PubMed

Eby, P; Gibson, J J; Yi, Y

2015-07-15

Storage trials were conducted for 2 to 3 months using a hydrocarbon and carbon dioxide gas mixture with known carbon isotopic composition to simulate typical hold times for gas samples prior to isotopic analysis. A range of containers (both pierced and unpierced) was periodically sampled to test for δ(13)C isotopic fractionation. Seventeen containers were tested for free-gas storage (20°C, 1 atm pressure) and 7 containers were tested for dissolved-gas storage, the latter prepared by bubbling free gas through tap water until saturated (20°C, 1 atm) and then preserved to avoid biological activity by acidifying to pH 2 with phosphoric acid and stored in the dark at 5°C. Samples were extracted using valves or by piercing septa, and then introduced into an isotope ratio mass spectrometer for compound-specific δ(13)C measurements. For free gas, stainless steel canisters and crimp-top glass serum bottles with butyl septa were most effective at preventing isotopic fractionation (pierced and unpierced), whereas silicone and PTFE-butyl septa allowed significant isotopic fractionation. FlexFoil and Tedlar bags were found to be effective only for storage of up to 1 month. For dissolved gas, crimp-top glass serum bottles with butyl septa were again effective, whereas silicone and PTFE-butyl were not. FlexFoil bags were reliable for up to 2 months. Our results suggest a range of preferred containers as well as several that did not perform very well for isotopic analysis. Overall, the results help establish better QA/QC procedures to avoid isotopic fractionation when storing environmental gas samples. Recommended containers for air transportation include steel canisters and glass serum bottles with butyl septa (pierced and unpierced). Copyright © 2015 John Wiley & Sons, Ltd.

Ratio between serum IL-8 and pepsinogen A/C: a marker for atrophic body gastritis.

PubMed

Sanduleanu, S; Bruïne, A D E; Biemond, I; Stridsberg, M; Jonkers, D; Lundqvist, G; Hameeteman, W; Stockbrügger, R W

2003-02-01

Elevated serum gastrin and a low pepsinogen A/C ratio are well-recognized markers for atrophic body gastritis (ABG). We have shown that the presence of body atrophy is also associated with elevated serum pro-inflammatory cytokines. This study tested the hypothesis that serum cytokines provide additional information to gastrin and pepsinogens in screening for ABG. Two hundred and twenty-six consecutive patients were investigated on referral for upper gastrointestinal endoscopy: 150 were patients with gastro-oesophageal reflux disease, receiving acid inhibitory medication either with proton pump inhibitors (n = 113) or with histamine2-receptor antagonists (n = 37), and 76 were nontreated controls, who had normal endoscopic findings. Gastric mucosal biopsies were sampled for histological examination (Sydney classification). Serum samples were analyzed for gastrin, chromogranin A (CgA), and pepsinogens A and C by RIA, and for the interleukins (IL)-1beta, IL-6, and IL-8 by ELISA. Subjects with ABG had significantly higher serum gastrin (P < 0.01) and serum CgA (P < 0.01) levels and significantly lower pepsinogen A/C ratios (P < 0.001) than those without ABG. Additionally, serum IL-1beta, IL-6 and, especially, IL-8 levels were significantly higher in the subjects with than in those without ABG (P < 0.0001, for all cytokines). To optimize the detection of body atrophy we defined the ABG index: the ratio between the simultaneously measured IL-8 and pepsinogen A/C. The area under the ROC curve for the ABG index was significantly greater than that for serum gastrin and for serum pepsinogen A/C alone (0.91 +/- 0.029 vs. 0.72 +/- 0.042, and vs. 0.83 +/- 0.031, P = 0.018 and P = 0.049). Using the ABG index at a cut-off value of 1.8 pg mL-1, 91% of the cases were classified correctly. The ratio between serum IL-8 and pepsinogen A/C accurately predicts the presence of ABG. We therefore propose the ABG index as a noninvasive screening test for ABG in population-based studies.

Clinical use of serum parvovirus and distemper virus antibody titers for determining revaccination strategies in healthy dogs.

PubMed

Twark, L; Dodds, W J

2000-10-01

To assess whether serum canine parvovirus (CPV) and canine distemper virus (CDV) antibody titers can be used to determine revaccination protocols in healthy dogs. Case series. 1,441 dogs between 6 weeks and 17 years old. CPV and CDV antibody titers in serum samples submitted to a commercial diagnostic laboratory were measured by use of indirect fluorescent antibody (IFA) tests. On the basis of parallel measurements of CPV and CDV serum antibody titers in 61 paired serum samples determined by use of hemagglutination inhibition and serum neutralization methods, respectively, we considered titers > or = 1:5 (IFA test) indicative of an adequate antibody response. Age, breed, and sex were not significantly associated with adequate CPV- or CDV-specific antibody responses. Of 1,441 dogs, 1,370 (95.1%) had adequate and 71 (4.9%) had inadequate antibody responses to CPV, whereas 1,346 of 1,379 (97.6%) dogs had adequate and 33 (2.4%) had inadequate responses to CDV. Vaccination histories were available for 468 dogs (468 for CPV, 457 for CDV). Interval between last vaccination and antibody measurement was 1 to 2 years for the majority (281/468; 60.0%) of dogs and 2 to 7 years for 142 of 468 (30.3%) dogs. Interval was < 1 year in only 45 of 468 (9.6%) dogs. The high prevalence of adequate antibody responses (CPV, 95.1%; CDV, 97.6%) in this large population of dogs suggests that annual revaccination against CPV and CDV may not be necessary.

Effects of oral administration of a commercial activated charcoal suspension on serum osmolality and lactate concentration in the dog.

PubMed

Burkitt, Jamie M; Haskins, Steve C; Aldrich, Janet; Jandrey, Karl E; Rezende, Marlis L; Boyle, Jennifer E

2005-01-01

The purpose of this investigation was to determine the effects of an activated charcoal (AC) suspension containing propylene glycol and glycerol on serum osmolality, osmolal gap, and lactate concentration in dogs. Six healthy adult dogs were administered 4 g/kg AC in a commercially available suspension that contained propylene glycol and glycerol as vehicles. Blood samples were taken before and 1, 4, 6, 8, 12, and 24 hours after the administration of the test suspension. Samples were analyzed for osmolality, blood gases, and concentrations of lactate, sodium, potassium, serum urea nitrogen, and glucose. Osmolal gaps were calculated for each time point. Mean serum osmolality, osmolal gap, and lactate concentration were significantly increased after suspension administration compared to baseline. Serum osmolality increased from 311 mOsm/kg at baseline to 353 mOsm/kg, osmolal gap increased from 5 to 52 mOsm/kg, and lactate concentration increased from 1.9 to 4.5 mmol/L after suspension administration (all P < .01). Three of the 6 dogs vomited between 1 and 3 hours after the administration of the test suspension, and 4 of 6 dogs were lethargic. All dogs drank frequently after AC administration. Commercial AC suspension administered at a clinically relevant dose increases serum osmolality, osmolal gap, and lactate concentration in dogs. These laboratory measures and the clinical signs of vomiting, lethargy, and increased frequency of drinking might complicate the diagnosis or monitoring of some intoxications (such as ethylene glycol) in dogs that have previously received AC suspension containing propylene glycol, glycerol, or both as vehicles.

Discovery and Validation of a Six-Marker Serum Protein Signature for the Diagnosis of Active Pulmonary Tuberculosis.

PubMed

De Groote, Mary A; Sterling, David G; Hraha, Thomas; Russell, Theresa M; Green, Louis S; Wall, Kirsten; Kraemer, Stephan; Ostroff, Rachel; Janjic, Nebojsa; Ochsner, Urs A

2017-10-01

New non-sputum biomarker tests for active tuberculosis (TB) diagnostics are of the highest priority for global TB control. We performed in-depth proteomic analysis using the 4,000-plex SOMAscan assay on 1,470 serum samples from seven countries where TB is endemic. All samples were from patients with symptoms and signs suggestive of active pulmonary TB that were systematically confirmed or ruled out for TB by culture and clinical follow-up. HIV coinfection was present in 34% of samples, and 25% were sputum smear negative. Serum protein biomarkers were identified by stability selection using L1-regularized logistic regression and by Kolmogorov-Smirnov (KS) statistics. A naive Bayes classifier using six host response markers (HR6 model), including SYWC, kallistatin, complement C9, gelsolin, testican-2, and aldolase C, performed well in a training set (area under the sensitivity-specificity curve [AUC] of 0.94) and in a blinded verification set (AUC of 0.92) to distinguish TB and non-TB samples. Differential expression was also highly significant ( P < 10 -20 ) for previously described TB markers, such as IP-10, LBP, FCG3B, and TSP4, and for many novel proteins not previously associated with TB. Proteins with the largest median fold changes were SAA (serum amyloid protein A), NPS-PLA2 (secreted phospholipase A2), and CA6 (carbonic anhydrase 6). Target product profiles (TPPs) for a non-sputum biomarker test to diagnose active TB for treatment initiation (TPP#1) and for a community-based triage or referral test (TPP#2) have been published by the WHO. With 90% sensitivity and 80% specificity, the HR6 model fell short of TPP#1 but reached TPP#2 performance criteria. In conclusion, we identified and validated a six-marker signature for active TB that warrants diagnostic development on a patient-near platform. Copyright © 2017 De Groote et al.

Discovery and Validation of a Six-Marker Serum Protein Signature for the Diagnosis of Active Pulmonary Tuberculosis

PubMed Central

De Groote, Mary A.; Sterling, David G.; Hraha, Thomas; Russell, Theresa M.; Green, Louis S.; Wall, Kirsten; Kraemer, Stephan; Ostroff, Rachel; Janjic, Nebojsa

2017-01-01

ABSTRACT New non-sputum biomarker tests for active tuberculosis (TB) diagnostics are of the highest priority for global TB control. We performed in-depth proteomic analysis using the 4,000-plex SOMAscan assay on 1,470 serum samples from seven countries where TB is endemic. All samples were from patients with symptoms and signs suggestive of active pulmonary TB that were systematically confirmed or ruled out for TB by culture and clinical follow-up. HIV coinfection was present in 34% of samples, and 25% were sputum smear negative. Serum protein biomarkers were identified by stability selection using L1-regularized logistic regression and by Kolmogorov-Smirnov (KS) statistics. A naive Bayes classifier using six host response markers (HR6 model), including SYWC, kallistatin, complement C9, gelsolin, testican-2, and aldolase C, performed well in a training set (area under the sensitivity-specificity curve [AUC] of 0.94) and in a blinded verification set (AUC of 0.92) to distinguish TB and non-TB samples. Differential expression was also highly significant (P < 10−20) for previously described TB markers, such as IP-10, LBP, FCG3B, and TSP4, and for many novel proteins not previously associated with TB. Proteins with the largest median fold changes were SAA (serum amyloid protein A), NPS-PLA2 (secreted phospholipase A2), and CA6 (carbonic anhydrase 6). Target product profiles (TPPs) for a non-sputum biomarker test to diagnose active TB for treatment initiation (TPP#1) and for a community-based triage or referral test (TPP#2) have been published by the WHO. With 90% sensitivity and 80% specificity, the HR6 model fell short of TPP#1 but reached TPP#2 performance criteria. In conclusion, we identified and validated a six-marker signature for active TB that warrants diagnostic development on a patient-near platform. PMID:28794177

Occupational rhinitis and bronchial asthma due to artichoke (Cynara scolymus).

PubMed

Miralles, Juan-Carlos; García-Sells, Javier; Bartolomé, Borja; Negro, José-María

2003-07-01

The artichoke is a perennial horticultural plant that belongs to the Compositae family. To present case studies of 2 vegetable warehouse workers who developed occupational rhinitis and bronchial asthma by sensitization to artichoke. Skin prick tests with common inhalants and foods were performed. Specific IgE to artichoke, Parietaria judaica pollen, and Olea europaea pollen extracts was measured by a specific IgE enzyme immunosorbent assay kit. Molecular mass of the allergens was studied by the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) immunoblotting technique. Patients underwent a nasal challenge test, and one patient provided peak expiratory flow rate (PEFR) measurements in her workplace. In both patients, results of skin prick tests to artichoke were positive. Levels of specific IgE for artichoke were 0.68 kU/L in patient 1 and 2.14 kU/L in patient 2. The protein composition of the artichoke extract, studied by SDS-PAGE, showed that most bands ranged from 30 to 14 kDa. The IgE-binding bands with the serum samples of patient 1 showed apparent molecular masses of 56, 48, 38, 31, 27, 25, 16, and 15 kDa; however, the serum samples of patient 2 showed IgE bands of 21 and 19 kDa. Western blotting of artichoke extract showed a complete inhibition of IgE-binding bands when serum samples were preincubated with P. judaica pollen extract. Nasal challenge with artichoke extract triggered a peak nasal inspiratory flow decrease of 81% and 85% in patient 1 and patient 2, respectively. Finally, patient 1 recorded a PEFR decrease of up to 36% after exposure to artichoke in her workplace. SDS-PAGE immunoblotting inhibition performed for the artichoke extract showed a total disappearance of the specific IgE binding bands when serum samples were previously incubated with P. judaica pollen extract, thus establishing the existence of a serologic cross-reactivity between artichoke and P. judaica pollen.

Seroprevalence of Chikungunya in Southern Odisha

PubMed Central

Mohanty, Indrani; Dash, Muktikesh; Sahu, Susmita; Narasimham, M.V.; Panda, Pritilata; Padhi, Sanghamitra

2013-01-01

Background: The emergence of chikungunya (CHIK) infection was observed in Odisha, India in 2006. Thereafter many cases with symptoms suggestive of CHIK were reported from different districts of Southern-Odisha. This study was aimed to know the seroprevalence, clinical presentations and seasonal trends of CHIK infection in this region. Materials and Methods: This study was conducted in a tertiary hospital of this region. Serum samples received in the Department of Microbiology from various districts of Southern-Odisha from April 2011 to March 2012 were included in the study. The samples were tested for CHIK and dengue Immunoglobin M (IgM) antibodies by enzyme-linked immunosorbent assay and malaria parasite by immunochromatographic test (ICT) method. Results: Out of the 678 serum samples tested, 174 were positive for CHIK, 15 for dengue and two samples were positive for both CHIK and dengue IgM antibodies. The most affected age group was 16-45 years. Females were more affected than males. Conclusion: The seroprevalence of CHIK among the suspected cases was 25.7%. Co-infection with CHIK and dengue was found to be 1.15%. The infection had spread to new areas during this outbreak. PMID:24479040

Improving serum calcium test ordering according to a decision algorithm.

PubMed

Faria, Daniel K; Taniguchi, Leandro U; Fonseca, Luiz A M; Ferreira-Junior, Mario; Aguiar, Francisco J B; Lichtenstein, Arnaldo; Sumita, Nairo M; Duarte, Alberto J S; Sales, Maria M

2018-05-18

To detect differences in the pattern of serum calcium tests ordering before and after the implementation of a decision algorithm. We studied patients admitted to an internal medicine ward of a university hospital on April 2013 and April 2016. Patients were classified as critical or non-critical on the day when each test was performed. Adequacy of ordering was defined according to adherence to a decision algorithm implemented in 2014. Total and ionised calcium tests per patient-day of hospitalisation significantly decreased after the algorithm implementation; and duplication of tests (total and ionised calcium measured in the same blood sample) was reduced by 49%. Overall adequacy of ionised calcium determinations increased by 23% (P=0.0001) due to the increase in the adequacy of ionised calcium ordering in non-critical conditions. A decision algorithm can be a useful educational tool to improve adequacy of the process of ordering serum calcium tests. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Evaluation of performance of bacterial culture of feces and serum ELISA across stages of Johne's disease in cattle using a Bayesian latent class model.

PubMed

Espejo, L A; Zagmutt, F J; Groenendaal, H; Muñoz-Zanzi, C; Wells, S J

2015-11-01

The objective of this study was to evaluate the performance of bacterial culture of feces and serum ELISA to correctly identify cows with Mycobacterium avium ssp. paratuberculosis (MAP) at heavy, light, and non-fecal-shedding levels. A total of 29,785 parallel test results from bacterial culture of feces and serum ELISA were collected from 17 dairy herds in Minnesota, Pennsylvania, and Colorado. Samples were obtained from adult cows from dairy herds enrolled for up to 10 yr in the National Johne's Disease Demonstration Herd Project. A Bayesian latent class model was fitted to estimate the probabilities that bacterial culture of feces (using 72-h sedimentation or 30-min centrifugation methods) and serum ELISA results correctly identified cows as high positive, low positive, or negative given that cows were heavy, light, and non-shedders, respectively. The model assumed that no gold standard test was available and conditional independency existed between diagnostic tests. The estimated conditional probabilities that bacterial culture of feces correctly identified heavy shedders, light shedders, and non-shedders were 70.9, 32.0, and 98.5%, respectively. The same values for the serum ELISA were 60.6, 18.7, and 99.5%, respectively. Differences in diagnostic test performance were observed among states. These results improve the interpretation of results from bacterial culture of feces and serum ELISA for detection of MAP and MAP antibody (respectively), which can support on-farm infection control decisions and can be used to evaluate disease-testing strategies, taking into account the accuracy of these tests. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Cells and biofluids analyzed in aqueous environment by infrared spectroscopy

NASA Astrophysics Data System (ADS)

Naumann, D.; Lasch, P.; Fabian, H.

2006-02-01

Infrared transmission/absorption measurements of cells and biofluids in water are restricted to very short optical pathlengths. When the amide I and amide II bands of protein constituents have to be analysed, path-lengths of less than 8 μm are necessary. Infrared spectra of cancer cells were collected from physiological buffer solutions utilizing custom-made mid-infrared compatible IR-cuvettes. The technology permitted to obtain cell-type specific spectral signatures and probe biochemical changes induced by varying temperatures or cell-drug interaction. Optical path-lengths of 8-30 μm were used on a set of microbial test strains to evaluate, whether the methodology can also be used to discriminate and identify micro-organisms. A semi-automatic methodology was developed for the analysis of liquid serum samples, which combines simple sample handling with high sample throughput and extreme measurement reproducibility. The applicability of this infrared technology to the analysis of liquid serum samples from cattle and human beings suffering from various acute viral or bacterial infections was explored testing the interrelationship between α-helical and β-sheet specific spectral signatures in the amide I band contour and total albumin and globulin content in serum. The technical details, advantages, and limitations of the new technology are described in the context of developing a routine, IR-based biodiagnostic technique for biofluids and biological cells.

Detection of autoantibodies against thyroid peroxidase in serum samples of hypothyroid dogs.

PubMed

Skopek, Evelyn; Patzl, Martina; Nachreiner, Raymond F

2006-05-01

To establish a sensitive test for the detection of autoantibodies against thyroid peroxidase (TPO) in canine serum samples. 365 serum samples from dogs with hypothyroidism as determined on the basis of serum concentrations of total and free triiodothyronine (T3), total and free thyroxine (T4), and thyroid-stimulating hormone, of which 195 (53%) had positive results for at least 1 of 3 thyroid autoantibodies (against thyroglobulin [Tg], T4, or T3) and serum samples from 28 healthy dogs (control samples). TPO was purified from canine thyroid glands by extraction with detergents, ultracentrifugation, and precipitation with ammonium sulfate. Screening for anti-TPO autoantibodies in canine sera was performed by use of an immunoblot assay. Thyroid extract containing TPO was separated electrophoretically, blotted, and probed with canine sera. Alkaline phosphatase-conjugated rabbit anti-dog IgG was used for detection of bound antibodies. TPO bands were observed at 110, 100, and 40 kd. Anti-TPO autoantibodies against the 40-kd fragment were detected in 33 (17%) sera of dogs with positive results for anti-Tg, anti-T4, or anti-T3 autoantibodies but not in sera of hypothyroid dogs without these autoantibodies or in sera of healthy dogs. The immunoblot assay was a sensitive and specific method for the detection of autoantibodies because it also provided information about the antigen. Anti-TPO autoantibodies were clearly detected in a fraction of hypothyroid dogs. The value of anti-TPO autoantibodies for use in early diagnosis of animals with thyroid gland diseases should be evaluated in additional studies.

Serology of Typhoid Fever in an Area of Endemicity and Its Relevance to Diagnosis

PubMed Central

House, Deborah; Wain, John; Ho, Vo A.; Diep, To S.; Chinh, Nguyen T.; Bay, Phan V.; Vinh, Ha; Duc, Minh; Parry, Christopher M.; Dougan, Gordon; White, Nicholas J.; Hien, Tran Tinh; Farrar, Jeremy J.

2001-01-01

Currently, the laboratory diagnosis of typhoid fever is dependent upon either the isolation of Salmonella enterica subsp. enterica serotype Typhi from a clinical sample or the detection of raised titers of agglutinating serum antibodies against the lipopolysaccharide (LPS) (O) or flagellum (H) antigens of serotype Typhi (the Widal test). In this study, the serum antibody responses to the LPS and flagellum antigens of serotype Typhi were investigated with individuals from a region of Vietnam in which typhoid is endemic, and their usefulness for the diagnosis of typhoid fever was evaluated. The antibody responses to both antigens were highly variable among individuals infected with serotype Typhi, and elevated antibody titers were also detected in a high proportion of serum samples from healthy subjects from the community. In-house enzyme-linked immunosorbent assays (ELISAs) for the detection of specific classes of anti-LPS and antiflagellum antibodies were compared with other serologically based tests for the diagnosis of typhoid fever (Widal TO and TH, anti-serotype Typhi immunoglobulin M [IgM] dipstick, and IDeaL TUBEX). At a specificity of ≥0.93, the sensitivities of the different tests were 0.75, 0.55, and 0.52 for the anti-LPS IgM, IgG, and IgA ELISAs, respectively; 0.28 for the antiflagellum IgG ELISA; 0.47 and 0.32 for the Widal TO and TH tests, respectively; and 0.77 for the anti-serotype Typhi IgM dipstick assay. The specificity of the IDeaL TUBEX was below 0.90 (sensitivity, 0.87; specificity, 0.76). The serological assays based on the detection of IgM antibodies against either serotype Typhi LPS (ELISA) or whole bacteria (dipstick) had a significantly higher sensitivity than the Widal TO test when used with a single acute-phase serum sample (P ≤ 0.007). These tests could be of use for the diagnosis of typhoid fever in patients who have clinical typhoid fever but are culture negative or in regions where bacterial culturing facilities are not available. PMID:11230418

Multicommuted flow system for the determination of glucose in animal blood serum exploiting enzymatic reaction and chemiluminescence detection

PubMed Central

Pires, Cherrine K.; Martelli, Patrícia B.; Lima, José L. F. C.; Saraiva, Maria Lúcia M. F. S.

2003-01-01

An automatic flow procedure based on multicommutation dedicated for the determination of glucose in animal blood serum using glucose oxidase with chemiluminescence detection is described. The flow manifold consisted of a set of three-way solenoid valves assembled to implement multicommutation. A microcomputer furnished with an electronic interface and software written in Quick BASIC 4.5 controlled the manifold and performed data acquisition. Glucose oxidase was immobilized on porous silica beads (glass aminopropyl) and packed in a minicolumn (15 × 5 mm). The procedure was based on the enzymatic degradation of glucose, producing hydrogen peroxide, which oxidized luminol in the presence of hexacyanoferrate(III), causing the chemiluminescence. The system was tested by analysing a set of serum animal samples without previous treatment. Results were in agreement with those obtained with the conventional method (LABTEST Kit) at the 95% confidence level. The detection limit and variation coefficient were estimated as 12.0 mg l−1 (99.7% confidence level) and 3.5% (n = 20), respectively. The sampling rate was about 60 determinations h−1 with sample concentrations ranging from 50 to 600 mg l−1 glucose. The consumptions of serum sample, hexacyanoferrate(III) and luminol were 46 μl, 10.0 mg and 0.2 mg/determination, respectively. PMID:18924619

An occupational exposure assessment of a perfluorooctanesulfonyl fluoride production site: biomonitoring.

PubMed

Olsen, Geary W; Logan, Perry W; Hansen, Kristen J; Simpson, Cathy A; Burris, Jean M; Burlew, Michele M; Vorarath, Phanasouk P; Venkateswarlu, Pothapragada; Schumpert, John C; Mandel, Jeffrey H

2003-01-01

This investigation randomly sampled a fluorochemical manufacturing employee population to determine the distribution of serum fluorochemical levels according to employees' jobs and work areas. Previous analyses of medical surveillance data have not shown significant associations between fluorochemical production employees' clinical chemistry and hematology tests and their serum PFOS and perfluorooctanoate (PFOA, C(7)F(15)COO(-)) concentrations, but may have been subject to nonparticipation bias. A random sample of the on-site film plant employee population, where fluorochemicals are not produced, determined their serum concentrations also. Of the 232 employees randomly selected for serum sampling, 186 (80%) employees participated (n=126 chemical plant; n=60 film plant). Sera samples were extracted using an ion-pairing extraction procedure and were quantitatively analyzed for seven fluorochemicals using high-pressure liquid chromatography electrospray tandem mass spectrometry methods. Geometric means (in parts per million) and 95% confidence intervals (in parentheses) of the random sample of 126 chemical plant employees were: PFOS 0.941 (0.787-1.126); PFOA 0.899 (0.722-1.120); perfluorohexanesulfonate 0.180 (0.145-0.223); N-ethyl perfluorooctanesulfonamidoacetate 0.008 (0.006-0.011); N-methyl perfluorooctanesulfonamidoacetate 0.081 (0.067-0.098); perfluorooctanesulfonamide 0.013 (0.009-0.018); and perfluorooctanesulfonamidoacetate 0.022 (0.018-0.029). These geometric means were approximately one order of magnitude higher than those observed for the film plant employees.

A cross-sectional analysis of dioxins and health effects in municipal and private waste incinerator workers in Japan

PubMed Central

YAMAMOTO, Kenya; KUDO, Mitsuhiro; ARITO, Heihachiro; OGAWA, Yasutaka; TAKATA, Tsutomu

2015-01-01

This cross-sectional study was intended to examine health effects of 678 male workers employed during an 8-yr period from 2000 to 2007 at 36 municipal and private waste incineration plants in Japan. Blood samples were obtained for analysis of concentrations of dioxins including coplanar polychlorinated biphenyls (coplanar PCBs) and evaluation of health effects. Health effects including diabetes were surveyed via a physician’s interview or clinical data from blood samples. There was a certain difference in serum concentrations of polychlorinated dibenzofurans (PCDFs) between the incinerator workers and Japanese general population, although no differences in the concentrations of total dioxins or polychlorinated dibenzo-p-dioxins (PCDDs) were found between the two groups. A few positive correlations between serum levels of PCDDs and PCDFs and the results of laboratory and physiological tests were found, but coplanar PCBs showed significant relations with 14 parameters of the tests. The background serum levels of PCDDs, PCDFs and total dioxins were significantly associated with the prevalence of diabetes. No essential differences in serum concentrations of total dioxins and in prevalence of diabetes between our subjects and the general population suggested that the incinerator workers were marginally exposed to dioxins in the workplace without any recognizable adverse health effects. PMID:26212412

Serosurvey of Crimean-Congo Hemorrhagic Fever Virus in Cattle, Mali, West Africa.

PubMed

Maiga, Ousmane; Sas, Miriam Andrada; Rosenke, Kyle; Kamissoko, Badian; Mertens, Marc; Sogoba, Nafomon; Traore, Abdallah; Sangare, Modibo; Niang, Mamadou; Schwan, Tom G; Maiga, Hamidou Moussa; Traore, Sekou F; Feldmann, Heinz; Safronetz, David; Groschup, Martin H

2017-06-01

AbstractCrimean-Congo hemorrhagic fever is a tick-borne disease caused by the arbovirus Crimean-Congo hemorrhagic fever virus (CCHFV, family Bunyaviridae, genus Nairovirus ). CCHFV can cause a severe hemorrhagic fever with high-case fatality rates in humans. CCHFV has a wide geographic range and has been described in around 30 countries in the Middle East, Asia, Europe, and Africa including Mali and neighboring countries. To date, little is known about the prevalence rates of CCHFV in Mali. Here, using banked bovine serum samples from across the country, we describe the results of a seroepidemiological study for CCHFV aimed at identifying regions of circulation in Mali. In total, 1,074 serum samples were tested by a modified in-house CCHFV-IgG-enzyme-linked immunosorbent assay (ELISA) with confirmatory testing by commercial ELISA and immunofluorescence assay. Overall, 66% of samples tested were positive for CCHFV-specific IgG antibodies. Regional seroprevalence rates ranged from 15% to 95% and seemed to correlate with cattle density. Our results demonstrate that CCHFV prevalence is high in many regions in Mali and suggest that CCHFV surveillance should be established.

Automation of serum (1→3)-beta-D-glucan testing allows reliable and rapid discrimination of patients with and without candidemia.

PubMed

Prüller, Florian; Wagner, Jasmin; Raggam, Reinhard B; Hoenigl, Martin; Kessler, Harald H; Truschnig-Wilders, Martie; Krause, Robert

2014-07-01

Testing for (1→3)-beta-D-glucan (BDG) is used for detection of invasive fungal infection. However, current assays lack automation and the ability to conduct rapid single-sample testing. The Fungitell assay was adopted for automation and evaluated using clinical samples from patients with culture-proven candidemia and from culture-negative controls in duplicate. A comparison with the standard assay protocol was made in order to establish analytical specifications. With the automated protocol, the analytical measuring range was 8-2500 pg/ml of BDG, and precision testing resulted in coefficients of variation that ranged from 3.0% to 5.5%. Samples from 15 patients with culture-proven candidemia and 94 culture-negative samples were evaluated. All culture-proven samples showed BDG values >80 pg/ml (mean 1247 pg/ml; range, 116-2990 pg/ml), which were considered positive. Of the 94 culture-negative samples, 92 had BDG values <60 pg/ml (mean, 28 pg/ml), which were considered to be negative, and 2 samples were false-positive (≥80 pg/ml; up to 124 pg/ml). Results could be obtained within 45 min and showed excellent agreement with results obtained with the standard assay protocol. The automated Fungitell assay proved to be reliable and rapid for diagnosis of candidemia. It was demonstrated to be feasible and cost efficient for both single-sample and large-scale testing of serum BDG. Its 1-h time-to-result will allow better support for clinicians in the management of antifungal therapy. © The Author 2014. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Colony-level assessment of Brucella and Leptospira in the Guadalupe fur seal, Isla Guadalupe, Mexico.

PubMed

Ziehl-Quirós, E Carolina; García-Aguilar, María C; Mellink, Eric

2017-01-24

The relatively small population size and restricted distribution of the Guadalupe fur seal Arctocephalus townsendi could make it highly vulnerable to infectious diseases. We performed a colony-level assessment in this species of the prevalence and presence of Brucella spp. and Leptospira spp., pathogenic bacteria that have been reported in several pinniped species worldwide. Forty-six serum samples were collected in 2014 from pups at Isla Guadalupe, the only place where the species effectively reproduces. Samples were tested for Brucella using 3 consecutive serological tests, and for Leptospira using the microscopic agglutination test. For each bacterium, a Bayesian approach was used to estimate prevalence to exposure, and an epidemiological model was used to test the null hypothesis that the bacterium was present in the colony. No serum sample tested positive for Brucella, and the statistical analyses concluded that the colony was bacterium-free with a 96.3% confidence level. However, a Brucella surveillance program would be highly recommendable. Twelve samples were positive (titers 1:50) to 1 or more serovars of Leptospira. The prevalence was calculated at 27.1% (95% credible interval: 15.6-40.3%), and the posterior analyses indicated that the colony was not Leptospira-free with a 100% confidence level. Serovars Icterohaemorrhagiae, Canicola, and Bratislava were detected, but only further research can unveil whether they affect the fur seal population.

[Isolation and immunology identification of spermagglutinating antibodies from human serum].

PubMed

Cibulka, J; Ulcova-Gallová, Z; Balvín, M; Bibková, K; Micanová, Z

2009-06-01

Isolation of spermagglutinating antibodies and their assesment. Retrospective study. Special consulting for reptoduction immunology, Department of Obstetrics and Gynecology, Charles University and Faculty Hospital, Plzen. Fractionation of serum samples by liquid exclusion chromatography, examination of full sera and their chromatographic fractions by Friberg teste (Tray Agglutination Test--TAT), indirect antiimmunoglobulin reaction test (i-MAR test) and by supplementar radial immunodiffusiona (RID). Isolation of spermagglutinating fractions of antisperm antibodies positive sera preserved spermagglutinating aktivity and confirmed great spermagglutinating potential of IgM. According to assesment of the presence of IgG and IgG we reported possible states of immunisation: actual immunisation with IgM activity, perpetual stimulation (IgG and IgM) and, finaly, anamnestic titres in IgG. These findings can help us to choose an optimal way of treatment. Excluding gel chromatography is suitable method for serum proteins fractionation, but not their identification--presence of antisperm antibodies does not affect the chromatographic spectrum, nor the RID patterns.

24-hour urinary aldosterone excretion test

MedlinePlus

Aldosterone - urine; Addison disease - urine aldosterone; Cirrhosis - serum aldosterone ... A 24-hour urine sample is needed. You will need to collect your urine over 24 hours . Your health care provider will tell ...

A Primary Investigation on Serum CTX-II Changes in Patients Infected with Brucellosis in Qinghai Plateau, China.

PubMed

Zhao, Zhi Jun; Li, Qiang; Zhou, Xin; Ma, Li; Xu, Li Qing; Yang, Pei Zhen; Meng, Xian Ya; Yu, Hui Zhen; Xu, Xiao Qing; Cao, Jian Ying

2016-03-01

Brucellosis is one of the most widespread zoonotic diseases, with the most frequent complication being osteoarticular changes. The aim of this study was to assess the changes of C-terminal telopeptide of type II collagen (CTX-II) in patients infected with brucellosis. A total of 84 brucellosis patients and 43 volunteers were selected and divided into brucellosis vs. control groups. Serum samples were subjected to serological tests for brucellosis, and CTX-II levels in all samples were measured simultaneously with ELISA. The results showed that serum CTX-II levels in human brucellosis were higher than those of healthy controls, without a statistically significant difference, but serum CTX-II levels in male patients were significantly higher than those of female patients (P<0.05). This finding could indicate the biological changes in the cartilage and bone in human brucellosis. Copyright © 2016 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

Physical Performance and Serum 25(OH)vitamin D Status in Community Dwelling Old Mobility Limited Adults: A Cross-Sectional Study.

PubMed

von Berens, Å; Cederholm, T; Fielding, R A; Gustafsson, T; Kirn, D; Laussen, J; Nydahl, M; Travison, T G; Reid, K; Koochek, A

2018-01-01

To examine the potential association between serum 25(OH) vitamin D and the performance on the Short Physical Performance Battery (SPPB) including the sub-components; five repeated chair stands test, 4 meters walk test and balance in older mobility-limited community-dwelling men and women. A cross sectional study was performed in American and Swedish subjects who were examined for potential participation in a combined exercise and nutrition intervention trial. Logistic regression analysis and linear regression analyses were performed to evaluate the association for 25(OH)D with the overall score on the SBBP, chair stand, gait speed and balance. Community-dwelling (mean age 77.6 ± 5.3 years) mobility limited American (n=494) and Swedish (n=116) females (59%) and males. The SPPB (0-12 points) includes chair stand (s), gait speed (m/s) and a balance test. Mobility limitation i.e., SPPB score ≤ 9 was an inclusion criterion. A blood sample was obtained to measure serum 25(OH)vitamin D concentrations. No clear association of 25(OH)D with SPPB scores was detected either when 25(OH)D was assessed as a continuous variable or when categorized according to serum concentrations of <50, 50-75 or <75 nmol/L. However, when analyzing the relationship between 25(OH)D and seconds to perform the chair stands, a significant quadratic relationship was observed. Thus, at serum levels of 25(OH)D above 74 nmol/L, higher concentrations appeared to be advantageous for the chair stand test, whereas for serum levels below 74 nmol/L this association was not observed. This cross- sectional study lacked clear association between serum 25(OH)D and physical performance in mobility limited adults. A potentially interesting observation was that at higher serum levels of 25(OH)D a better performance on the chair stand test was indicated.

Brominated flame retardant (BFRs) and Dechlorane Plus (DP) in paired human serum and segmented hair.

PubMed

Qiao, Lin; Zheng, Xiao-Bo; Yan, Xiao; Wang, Mei-Huang; Zheng, Jing; Chen, She-Jun; Yang, Zhong-Yi; Mai, Bi-Xian

2018-01-01

Brominated flame retardants (BFRs) and Dechlorane Plus (DP) were measured in both human hair and paired serum samples from a cohort of university students in South China. Segmental analysis was conducted to explore gender difference and the relationships between the hair and serum. The concentrations of total PBDEs in the hair and serum samples were in a range of 0.28-34.1ng/g dry weight (dw) and 0.16-156ng/g lipid weight (lw), respectively. Concentrations of ∑DPs (sum of the syn-DP and anti-DP isomers) in all hair samples ranged from nd-5.45ng/g dry weight. Concentrations of most PBDEs and decabromodiphenylethane (DBDPE) in distal segments (5-10cm from the scalp) were higher than those in the proximal segments (0-5cm from the scalp) (t-test, p < 0.05), which could be due to the longer exposure time of distal segments. The proximal segments exhibited a unique congener profile, more close to that in the serum rather than the distal segments of hair. An obvious gender difference was found in the levels of ∑PBDEs using integrated hair samples, while the difference disappeared when considering alone the proximal segments of hair (0-5cm from scalp) for both genders. This paper provides supplement to the current knowledge on sources of BFRs and DPs in hair and declares the importance of segmental analysis. Copyright © 2017 Elsevier Inc. All rights reserved.

Comparison of Serum Autoantibodies to Desmogleins I, III in Patients with Oral Lichen Planus and Healthy Controls.

PubMed

Gholizadeh, Narges; Khoini Poorfar, Hossein; TaghaviZenouz, Ali; Vatandoost, Masoumeh; Mehdipour, Masoumeh

2015-01-01

Lichen planus is a mucocutaneous disease which is relatively common and in 30-70% of patients, mucosal lesions can be seen and known as a precancerous lesion but its etiology is not completely understood. Desmogleins I and III are the main desmosomal transmembrane proteins. These proteins have been identified as the autoantigen of the autoimmune disease. The aim of this study was evaluation of serum autoantibodies against desmogleins Ι, ΙΙΙ in oral lichen planus . We attempted to determine the etiology of this disease with evaluation of these serum factors. Thirty-five patients with oral lichen planus and 35 healthy controls were recruited and tested for serum autoantibodies against desmogleins Ι, ΙΙΙ and indirect immunofluorescence also performed. Data were analyzed by statistical-analytical methods (Independent sample t -test) with using the SPSS.15 software. Serum autoantibody against desmoglein Ι had no significant difference in the two groups ( P =0.31 ) but significant increase in serum autoantibody to desmoglein ΙΙΙ was found in patients with oral lichen planus ( P =0.00) . It seems that autoantibody against desmoglein III has a significant role in the pathogenesis of oral lichen planus.

Long-term stability of glucose: glycolysis inhibitor vs. gel barrier tubes.

PubMed

Winter, Theresa; Hannemann, Anke; Suchsland, Juliane; Nauck, Matthias; Petersmann, Astrid

2018-03-12

Measuring the glucose concentration in whole blood samples is critical due to unsatisfactory glycolysis inhibition. Previous studies showed that Terumo tubes were superior, but they were taken off the European market in 2016 and alternatives were required. This initiated the present evaluation of glucose stability in five available tube types. Venous blood samples were collected from 61 healthy volunteers to test tubes supplied by Terumo (two sets), Greiner FC-Mix, BD FX-Mixture and BD serum. After sampling, the contents were thoroughly mixed and centrifuged within an hour. The glucose concentrations were determined and the samples resuspended except for BD serum tubes (gel barrier). The first 30 samples were stored at room temperature and the remaining 31 at 4°C. After 24, 48, 72 and 96 h, all tubes were (re)centrifuged, and glucose concentration measurements were repeated. Changes in glucose concentrations over time differed significantly between the investigated tube types and to a certain extent between the two storing conditions. Glycolysis was most evident in the BD FX-mixture tubes. Good glucose stability was observed in samples retrieved form BD serum and Greiner tubes. The stability in both Terumo tubes was comparable to that in other studies. Although Greiner and both Terumo tubes are supposed to contain the same glycolysis inhibitor, glucose stability differed between these tubes. We showed that Greiner is an acceptable alternative to Terumo and that glucose in serum that was rapidly separated from corpuscles by a gel barrier is stable for an extended time.

Detection of Japanese Encephalitis Virus RNA in Human Throat Samples in Laos - A Pilot study.

PubMed

Bharucha, Tehmina; Sengvilaipaseuth, Onanong; Seephonelee, Malee; Vongsouvath, Malavanh; Vongsouvath, Manivanh; Rattanavong, Sayaphet; Piorkowski, Géraldine; Lecuit, Marc; Gorman, Christopher; Pommier, Jean-David; Newton, Paul N; de Lamballerie, Xavier; Dubot-Pérès, Audrey

2018-05-22

Japanese encephalitis virus (JEV) is the most commonly identified cause of acute encephalitis syndrome (AES) in Asia. The WHO recommended test is anti-JEV IgM-antibody-capture-enzyme-linked-immunosorbent-assay (JEV MAC-ELISA). However, data suggest this has low positive predictive value, with false positives related to other Flavivirus infections and vaccination. JEV RT-PCR in cerebrospinal fluid (CSF) and/or serum is highly specific, but is rarely positive; 0-25% of patients that fulfil the WHO definition of JE (clinical Acute Encephalitis Syndrome (AES) and JEV MAC-ELISA positive). Testing other body fluids by JEV RT-qPCR may improve the diagnosis. As a pilot study thirty patients admitted to Mahosot Hospital 2014-2017, recruited to the South-East-Asia-Encephalitis study, were tested by JEV MAC-ELISA and two JEV real-time RT-PCR (RT-qPCR) assays (NS2A and NS3). Eleven (36.7%) were JEV MAC-ELISA positive. Available CSF and serum samples of these patients were JEV RT-qPCR negative but 2 (7%) had JEV RNA detected in their throat swabs. JEV RNA was confirmed by re-testing, and sequencing of RT-qPCR products. As the first apparent report of JEV RNA detection in human throat samples, the provides new perspectives on human JEV infection, potentially informing improving JEV detection. We suggest that testing patients' throat swabs for JEV RNA is performed, in combination with molecular and serological CSF and serum investigations, on a larger scale to investigate the epidemiology of the presence of JEV in human throats. Throat swabs are an easy and non-invasive tool that could be rolled out to a wider population to improve knowledge of JEV molecular epidemiology.

Evaluation of three high abundance protein depletion kits for umbilical cord serum proteomics

PubMed Central

2011-01-01

Background High abundance protein depletion is a major challenge in the study of serum/plasma proteomics. Prior to this study, most commercially available kits for depletion of highly abundant proteins had only been tested and evaluated in adult serum/plasma, while the depletion efficiency on umbilical cord serum/plasma had not been clarified. Structural differences between some adult and fetal proteins (such as albumin) make it likely that depletion approaches for adult and umbilical cord serum/plasma will be variable. Therefore, the primary purposes of the present study are to investigate the efficiencies of several commonly-used commercial kits during high abundance protein depletion from umbilical cord serum and to determine which kit yields the most effective and reproducible results for further proteomics research on umbilical cord serum. Results The immunoaffinity based kits (PROTIA-Sigma and 5185-Agilent) displayed higher depletion efficiency than the immobilized dye based kit (PROTBA-Sigma) in umbilical cord serum samples. Both the PROTIA-Sigma and 5185-Agilent kit maintained high depletion efficiency when used three consecutive times. Depletion by the PROTIA-Sigma Kit improved 2DE gel quality by reducing smeared bands produced by the presence of high abundance proteins and increasing the intensity of other protein spots. During image analysis using the identical detection parameters, 411 ± 18 spots were detected in crude serum gels, while 757 ± 43 spots were detected in depleted serum gels. Eight spots unique to depleted serum gels were identified by MALDI- TOF/TOF MS, seven of which were low abundance proteins. Conclusions The immunoaffinity based kits exceeded the immobilized dye based kit in high abundance protein depletion of umbilical cord serum samples and dramatically improved 2DE gel quality for detection of trace biomarkers. PMID:21554704

Identification of novel serum peptide biomarkers for high-altitude adaptation: a comparative approach

PubMed Central

Yang, Juan; Li, Wenhua; Liu, Siyuan; Yuan, Dongya; Guo, Yijiao; Jia, Cheng; Song, Tusheng; Huang, Chen

2016-01-01

We aimed to identify serum biomarkers for screening individuals who could adapt to high-altitude hypoxia at sea level. HHA (high-altitude hypoxia acclimated; n = 48) and HHI (high-altitude hypoxia illness; n = 48) groups were distinguished at high altitude, routine blood tests were performed for both groups at high altitude and at sea level. Serum biomarkers were identified by comparing serum peptidome profiling between HHI and HHA groups collected at sea level. Routine blood tests revealed the concentration of hemoglobin and red blood cells were significantly higher in HHI than in HHA at high altitude. Serum peptidome profiling showed that ten significantly differentially expressed peaks between HHA and HHI at sea level. Three potential serum peptide peaks (m/z values: 1061.91, 1088.33, 4057.63) were further sequence identified as regions of the inter-α trypsin inhibitor heavy chain H4 fragment (ITIH4 347–356), regions of the inter-α trypsin inhibitor heavy chain H1 fragment (ITIH1 205–214), and isoform 1 of fibrinogen α chain precursor (FGA 588–624). Expression of their full proteins was also tested by ELISA in HHA and HHI samples collected at sea level. Our study provided a novel approach for identifying potential biomarkers for screening people at sea level who can adapt to high altitudes. PMID:27150491

Coxiella burnetii Circulation in a Naturally Infected Flock of Sheep: Individual Follow-Up of Antibodies in Serum and Milk

PubMed Central

Joulié, A.; Rousset, E.; Gasqui, P.; Lepetitcolin, E.; Leblond, A.; Sidi-Boumedine, K.

2017-01-01

ABSTRACT The control of Q fever, a zoonotic disease caused by the Coxiella burnetii bacterium, remains a scientific challenge. Domestic ruminants are considered the main reservoir, shedding C. burnetii essentially through parturition products during abortion or birth. Sheep are particularly frequently associated with human outbreaks, but there are insufficient field data to fully understand disease dynamics and to instigate efficient control measures. A longitudinal follow-up study of a naturally infected sheep flock was performed (i) to investigate relationships between seropositivity and bacterial shedding in the vaginal mucus, (ii) to describe the kinetics of antibodies, including responses to vaccination, (iii) to monitor maternal antibodies in ewe lambs, and (iv) to compare serological results for milk and serum samples. For 8 months, we collected blood samples every 3 weeks from 11 aborting and 26 nonaborting dairy ewes, 20 nonaborting suckler ewes, and 9 ewe lambs. Individual milk samples were also obtained from lactating females. All serum and milk samples were tested by enzyme-linked immunosorbent assay (ELISA), whereas vaginal swabs were tested by quantitative PCR. We found that some dairy females did not seroconvert despite shedding C. burnetii in their vaginal mucus. Overall, antibody levels in adult females were found to remain stable over time, with exceptions during the mating and lambing periods. Maternal antibodies decreased during the first month after birth. Interestingly, antibody levels in milk were correlated with those in serum. This study provides valuable field data that will help improve Q fever surveillance and within-flock management measures. IMPORTANCE Field data are necessary to improve the surveillance, diagnosis, and sanitary management of Q fever in livestock. Here, we provide extensive serological data obtained from serum and milk samples from infected and vaccinated ewes belonging to a naturally infected flock of sheep. We show that antibody levels are stable over time and seropositivity and vaginal shedding are not clearly correlated, whereas antibody levels in milk are strongly correlated with those in serum. Accordingly, we find that antibody levels in bulk tank milk are consistent with the variations observed in the serum of dairy females over time. We report the existence of maternal antibody transmission to ewe lambs and we show that the presence of maternal antibodies at birth does not prevent the development of a serological response to vaccination at the age of 4 months. Finally, we report that adult ewes generally seroconvert after vaccination, including during pregnancy. PMID:28455328

Coxiella burnetii Circulation in a Naturally Infected Flock of Sheep: Individual Follow-Up of Antibodies in Serum and Milk.

PubMed

Joulié, A; Rousset, E; Gasqui, P; Lepetitcolin, E; Leblond, A; Sidi-Boumedine, K; Jourdain, E

2017-07-01

The control of Q fever, a zoonotic disease caused by the Coxiella burnetii bacterium, remains a scientific challenge. Domestic ruminants are considered the main reservoir, shedding C. burnetii essentially through parturition products during abortion or birth. Sheep are particularly frequently associated with human outbreaks, but there are insufficient field data to fully understand disease dynamics and to instigate efficient control measures. A longitudinal follow-up study of a naturally infected sheep flock was performed (i) to investigate relationships between seropositivity and bacterial shedding in the vaginal mucus, (ii) to describe the kinetics of antibodies, including responses to vaccination, (iii) to monitor maternal antibodies in ewe lambs, and (iv) to compare serological results for milk and serum samples. For 8 months, we collected blood samples every 3 weeks from 11 aborting and 26 nonaborting dairy ewes, 20 nonaborting suckler ewes, and 9 ewe lambs. Individual milk samples were also obtained from lactating females. All serum and milk samples were tested by enzyme-linked immunosorbent assay (ELISA), whereas vaginal swabs were tested by quantitative PCR. We found that some dairy females did not seroconvert despite shedding C. burnetii in their vaginal mucus. Overall, antibody levels in adult females were found to remain stable over time, with exceptions during the mating and lambing periods. Maternal antibodies decreased during the first month after birth. Interestingly, antibody levels in milk were correlated with those in serum. This study provides valuable field data that will help improve Q fever surveillance and within-flock management measures. IMPORTANCE Field data are necessary to improve the surveillance, diagnosis, and sanitary management of Q fever in livestock. Here, we provide extensive serological data obtained from serum and milk samples from infected and vaccinated ewes belonging to a naturally infected flock of sheep. We show that antibody levels are stable over time and seropositivity and vaginal shedding are not clearly correlated, whereas antibody levels in milk are strongly correlated with those in serum. Accordingly, we find that antibody levels in bulk tank milk are consistent with the variations observed in the serum of dairy females over time. We report the existence of maternal antibody transmission to ewe lambs and we show that the presence of maternal antibodies at birth does not prevent the development of a serological response to vaccination at the age of 4 months. Finally, we report that adult ewes generally seroconvert after vaccination, including during pregnancy. Copyright © 2017 American Society for Microbiology.

Determination of bisphenol A in human serum by high-performance liquid chromatography with multi-electrode electrochemical detection.

PubMed

Inoue, K; Kato, K; Yoshimura, Y; Makino, T; Nakazawa, H

2000-11-10

A simple and sensitive method using high-performance liquid chromatography with multi-electrode electrochemical detection (HPLC-ED) including a coulometric array of four electrochemical sensors has been developed for the determination of bisphenol A in water and human serum. For good separation and detection of bisphenol A, a CAPCELL PAK UG 120 C18 reversed-phase column and a mobile phase consisting of 0.3% phosphoric acid-acetonitrile (60:40) were used. The detection limit obtained by the HPLC-ED method was 0.01 ng/ml (0.5 pg), which was more than 3000-times higher than the detection limit obtained by the ultraviolet (UV) method, and more than 200-times higher than the detection limit obtained by the fluorescence (FL) method. Bisphenol A in water and serum samples was pretreated by solid-phase extraction (SPE) after removing possible contamination derived from a plastic SPE cartridges and water used for the pretreatment. A trace amount (ND approximately 0.013 ng/ml) of bisphenol A was detected from the parts of cartridges (filtration column, sorbent bed and frits) by extraction with methanol, and it was completely removed by washing with at least 15 ml of methanol in the operation process. The concentrations of bisphenol A in tap water and Milli-Q-purified water were found to be 0.01 and 0.02 ng/ml, respectively. For that reason, bisphenol A-free water was made to trap bisphenol A in water using an Empore disk. In every pretreatment, SPE methods using bisphenol A-free water and washing with 15 ml of methanol were done in water and serum samples. The yields obtained from the recovery tests using water to which 0.5 or 0.05 ng/ml of bisphenol A was added were 83.8 to 98.2%, and the RSDs were 3.4 to 6.1%, respectively. The yields obtained from the recovery tests by OASIS HLB using serum to which 1.0 ng/ml or 0.1 ng/ml of bisphenol A was added were 79.0% and 87.3%, and the RSDs were 5.1% and 13.5%, respectively. The limits of quantification in water and serum sample were 0.01 ng/ml and 0.05 ng/ml, respectively. The method was applied to the determination of bisphenol A in healthy human serum sample, and the obtained detection was 0.32 ng/ml. From these results, the HPLC-ED method should be the most useful in the determination of bisphenol A at low concentration levels in water and biological samples.

Evaluation of the Efficacy, Potential for Vector Transmission, and Duration of Immunity of MP-12, an Attenuated Rift Valley Fever Virus Vaccine Candidate, in Sheep

DTIC Science & Technology

2015-08-01

testing by virus isolation (VI). Positive-control mosquitoes were sampled whole on the day of the spiked blood meal. b NA, not applicable. Miller et al ...percent inhibition of the negative control, calculated as 1 (test serum OD/negative serum OD) 100. Miller et al . 934 cvi.asm.org August 2015 Volume 22...Trop Med Hyg 44:278 –282. 34. Morrill JC, Mebus CA, Peters CJ. 1997. Safety of a mutagen-attenuated Miller et al . 936 cvi.asm.org August 2015 Volume 22

Optimization and Validation of a Plaque Reduction Neutralization Test for the Detection of Neutralizing Antibodies to Four Serotypes of Dengue Virus Used in Support of Dengue Vaccine Development

PubMed Central

Timiryasova, Tatyana M.; Bonaparte, Matthew I.; Luo, Ping; Zedar, Rebecca; Hu, Branda T.; Hildreth, Stephen W.

2013-01-01

A dengue plaque reduction neutralization test (PRNT) to measure dengue serotype–specific neutralizing antibodies for all four virus serotypes was developed, optimized, and validated in accordance with guidelines for validation of bioanalytical test methods using human serum samples from dengue-infected persons and persons receiving a dengue vaccine candidate. Production and characterization of dengue challenge viruses used in the assay was standardized. Once virus stocks were characterized, the dengue PRNT50 for each of the four serotypes was optimized according to a factorial design of experiments approach for critical test parameters, including days of cell seeding before testing, percentage of overlay carboxymethylcellulose medium, and days of incubation post-infection to generate a robust assay. The PRNT50 was then validated and demonstrated to be suitable to detect and measure dengue serotype-specific neutralizing antibodies in human serum samples with acceptable intra-assay and inter-assay precision, accuracy/dilutability, specificity, and with a lower limit of quantitation of 10. PMID:23458954

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

PubMed

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

2014-04-01

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

A rapid field test for sylvatic plague exposure in wild animals

USGS Publications Warehouse

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

2014-01-01

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

A macro-enzyme cause of an isolated increase of alkaline phosphatase.

PubMed

Cervinski, Mark A; Lee, Hong Kee; Martin, Isabella W; Gavrilov, Dimitar K

2015-02-02

Macroenzyme complexes of serum enzymes and antibody can increase the circulating enzymatic activity and may lead to unnecessary additional testing and procedures. Laboratory physicians and scientists need to be aware of techniques to identify macroenzyme complexes when suspected. To investigate the possibility of a macro-alkaline phosphatase in the serum of a 74 year old male with persistently increased alkaline phosphatase we coupled a protein A/G agarose affinity chromatography technique with isoenzyme electrophoresis to look for the presence of macro-alkaline phosphatase. The majority of the alkaline phosphatase activity in the patient's serum sample was bound to the column and only a minor fraction (25%) of alkaline phosphatase activity was present in the column flow-through. The alkaline phosphatase activity was also found to co-elute with the immunoglobulins in the patient sample. The alkaline phosphatase activity in a control serum sample concurrently treated in the same manner did not bind to the column and was found in the column flow-through. The use of protein A/G agarose affinity chromatography is a rapid and simple method that can be applied to the investigation of other macro-enzyme complexes. Copyright © 2014 Elsevier B.V. All rights reserved.

A Human Serum-Based Enzyme-Free Continuous Glucose Monitoring Technique Using a Needle-Type Bio-Layer Interference Sensor.

PubMed

Seo, Dongmin; Paek, Sung-Ho; Oh, Sangwoo; Seo, Sungkyu; Paek, Se-Hwan

2016-09-24

The incidence of diabetes is continually increasing, and by 2030, it is expected to have increased by 69% and 20% in underdeveloped and developed countries, respectively. Therefore, glucose sensors are likely to remain in high demand in medical device markets. For the current study, we developed a needle-type bio-layer interference (BLI) sensor that can continuously monitor glucose levels. Using dialysis procedures, we were able to obtain hypoglycemic samples from commercial human serum. These dialysis-derived samples, alongside samples of normal human serum were used to evaluate the utility of the sensor for the detection of the clinical interest range of glucose concentrations (70-200 mg/dL), revealing high system performance for a wide glycemic state range (45-500 mg/dL). Reversibility and reproducibility were also tested over a range of time spans. Combined with existing BLI system technology, this sensor holds great promise for use as a wearable online continuous glucose monitoring system for patients in a hospital setting.

Equine infectious anemia prevalence in feral donkeys from Northeast Brazil.

PubMed

Oliveira, Fernanda G; Cook, R Frank; Naves, João H F; Oliveira, Cairo H S; Diniz, Rejane S; Freitas, Francisco J C; Lima, Joseney M; Sakamoto, Sidnei M; Leite, Rômulo C; Issel, Charles J; Reis, Jenner K P

2017-05-01

Equine infectious anemia virus (EIAV) is an important cause of morbidity and mortality throughout the world. Although the virus infects all members of the Equidae the vast majority of studies have been conducted in horses (Equus caballus) with comparatively little information available for other equid species. Brazil has one of the most abundant donkey (E. asinus) populations of any nation although the economic importance of these animals is declining as transportation becomes increasingly mechanized. As a result, considerable numbers of donkeys especially in the Northeast of the country have been released and allowed pursue an almost feral existence. Consequently, this large and growing population constitutes a significant risk as a reservoir for the maintenance and transmission of important equine infectious diseases such as glanders and equine arteritis virus in addition to EIAV. This study examines the prevalence of EIA in a semi-wild donkey population from Mossoró city, in Northeast Brazil, using AGID followed by cELISA, rgp90 ELISA and immunoblot (IB). Serum samples were collected from 367 donkeys without obvious EIA clinical signs. Subsequent testing revealed seropositive rates of 1.6% (6/367) in officially approved AGID tests, 3.3% (12/367) in cELISA and 14.4% (53/367) in the rgp90 ELISA. However, 88.7% (47/53) of the rgp90 ELISA positive samples were almost certainly false reactions because they failed to react with two or more antigens in IB. Consequently, the rpg90 ELISA has a similar sensitivity to AGID with donkey serum samples. Such high false positive rates have not been observed previously with serum samples from horses. Another highly significant finding is that 56.9% (33/58) of the donkey serum samples tested in IB had reactivity to EIAV p26 only. Although this could result from recent infection with the virus, it has been found that in some equids p26 only reactivity persists for extensive periods of time suggesting exposure to antigens possessing cross-reactive determinants or EIAV strains with envelope glycoproteins that are different from any that have been previously characterized and so undetectable by current IB techniques. Copyright © 2017 Elsevier B.V. All rights reserved.

Mycothiol acetyltransferase (Rv0819) of Mycobacterium tuberculosis is a potential biomarker for direct diagnosis of tuberculosis using patient serum specimens.

PubMed

Zeitoun, H; Bahey-El-Din, M; Kassem, M A; Aboushleib, H M

2017-12-01

Mycobacterium tuberculosis infection constitutes a global threat that results in significant morbidity and mortality worldwide. Efficient and early diagnosis of tuberculosis (TB) is of paramount importance for successful treatment. The aim of the current study is to investigate the mycobacterial mycothiol acetyltransferase Rv0819 as a potential novel biomarker for the diagnosis of active TB infection. The gene encoding Rv0819 was cloned and successfully expressed in Escherichia coli. The recombinant Rv0819 was purified using metal affinity chromatography and was used to raise murine polyclonal antibodies against Rv0819. The raised antibodies were employed for direct detection of Rv0819 in patient serum samples using dot blot assay and competitive enzyme-linked immunosorbent assay (ELISA). Serum samples were obtained from 68 confirmed new TB patients and 35 healthy volunteers as negative controls. The dot blot assay showed sensitivity of 64·7% and specificity of 100%, whereas the competitive ELISA assay showed lower sensitivity (54·4%) and specificity (88·57%). The overall sensitivity of the combined results of the two tests was found to be 89·7%. Overall, the mycobacterial Rv0819 is a potential TB serum biomarker that can be exploited, in combination with other TB biomarkers, for efficient and reliable diagnosis of active TB infection. The early and accurate diagnosis of tuberculosis infection is of paramount importance for initiating treatment and avoiding clinical complications. Most current diagnostic tests have poor sensitivity and/or specificity and in many cases they are too expensive for routine diagnostic testing in resource-limited settings. In the current study, we examined a novel mycobacterial serum biomarker, namely mycothiol acetyltransferase Rv0819. The antigen was detectable in serum specimens of a significant number of tuberculosis patients. This article proves the importance of Rv0819 and paves the way towards its future use as a useful diagnostic marker for tuberculosis infection. © 2017 The Society for Applied Microbiology.

Development and evaluation of a truncated recombinant NS3 antigen-based indirect ELISA for detection of pestivirus antibodies in sheep and goats.

PubMed

Kalaiyarasu, Semmannan; Mishra, Niranjan; Rajukumar, Katherukamem; Nema, Ram Kumar; Behera, Sthita Pragnya

2015-01-01

The aim of this study was to develop an indirect ELISA using the helicase domain of bovine viral diarrhoea virus (BVDV) NS3 protein instead of full-length NS3 protein for detection of BVDV and BDV antibodies in sheep and goats and its validation by comparing its sensitivity and specificity with virus neutralization test (VNT) as the reference test. The purified 50 kDa recombinant NS3 protein was used as the coating antigen in the ELISA. The optimal concentration of antigen was 320 ng/well at a serum dilution of 1:20 and the optimal positive cut-off optical density value was 0.40 based on test results of 418 VNT negative sheep and goat sera samples. When 569 serum samples from sheep (463) and goats (106) were tested, the ELISA showed a sensitivity of 91.71% and specificity of 94.59% with BVDV VNT. A good correlation (93.67%) was observed between the two tests. It showed a sensitivity of 85% and specificity of 86.6% with VNT in detecting BDV antibody positive or negative samples. This study demonstrates the efficacy of truncated recombinant NS3 antigen based ELISA for seroepidemiological study of pestivirus infection in sheep and goats.

Serum neuron-specific enolase levels from the same patients differ between laboratories: assessment of a prospective post-cardiac arrest cohort.

PubMed

Mlynash, Michael; Buckwalter, Marion S; Okada, Ami; Caulfield, Anna Finley; Venkatasubramanian, Chitra; Eyngorn, Irina; Verbeek, Marcel M; Wijman, Christine A C

2013-10-01

In comatose post-cardiac arrest patients, a serum neuron-specific enolase (NSE) level of >33 μg/L within 72 h was identified as a reliable marker for poor outcome in a large Dutch study (PROPAC), and this level was subsequently adopted in an American Academy of Neurology practice parameter. Later studies reported that NSE >33 μg/L is not a reliable predictor of poor prognosis. To test whether different clinical laboratories contribute to this variability, we compared NSE levels from the laboratory used in the PROPAC study (DLM-Nijmegen) with those of our hospital's laboratory (ARUP) using paired blood samples. We prospectively enrolled cardiac arrest patients who remained comatose after resuscitation. During the first 3 days, paired blood samples for serum NSE were drawn at a median of 10 min apart. After standard preparation for each lab, one sample was sent to ARUP laboratories and the other to DLM-Nijmegen. Fifty-four paired serum samples from 33 patients were included. Although the serum NSE measurements correlated well between laboratories (R = 0.91), the results from ARUP were approximately 30% lower than those from DLM-Nijmegen. Therapeutic hypothermia did not affect this relationship. Two patients had favorable outcomes after hypothermia despite NSE levels measured by DLM-Nijmegen as >33 μg/L. Absolute serum NSE levels of comatose cardiac arrest patients differ between laboratories. Any specific absolute cut-off levels proposed to prognosticate poor outcome should not be used without detailed data on how neurologic outcomes correspond to a particular laboratory's method, and even then only in conjunction with other prognostic variables.

Serum herpes simplex antibodies

MedlinePlus

... causes cold sores (oral herpes). HSV-2 causes genital herpes. How the Test is Performed A blood sample ... person has ever been infected with oral or genital herpes . It looks for antibodies to herpes simplex virus ...

[Detecting the markers of HIV infection with the new enzyme immunoassay diagnostic kit "DS-EIA-HIV-AB-AG-SPECTRUM" at the laboratories of AIDS prevention and control centers in the Volga Federal District].

PubMed

Ivanova, N I; Peksheva, O Iu

2009-03-01

A possibility of simultaneously detecting specific antibodies to HIV-1 and HIV-2 by enzyme immunoassay (EIA) at lower concentrations than those by immunoblotting (IB), and well as an additional possibility of earlier diagnosis of HIV infection, by identifying the HIV-1 antigen p24 lay the foundation of the "DS-EIA-HIV-AB-AG-SPECTRUM" test system made by OOO "Research-and-Production Association "Diagnosticheskiye Sistemy" (Diagnostic Systems). These peculiarities were compared with those of IB at a number of laboratories of AIDS prevention and control centers in the Volga Federal District, by using native serum/plasma samples and a specially designed control panel. The analysis of the conducted studies to identify HIV-1 and HIV-2 antibodies and HIV-1 antigen p24 in 65 plasma/serum samples in the "DS-EIA-HIV-AB-AG-SPECTRUM" and "LIA-HIV-1/2" (OOO "Niarmedik plus") test systems while confirming the positive result indicated agreement in 57 (87.7%) cases. The diagnostic possibilities of the "DS-EIA-HIV-AB-AG-SPECTRUM" test system versus the "New Lav-Blot I" one to make a laboratory diagnosis of HIV infection were studied. Irrefragable answers as to the availability of HIV-1 markers in the study serum samples on the enciphered panel were provided by IB in 73.3% of cases and EIA in 92%.

Evaluation of diagnostic assays for the serological detection of Actinobacillus pleuropneumoniae on samples of known or unknown exposure.

PubMed

Opriessnig, Tanja; Hemann, Michelle; Johnson, John K; Heinen, Sheila; Giménez-Lirola, Luis G; O'Neill, Kevin C; Hoang, Hai; Yoon, Kyoung-Jin; Gottschalk, Marcelo; Halbur, Patrick G

2013-01-01

Accurate diagnosis of exposure to Actinobacillus pleuropneumoniae is important for maintaining negative farms. In the present study, the ability of a dual-plate complement fixation (CF) assay and 3 commercially available enzyme-linked immunosorbent assays (ELISAs; quad-plate ELISA-1, single-plate ELISA-2, and single-plate ELISA-3) in detecting serological evidence of A. pleuropneumoniae exposure was compared using serum samples of experimentally infected or vaccinated pigs, or field samples from the United States. Forty-two pigs were divided into groups of 2 pigs and were inoculated with 1 of 15 A. pleuropneumoniae strains representing all known serovars of A. pleuropneumoniae, or with Actinobacillus suis, or were vaccinated with a bacterin containing A. pleuropneumoniae serovar 1, 3, 5, or 7. Serum samples collected at the day of inoculation or vaccination and 7, 14, 21, and 28 days later were used to compare the assays. On samples from experimentally infected pigs, the dual-plate CF assay, quad-plate ELISA-1, single-plate ELISA-2, and single-plate ELISA-3 had sensitivities of 0.46, 0.74, 0.13, and 0.13 and specificities of 0.90, 1.0, 1.0, and 1.0, respectively. Vaccinated pigs were identified only by the dual-plate CF assay and the quad-plate ELISA-1. In addition, 90 serum samples with unknown A. pleuropneumoniae exposure collected under field conditions were tested with all assays. The agreement of the 4 assays on field samples was slight to fair. While several assays are available for demonstration of A. pleuropneumoniae exposure, differences in assay targets complicate test choices. Decisions on which assay or combination of assays to use depend on the specific reasons for running the assays.

Exploration of attenuated total reflectance mid-infrared spectroscopy and multivariate calibration to measure immunoglobulin G in human sera.

PubMed

Hou, Siyuan; Riley, Christopher B; Mitchell, Cynthia A; Shaw, R Anthony; Bryanton, Janet; Bigsby, Kathryn; McClure, J Trenton

2015-09-01

Immunoglobulin G (IgG) is crucial for the protection of the host from invasive pathogens. Due to its importance for human health, tools that enable the monitoring of IgG levels are highly desired. Consequently there is a need for methods to determine the IgG concentration that are simple, rapid, and inexpensive. This work explored the potential of attenuated total reflectance (ATR) infrared spectroscopy as a method to determine IgG concentrations in human serum samples. Venous blood samples were collected from adults and children, and from the umbilical cord of newborns. The serum was harvested and tested using ATR infrared spectroscopy. Partial least squares (PLS) regression provided the basis to develop the new analytical methods. Three PLS calibrations were determined: one for the combined set of the venous and umbilical cord serum samples, the second for only the umbilical cord samples, and the third for only the venous samples. The number of PLS factors was chosen by critical evaluation of Monte Carlo-based cross validation results. The predictive performance for each PLS calibration was evaluated using the Pearson correlation coefficient, scatter plot and Bland-Altman plot, and percent deviations for independent prediction sets. The repeatability was evaluated by standard deviation and relative standard deviation. The results showed that ATR infrared spectroscopy is potentially a simple, quick, and inexpensive method to measure IgG concentrations in human serum samples. The results also showed that it is possible to build a united calibration curve for the umbilical cord and the venous samples. Copyright © 2015 Elsevier B.V. All rights reserved.

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

PubMed Central

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

2017-01-01

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

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

PubMed

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

2017-01-01

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

Posttraumatic Stress Disorder, Exposure to Combat, and Lower Plasma Cortisol among Vietnam Veterans: Findings and Clinical Implications.

ERIC Educational Resources Information Center

Boscarino, Joseph A.

1996-01-01

Clinical studies suggest individuals with posttraumatic stress disorder (PSD) experience neuroendocrine systems alterations, resulting in significantly lower plasma cortisol. To test this hypothesis, morning serum cortisol was compared among a national sample of Vietnam "theater" veterans (n=2,490) and a sample of Vietnam "era"…

Co-infections with Chikungunya and Dengue Viruses, Guatemala, 2015.

PubMed

Edwards, Thomas; Signor, Leticia Del Carmen Castillo; Williams, Christopher; Donis, Evelin; Cuevas, Luis E; Adams, Emily R

2016-11-01

We screened serum samples referred to the national reference laboratory in Guatemala that were positive for chikungunya or dengue viruses in June 2015. Co-infection with both viruses was detected by reverse transcription PCR in 46 (32%) of 144 samples. Specimens should be tested for both arboviruses to detect co-infections.

Seroprevalence of Toxoplasma gondii antibodies in captive wild mammals and birds in Brazil.

USDA-ARS?s Scientific Manuscript database

In this study serum samples of 203 animals from different locations from zoos and breeding facilities from the north and northeast region of Brazil were analyzed for the presence of anti-Toxoplasma gondii antibodies by the modified agglutination test (MAT) with a cutoff of 1:25. Of the sampled anima...

Borrelia miyamotoi sensu lato seroreactivity and seroprevalence in the northeastern United States.

PubMed

Krause, Peter J; Narasimhan, Sukanya; Wormser, Gary P; Barbour, Alan G; Platonov, Alexander E; Brancato, Janna; Lepore, Timothy; Dardick, Kenneth; Mamula, Mark; Rollend, Lindsay; Steeves, Tanner K; Diuk-Wasser, Maria; Usmani-Brown, Sahar; Williamson, Phillip; Sarksyan, Denis S; Fikrig, Erol; Fish, Durland

2014-07-01

Borrelia miyamotoi sensu lato, a relapsing fever Borrelia sp., is transmitted by the same ticks that transmit B. burgdorferi (the Lyme disease pathogen) and occurs in all Lyme disease-endemic areas of the United States. To determine the seroprevalence of IgG against B. miyamotoi sensu lato in the northeastern United States and assess whether serum from B. miyamotoi sensu lato-infected persons is reactive to B. burgdorferi antigens, we tested archived serum samples from area residents during 1991-2012. Of 639 samples from healthy persons, 25 were positive for B. miyamotoi sensu lato and 60 for B. burgdorferi. Samples from ≈10% of B. miyamotoi sensu lato-seropositive persons without a recent history of Lyme disease were seropositive for B. burgdorferi. Our results suggest that human B. miyamotoi sensu lato infection may be common in southern New England and that B. burgdorferi antibody testing is not an effective surrogate for detecting B. miyamotoi sensu lato infection.

Attempts to identify Clostridium botulinum toxin in milk from three experimentally intoxicated Holstein cows

USGS Publications Warehouse

Moeller, R.B.; Puschner, B.; Walker, R.L.; Rocke, T.E.; Smith, S.R.; Cullor, J.S.; Ardans, A.A.

2009-01-01

Three adult lactating Holstein cows were injected in the subcutaneous abdominal vein with 175 ng/kg of body weight of Clostridium botulinum type C toxin (451 cow median toxic doses) to determine if this botulinum toxin crosses the blood–milk barrier. Whole blood (in sodium heparin) and clotted blood serum samples were taken at 0 min, 10 min, and 3, 6, 9, and 12 h postinoculation. Milk samples were taken at 0 min and at 3, 6, 9 and 12 h postinoculation. All samples were tested for the presence of the toxin using the mouse bioassay and immunostick ELISA test. The immunostick ELISA identified the toxin in whole blood and the mouse bioassay identified the toxin in serum at all times examined in all 3 animals. Toxin was not identified by either detection method in milk samples collected from the 3 animals. From these results, it appears that Clostridium botulinum type C toxin does not cross from the blood to the milk in detectable concentrations.

Identification of periparturient mare and foal associated predictors of post parturient immunoglobulin A concentrations in Thoroughbred foals.

PubMed

Jenvey, C; Caraguel, C; Howarth, G B; Riley, C B

2012-12-01

Prior to the start of endogenous production of immunoglobulins (Igs), absorption of maternal Igs is important to protect against pathogens in the early neonatal period. It is possible that mare- or foal-associated factors may influence neonatal IgA concentrations. The temporal relationships among serum and milk IgA concentrations in Thoroughbred mare-foal pairs were explored to determine if periparturient mare- and foal-associated factors contribute to the prediction of foal serum IgA concentrations. Blood and milk samples as well as complete veterinary records, were collected for 84 Thoroughbred mare-foal pairs from one month before to 2 months after parturition. Samples were tested using enzyme-linked immunosorbent assay (ELISA) for concentrations of IgA. Pairwise correlation coefficients were estimated (P < 0.01) and simple linear regression used to investigate unconditional associations between mare IgA levels, mare and foal risk factors and foal serum IgA concentration at 12 h. Backwards, stepwise elimination of nonsignificant factors was used to create a final model. There were significant temporal relationships among mare serum IgA and among colostrum and milk IgA concentrations within mares (P < 0.01). Mare serum IgA concentrations up to one month before parturition were associated with foal serum IgA concentrations at all time points and with colostrum and milk IgA concentrations. Mare serum IgA at -28 days and parity were associated with foal serum IgA concentration at 12 h (P < 0.001). Mare serum IgA concentrations up to 28 days before parturition, together with mare parity, are indicative of neonatal foal serum IgA concentrations. Mare serum and colostrum IgA concentrations may be useful peripartum predictors of neonatal mucosal immune status, enabling earlier intervention to prevent the consequences of mucosal infections.

The Platelia Aspergillus ELISA in diagnosis of invasive pulmonary aspergilosis (IPA).

PubMed

Siemann, M; Koch-Dörfler, M

2001-01-01

The sensitivity of a sandwich enzyme-linked immunosorbent assay (ELISA) for detecting Aspergillus galactomannan was evaluated with 66 serum samples and 113 specimens of the respiratory tract obtained from 52 patients with pulmonary diseases. The patients were divided into five groups: proven invasive pulmonary aspergillosis (IPA) (five patients), probable IPA (seven patients), Aspergillus colonization (eight patients) or unlikely Aspergillus infection (27 patients). Another five patients with doubtful diagnostic test results are discussed in detail. The results of the Platelia Aspergillus ELISA (Sanofi Pasteur, Freiburg, Germany) in testing specimens of the respiratory tract were 90% sensitivity in proven (serum 38%), 60% in probable (serum 37%) and 71% in Aspergillus colonization (serum 0%). Furthermore, 85% of the Aspergillus spp. from positive cultures of specimens of the respiratory tract were also detected in the ELISA. A total of 57% of the culture negative specimens of patients with a least one positive culture or proven aspergillosis in a series of specimens were positive in the ELISA.

Serum creatinine and alkaline phosphatase levels are associated with severe chronic periodontitis.

PubMed

Caúla, A L; Lira-Junior, R; Tinoco, E M B; Fischer, R G

2015-12-01

Periodontitis may alter systemic homeostasis and influence creatinine and alkaline phosphatase levels. Therefore, the aim of this study was to evaluate the relationship between severe chronic periodontitis and serum creatinine and alkaline phosphatase levels. One hundred patients were evaluated, 66 with severe chronic periodontitis (test group) and 34 periodontally healthy controls (control group). Medical, demographic and periodontal parameters were registered. Blood sample was collected after an overnight fast and serum creatinine and alkaline phosphatase levels were determined. There were significant differences between test and control groups in ethnicity, gender and educational level (p < 0.05). Patients with periodontitis showed a lower mean creatinine level (p < 0.05) and higher mean alkaline phosphatase level (p < 0.001) than the control group. There were significant correlations between periodontal parameters and serum creatinine and alkaline phosphatase levels. Severe chronic periodontitis was associated to lower creatinine and higher alkaline phosphatase levels. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

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

PubMed

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

2006-06-01

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

Comparison of specimens for detection of porcine reproductive and respiratory syndrome virus infection in boar studs.

PubMed

Pepin, B J; Kittawornrat, A; Liu, F; Gauger, P C; Harmon, K; Abate, S; Main, R; Garton, C; Hargrove, J; Rademacher, C; Ramirez, A; Zimmerman, J

2015-06-01

Porcine reproductive and respiratory syndrome virus (PRRSV)-contaminated semen from boars is a route of transmission to females, and early detection of PRRSV infection in boars is a key component in sow farm biosecurity. The purpose of this study was to determine the optimum diagnostic specimen(s) for the detection of acute PRRSV infection in boars. Individually housed boars (n = 15) were trained for semen and oral fluid collection and then vaccinated with a commercial PRRSV modified live virus vaccine. Starting on the day of vaccination and for 14 days thereafter, oral fluid specimens were collected daily from all boars. The 15 boars were subdivided into three groups of 5, and serum, blood swabs and 'frothy saliva' were collected at the time of semen collection on a 3-day rotation. Frothy saliva, derived from the submandibular salivary gland, is produced by aroused boars. Semen was centrifuged, and semen supernatant and cell fractions were tested separately. All samples were randomly ordered and then tested by PRRSV real-time quantitative reverse-transcription polymerase chain reaction assay (rRT-PCR) and PRRSV antibody ELISA. In this study, a comparison of serum, blood swab, and oral fluid rRT-PCR results found no statistically significant differences in the onset of detection or proportion of positives, but serum was numerically superior to oral fluids for early detection. Serum and oral fluid provided identical rRT-PCR results at ≥ 5 day post-vaccination. Likewise, the onset of detection of PRRSV antibody in serum, oral fluid and frothy saliva was statistically equivalent, with serum results again showing a numerical advantage. These results showed that the highest assurance of providing PRRSV-negative semen to sow farms should be based on rRT-PCR testing of serum collected at the time of semen collection. This approach can be augmented with oral fluid sampling from a random selection of uncollected boars to provide for statistically valid surveillance of the boar stud. © 2013 Blackwell Verlag GmbH.

Canine cancer screening via ultraviolet absorbance and fluorescence spectroscopy of serum proteins

NASA Astrophysics Data System (ADS)

Dickerson, Bryan D.; Geist, Brian L.; Spillman, William B., Jr.; Robertson, John L.

2007-11-01

A cost-effective optical cancer screening and monitoring technique was demonstrated in a pilot study of canine serum samples and was patented for commercialization. Compared to conventional blood chemistry analysis methods, more accurate estimations of the concentrations of albumin, globulins, and hemoglobin in serum were obtained by fitting the near UV absorbance and photoluminescence spectra of diluted serum as a linear combination of component reference spectra. Tracking these serum proteins over the course of treatment helped to monitor patient immune response to carcinoma and therapy. For cancer screening, 70% of dogs with clinical presentation of cancer displayed suppressed serum hemoglobin levels (below 20 mg/dL) in combination with atypical serum protein compositions, that is, albumin levels outside of a safe range (from 4 to 8 g/dL) and globulin levels above or below a more normal range (from 1.7 to 3.7 g/dL). Of the dogs that met these criteria, only 20% were given a false positive label by this cancer screening test.

An Immunoassay to Rapidly Measure Acetaminophen Protein Adducts Accurately Identifies Patients with Acute Liver Injury or Failure

PubMed Central

Roberts, Dean W.; Lee, William M.; Hinson, Jack A.; Bai, Shasha; Swearingen, Christopher J.; Stravitz, R. Todd; Reuben, Adrian; Letzig, Lynda; Simpson, Pippa M.; Rule, Jody; Fontana, Robert J.; Ganger, Daniel; Reddy, K. Rajender; Liou, Iris; Fix, Oren; James, Laura P.

2017-01-01

Background & Aims A rapid, reliable point-of-care assay to detect acetaminophen protein adducts in serum of patients with acute liver injury could improve diagnosis and management. AcetaSTAT is a competitive immunoassay used to measure acetaminophen protein adducts formed by toxic metabolites in serum samples from patients. We compared the accuracy of AcetaSTAT vs high-pressure liquid chromatography with electrochemical detection (HPLC-EC, a sensitive and specific quantitative analytical assay) to detect acetaminophen protein adducts. Methods We collected serum samples from 19 healthy individuals (no liver injury, no recent acetaminophen use), 29 patients without acetaminophen-associated acute liver injury, and 33 patients with acetaminophen-associated acute liver injury participating in the Acute Liver Failure Study Group registry. Each serum sample was analyzed by AcetaSTAT (reported as test band amplitude) and HPLC-EC (the reference standard). We also collected data on patient age, sex, weight, level of alanine aminotransferase on test day and peak values, concentration of acetaminophen, diagnoses (by site investigator and causality review committee), and outcome after 21 days. Differences between groups were analyzed using Fisher’s Exact for categorical variables and Kruskal-Wallis Test or Rank-Sum test for continuous variables. Results AcetaSTAT discriminated between patients with and without acetaminophen-associated acute liver injury; the median (and range) AcetaSTAT test band amplitude for patients with acetaminophen-associated acute liver injury was 584 (range, 222–1027) vs 3678 (range, 394–8289) for those without (P<.001). AcetaSTAT identified patients with acetaminophen-associated acute liver injury with 100% sensitivity, 86.2% specificity, a positive-predictive value of 89.2%, and a negative-predictive value of 100%. Results from AcetaSTAT were positive in 4 subjects who received a causality review committee diagnosis of non-acetaminophen–associated acute liver injury; HPLC-EC and biochemical profiles were consistent with acetaminophen-associated acute liver injury in 3 of these cases. Conclusion The competitive immunoassay AcetaSTAT demonstrates a high degree of concordance with HPLC-EC results in identifying patients with acetaminophen-associated acute liver injury. This rapid and simple assay could increase early detection of this disorder and aid clinical management. PMID:27641661

Comparison of different commercial ELISAs for detection of antibodies against porcine respiratory and reproductive syndrome virus in serum.

PubMed

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

2014-12-18

In recent years, several new ELISAs for the detection of antibodies against the porcine reproductive and respiratory disease virus (PRRSV) in pig serum have been developed. To interpret the results, specificity and sensitivity data as well as agreement to a reference ELISA must be available. In this study, three commercial ELISAs (INgezim PRRS 2.0 - ELISA II, Priocheck® PRRSV Ab porcine - ELISA III and CIVTEST suis PRRS E/S PLUS - ELISA IV, detecting PRRSV type 1 antibodies) were compared to a standard ELISA (IDEXX PRRS X3 Ab Test - ELISA I). The serum of three pigs vaccinated with an attenuated PRRSV live vaccine (genotype 2) was tested prior to and several times after the vaccination. Furthermore, serum samples of 245 pigs of PRRSV positive herds, 309 pigs of monitored PRRSV negative herds, 256 fatteners of assumed PRRSV negative herds with unknown herd history and 92 wild boars were tested with all four ELISAs. ELISAs II and III were able to detect seroconversion of vaccinated pigs with a similar reliability. According to kappa coefficient, the results showed an almost perfect agreement between ELISA I as reference and ELISA II and III (kappa > 0.8), and substantial agreement between ELISA I and ELISA IV (kappa = 0.71). Sensitivity of ELISA II, III and IV was 96.0%, 100% and 91.5%, respectively. The specificity of the ELISAs determined in samples of monitored PRRSV negative herds was 99.0%, 95.1% and 96.4%, respectively. In assumed negative farms that were not continually monitored, more positive samples were found with ELISA II to IV. The reference ELISA I had a specificity of 100% in this study. All tested ELISAs were able to detect a PRRSV positive herd. The specificity and sensitivity of the tested commercial ELISAs, however, differed. ELISA II had the highest specificity and ELISA III had the highest sensitivity in comparison to the reference ELISA. ELISA IV had a lower sensitivity and specificity than the other ELISAs.

Correlation of serum biomarkers (TSA & LSA) and epithelial dysplasia in early diagnosis of oral precancer and oral cancer.

PubMed

Sawhney, Hemant; Kumar, C Anand

Oral cancer is currently the most frequent cause of cancer-related deaths, which is usually preceded by oral pre-cancerous lesions and conditions. Altered glycosylation of glycoconjugates, such as sialic acid, fucose, etc. are amongst the important molecular changes that accompany malignant transformation. The purpose of our study was to evaluate usefulness of serum Total Sialic Acid (TSA) and serum Lipid-Bound Sialic Acid (LSA) as markers of oral precancerous lesions and histopathologically correlating them with grades of epithelial dysplasia. Blood samples were collected from 50 patients with oral precancer (Leukoplakia & OSMF), 25 patients with untreated oral cancer and 25 healthy subjects. Serum sialic acid (total and lipid bound) levels were measured spectrophotometrically. Tissue samples from all the patients were evaluated for dysplasia. Serum levels of total and lipid bound sialic acid were significantly elevated in patients with oral precancer and cancer when compared with healthy subjects. Analysis of variance test documented that there is progressive rise in serum levels of sialic acid with the degree of dysplastic changes in oral precancer patients. We observed positive correlation between serum levels of the markers and the extent of malignant disease (TNM Clinical staging) as well as histopathological grades. The results suggested that serum levels of TSA and LSA progressively increases with grades of dysplasia in precancerous groups and cancer group, when compared with healthy controls. These glycoconjugates, especially LSA has the clinical utility in indicating a premalignant change.

Serum and saliva magnesium in postmenopausal women with xerostomia.

PubMed

Agha-Hosseini, F; Mirzaii-Dizgah, I

2012-10-01

The aim of this study was to investigate serum, stimulated and unstimulated salivary magnesium in postmenopausal women with xerostomia. A case-control study was carried out on 60 selected postmenopausal women aged 41-77 years with or without xerostomia (30 as cases with xerostomia and 30 as controls without xerostomia), conducted at the Clinic of Oral Medicine, Tehran University of Medical Sciences. Unstimulated and paraffin-stimulated saliva samples were obtained by expectoration. Magnesium concentration was determined by the spectrophotometer method. Statistical analysis was carried out using Student's t-test. The mean serum concentration, but not stimulated and unstimulated whole saliva magnesium concentrations, was significantly higher in the cases than in the controls. Serum magnesium level appears to be associated with xerostomia in menopause.

Antioxidant capacity and oxygen radical diseases in the preterm newborn.

PubMed

Rogers, S; Witz, G; Anwar, M; Hiatt, M; Hegyi, T

2000-06-01

Bronchopulmonary dysplasia, intraventricular hemorrhage, necrotizing enterocolitis, and retinopathy of prematurity may be different manifestations of oxygen radical diseases of prematurity (ORDP). To test the hypothesis that the antioxidant capacity of cord blood serum will predict risk of ORDP. An inception cohort of premature neonates was followed up from birth until discharge or death to determine if outcome was related to cord blood serum antioxidant capacity, as determined by a manual assay measuring the relative inhibition of oxidation of 2,2'-azino-di-(3-ethylbenzthiazoline)-6 sulfonic acid (ABTS). Possible correlations between antioxidant capacity and various perinatal factors were also tested. Level 3 newborn intensive care unit. All inborn very low-birth-weight neonates from whom cord blood was available and for whom maternal consent was obtained were included. Newborns who died in the first week of life or who had major congenital malformations were excluded. A convenience sample of newborns weighing more than 1500 g was used to perfect assay and explore confounders. Significant ORDP was defined as the presence of intraventricular hemorrhage greater than grade 2, retinopathy of prematurity greater than stage 1, bronchopulmonary dysplasia at the postconceptional age of 36 weeks, or necrotizing enterocolitis with the hypothesis that neonates with ORDP will have lower antioxidant capacity in cord blood serum. Serum antioxidant capacity at birth correlated with gestational age for the entire sample of 41 neonates and for the 26 neonates born before 32 weeks' gestation. After correction for gestational age, cord serum antioxidant capacity did not correlate with maternal smoking, preeclampsia, chorioamnionitis, cord pH Apgar scores, or any of the ORDP studied. Cord serum antioxidant capacity correlates with gestational age but does not predict ORDP risk.

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

PubMed

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

2014-02-01

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

Prevalence of Parvovirus B19 and Parvovirus V9 DNA and Antibodies in Paired Bone Marrow and Serum Samples from Healthy Individuals

PubMed Central

Heegaard, Erik D.; Petersen, Bodil Laub; Heilmann, Carsten J.; Hornsleth, Allan

2002-01-01

Parvovirus B19 (hereafter referred to as B19) exhibits a marked tropism to human bone marrow (BM), and infection may lead to erythema infectiosum, arthropathy, hydrops fetalis, and various hematologic disorders. Recently, a distinct parvovirus isolate termed V9 with an unknown clinical spectrum was discovered. In contrast to the many studies of B19 serology and viremia, valid information on the frequency of B19 or V9 DNA in the BM of healthy individuals is limited. To develop a reference value, paired BM and serum samples from healthy subjects were tested for the presence of B19 and V9 DNA and specific antibodies. Immunoglobulin M (IgM) was not found in any of the serum samples. The prevalence of IgG showed a gradual and steady increase from 37% in children aged 1 to 5 years to 87% in people aged >50 years. When 190 well-characterized subjects were examined, B19 DNA was detected in the BM of 4 individuals (2.1%; 95% confidence interval, 0.58 to 5.3%) while none of the paired serum samples showed evidence of circulating viral DNA. V9 DNA was not found in any of the BM or serum samples. The finding of B19 DNA probably indicated a primary infection in one 7-year-old individual and reinfection or reactivation of persistent infection in the remaining three persons, aged 47 to 58 years. Serving as a benchmark for future studies, these findings are useful when interpreting epidemiologic data, performing BM transplantation, or considering clinical implications of parvovirus infection. PMID:11880419

Prevalence of parvovirus B19 and parvovirus V9 DNA and antibodies in paired bone marrow and serum samples from healthy individuals.

PubMed

Heegaard, Erik D; Petersen, Bodil Laub; Heilmann, Carsten J; Hornsleth, Allan

2002-03-01

Parvovirus B19 (hereafter referred to as B19) exhibits a marked tropism to human bone marrow (BM), and infection may lead to erythema infectiosum, arthropathy, hydrops fetalis, and various hematologic disorders. Recently, a distinct parvovirus isolate termed V9 with an unknown clinical spectrum was discovered. In contrast to the many studies of B19 serology and viremia, valid information on the frequency of B19 or V9 DNA in the BM of healthy individuals is limited. To develop a reference value, paired BM and serum samples from healthy subjects were tested for the presence of B19 and V9 DNA and specific antibodies. Immunoglobulin M (IgM) was not found in any of the serum samples. The prevalence of IgG showed a gradual and steady increase from 37% in children aged 1 to 5 years to 87% in people aged >50 years. When 190 well-characterized subjects were examined, B19 DNA was detected in the BM of 4 individuals (2.1%; 95% confidence interval, 0.58 to 5.3%) while none of the paired serum samples showed evidence of circulating viral DNA. V9 DNA was not found in any of the BM or serum samples. The finding of B19 DNA probably indicated a primary infection in one 7-year-old individual and reinfection or reactivation of persistent infection in the remaining three persons, aged 47 to 58 years. Serving as a benchmark for future studies, these findings are useful when interpreting epidemiologic data, performing BM transplantation, or considering clinical implications of parvovirus infection.

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

PubMed

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

2013-12-01

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

Field evaluation of two point-of-care tests for syphilis among men who have sex with men, Verona, Italy.

PubMed

Zorzi, Antonella; Cordioli, Maddalena; Gios, Lorenzo; Del Bravo, Paola; Toskin, Igor; Peeling, Rosanna W; Blondeel, Karel; Cornaglia, Giuseppe; Kiarie, James; Ballard, Ronald; Mirandola, Massimo

2017-12-01

The incidence of HIV and syphilis among men who have sex with men (MSM) in Europe has recently increased. Rapid point-of-care tests (POCTs) for syphilis can improve access to screening. The purpose of this study was to evaluate the performance of two syphilis POCTs compared with laboratory tests among MSM. The study was undertaken in Verona, Italy. Asymptomatic MSM, potentially exposed to syphilis, were enrolled prospectively. The POCTs evaluated were SD Bioline Syphilis 3.0 and Chembio DPP Syphilis Screen & Confirm Assay on both serum and fingerprick blood. The results of the POCTs were read by the naked eye by two independent readers and their concordance assessed. A total of 289 MSM were enrolled in the study. Based on laboratory tests, 35 MSM (12.1%) were TPPA-positive alone and 16 (5.5%) were both Treponema pallidum particle agglutination test (TPPA) and rapid plasma reagin (RPR)-positive. The specificities of both POCTs were above 99% on both serum and fingerstick blood specimens, while sensitivities varied considerably. The sensitivity of the SD Bioline test was lower on fingerprick blood (51.4% and 54.3%, readers 1 and 2, respectively) compared with that on serum (80.0% and 82.9%). In contrast, the Chembio test exhibited similar sensitivity values for serum and fingerprick samples (57.7% and 64.0% on serum vs 65.4% and 69.2% on fingerprick for the treponemal component; 63.6% on both samples by both readers for the non-treponemal component). The positive predictive value ranged between 100% and 93.9% for the treponemal component of both syphilis POCTs, but was lower (76.3%-100%)%) for the non-treponemal component of the Chembio POCT. The negative predictive value surpassed 90% for both tests on both samples. The agreement between readers was very high (>99%). The diagnostic performance of the syphilis POCTs was lower than expected; however, considering the prevalence of syphilis among MSM, POCTs should be recommended to improve syphilis detection among MSM. © World Health Organization [2017]. Licensee BMJ Publishing Group Limited. This is an open access article distributed under the terms of the Creative Commons Attribution IGO License (CC BY 4.0), which permits use, distribution, and reproduction for non-commercial purposes in any medium, provided the original work is properly cited. In any reproduction of this article there should not be any suggestion that WHO or this article endorse any specific organization or products. The use of the WHO logo is not permitted. This notice should be preserved along with the article’s original URL.

Albumin - blood (serum)

MedlinePlus

The normal range is 3.4 to 5.4 g/dL (34 to 54 g/L). Normal value ranges may vary slightly among different laboratories. Some labs use different measurements or test different samples. Talk to your provider about ...

Evaluation of the BT-1 serum assay for breast cancer.

PubMed

Whitehurst, M M; Aldenderfer, P H; Sooy, M M; Strelkauskas, A J

1999-01-01

The BT-1 assay which identifies a novel breast tumor associated serum analyte was performed for 143 patients previously diagnosed with breast cancer. Mucin tumor markers CA15-3/CA27-29 values were available for 50 patients and there was very minor overlap between patients positive by both tests. Patients' follow-up clinical status at sample draw was compared to BT-1 assay results. 27% of patients originally diagnosed as Stage II and 20% patients originally diagnosed as Stage III who were evaluated 'no disease' had positive BT-1 values. 8% patients diagnosed as Stage II had negative BT-1 results in samples drawn within 90 days of chemotherapy initiation, whereas 23% of patients diagnosed as Stage III cancer were BT-1 test positive within 90 days of chemotherapy initiation. 50% of patients tested before initial breast cancer surgery had positive BT-1 values, suggesting that the BT-1 assay may be useful in identification women with more advanced disease at diagnosis.

Serological evidence for hepatitis e virus infection in laboratory monkeys and pigs in animal facilities in Japan.

PubMed

Yamamoto, Hiroshi; Li, Tian-Cheng; Koshimoto, Chihiro; Ito, Kaori; Kita, Masakazu; Miyashita, Nobumoto; Arikawa, Jiro; Yagami, Kenichi; Asano, Masahide; Tezuka, Hideo; Suzuki, Noboru; Kurosawa, Tsutomu; Shibahara, Toshiyuki; Furuya, Masato; Mohri, Shirou; Sato, Hiroshi; Ohsawa, Kazutaka; Ibuki, Kentaro; Takeda, Naokazu

2008-07-01

In laboratory animal facilities, monkeys and pigs are used for animal experiments, but the details of hepatitis E virus (HEV) infection in these animals are unknown. The risk of infection from laboratory animals to humans has become a concern; therefore, much attention should be paid to the handling of these animals during their care and use, including surgical procedures performed on infected animals. In this connection, serum samples collected from 916 monkeys and 77 pigs kept in 23 animal facilities belonging to the Japanese Association of Laboratory Animal Facilities of National University Corporations (JALAN) and the Japanese Association of Laboratory Animal Facilities of Public and Private Universities (JALAP) in Japan were examined for the purpose of detecting antibodies to HEV and HEV RNA by using ELISA and RT-PCR, respectively. One hundred and seven serum samples of 916 (11.7%) monkeys were positive for anti-HEV IgG, and 7 and 17 serum samples of 916 (0.8% and 5.3%) monkeys were positive for anti-HEV IgM and IgA, respectively. Thirty-six samples from 62 (58.1%) farm pigs were positive for anti-HEV IgG, whereas all samples tested from miniature pigs were negative (0/15, 0%). Seven samples from 62 (9.1%) farm pigs and 7 samples from 916 (0.8%) monkeys were positive for IgM antibody, but these HEV-IgM antibody positive serum samples were HEV-RNA negative by RT-PCR. The IgM antibody positive rate (9.1%) of farm pigs was much higher than that of monkeys (0.8%). These results suggest the relative levels of risk of HEV infection from these animals to animal handlers and researchers who work with them in laboratory animal facilities.

Changes in Peripheral Serum Creatine Phosphokinase (CPK) and Lactic Dehydrogenase (LDH) in Acute Experimental Colonic Infarction

PubMed Central

Graeber, Geoffrey M.; Wukich, Dane K.; Cafferty, Patrick J.; O'Neill, John F.; Wolf, Robert E.; Ackerman, Norman B.; Harmon, John W.

1981-01-01

No satisfactory laboratory test for the early diagnosis of bowel infarction exists at this time. We have delineated changes in serum CPK levels after acute superior mesenteric artery infarction; whether or not comparable changes occur with inferior mesenteric artery infarction has not yet been determined. Furthermore, the changes in LDH associated with acute bowel infarction have not been documented. To determine the changes in serum CPK and LDH in acute colonic infarction, laparotomies were performed on dogs after peripheral baseline blood samples were drawn and each subject was randomly placed in one of three groups: laparotomy alone, acute colonic obstruction, and acute colonic infarction by ligation of the inferior mesenteric artery. The marginal artery of the colon was ligated at the peritoneal reflection and at the cecum to interrupt arterial collaterals. Blood samples were taken from each subject at intervals of three hours for 48 hours after injury. Serum from each sample was analyzed for total CPK and LDH by automated spectrophotometry. Isoenzymes were determined by agarose gel electrophoresis. Necropsies were conducted on all the dogs to confirm that the intended condition had been produced and that no intercurrent disease was present. The data support the conclusion that total CPK, total LDH and their isoenzymes become elevated in the peripheral serum after colonic infarction. The maximal elevations were all seen within the first 12 hours after acute colonic infarction. Total LDH and LDH3, the most prevalent isoenzyme of LDH in bowel, do not become elevated in the serum to as high a level as CPK, but the combination of serum elevations in both enzyme systems may prove to be of diagnostic significance. PMID:7305484

Comparative analysis of EV isolation procedures for miRNAs detection in serum samples.

PubMed

Andreu, Zoraida; Rivas, Eva; Sanguino-Pascual, Aitana; Lamana, Amalia; Marazuela, Mónica; González-Alvaro, Isidoro; Sánchez-Madrid, Francisco; de la Fuente, Hortensia; Yáñez-Mó, María

2016-01-01

Extracellular vesicles (EVs) are emerging as potent non-invasive biomarkers. However, current methodologies are time consuming and difficult to translate to clinical practice. To analyse EV-encapsulated circulating miRNA, we searched for a quick, easy and economic method to enrich frozen human serum samples for EV. We compared the efficiency of several protocols and commercial kits to isolate EVs. Different methods based on precipitation, columns or filter systems were tested and compared with ultracentrifugation, which is the most classical protocol to isolate EVs. EV samples were assessed for purity and quantity by nanoparticle tracking analysis and western blot or cytometry against major EV protein markers. For biomarker validation, levels of a set of miRNAs were determined in EV fractions and compared with their levels in total serum. EVs isolated with precipitation-based methods were enriched for a subgroup of miRNAs that corresponded to miRNAs described to be encapsulated into EVs (miR-126, miR-30c and miR-143), while the detection of miR-21, miR-16-5p and miR-19a was very low compared with total serum. Our results point to precipitation using polyethylene glycol (PEG) as a suitable method for an easy and cheap enrichment of serum EVs for miRNA analyses. The overall performance of PEG was very similar, or better than other commercial precipitating reagents, in both protein and miRNA yield, but in comparison to them PEG is much cheaper. Other methods presented poorer results, mostly when assessing miRNA by qPCR analyses. Using PEG precipitation in a longitudinal study with human samples, we demonstrated that miRNA could be assessed in frozen samples up to 8 years of storage. We report a method based on a cut-off value of mean of fold EV detection versus serum that provides an estimate of the degree of encapsulation of a given miRNA.

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

DOEpatents

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

1985-08-05

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

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

DOEpatents

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

1988-01-01

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

A bacterial engineered glycoprotein as a novel antigen for diagnosis of bovine brucellosis.

PubMed

Ciocchini, Andrés E; Serantes, Diego A Rey; Melli, Luciano J; Guidolin, Leticia S; Iwashkiw, Jeremy A; Elena, Sebastián; Franco, Cristina; Nicola, Ana M; Feldman, Mario F; Comerci, Diego J; Ugalde, Juan E

2014-08-27

Brucellosis is a highly contagious zoonosis that affects livestock and human beings. Laboratory diagnosis of bovine brucellosis mainly relies on serological diagnosis using serum and/or milk samples. Although there are several serological tests with different diagnostic performance and capacity to differentiate vaccinated from infected animals, there is still no standardized reference antigen for the disease. Here we validate the first recombinant glycoprotein antigen, an N-formylperosamine O-polysaccharide-protein conjugate (OAg-AcrA), for diagnosis of bovine brucellosis. This antigen can be produced in homogeneous batches without the need of culturing pathogenic brucellae; all characteristics that make it appropriate for standardization. An indirect immunoassay based on the detection of anti O-polysaccharide IgG antibodies in bovine samples was developed coupling OAg-AcrA to magnetic beads or ELISA plates. As a proof of concept and to validate the antigen, we analyzed serum, whole blood and milk samples obtained from non-infected, experimentally infected and vaccinated animals included in a vaccination/infection trial performed in our laboratory as well as more than 1000 serum and milk samples obtained from naturally infected and S19-vaccinated animals from Argentina. Our results demonstrate that OAg-AcrA-based assays are highly accurate for diagnosis of bovine brucellosis, even in vaccinated herds, using different types of samples and in different platforms. We propose this novel recombinant glycoprotein as an antigen suitable for the development of new standard immunological tests for screening and confirmatory diagnosis of bovine brucellosis in regions or countries with brucellosis-control programs. Copyright © 2014 Elsevier B.V. All rights reserved.

Epidemiology of Sarcocystis neurona infections in domestic cats (Felis domesticus) and its association with equine protozoal myeloencephalitis (EPM) case farms and feral cats from a mobile spay and neuter clinic.

PubMed

Stanek, J F; Stich, R W; Dubey, J P; Reed, S M; Njoku, C J; Lindsay, D S; Schmall, L M; Johnson, G K; LaFave, B M; Saville, W J A

2003-11-28

Equine protozoal myeloencephalitis (EPM) is a serious neurologic disease in the horse most commonly caused by Sarcocystis neurona. The domestic cat (Felis domesticus) is an intermediate host for S. neurona. In the present study, nine farms, known to have prior clinically diagnosed cases of EPM and a resident cat population were identified and sampled accordingly. In addition to the farm cats sampled, samples were also collected from a mobile spay and neuter clinic. Overall, serum samples were collected in 2001 from 310 cats, with samples including barn, feral and inside/outside cats. Of these 310 samples, 35 were from nine horse farms. Horse serum samples were also collected and traps were set for opossums at each of the farms. The S. neurona direct agglutination test (SAT) was used for both the horse and cat serum samples (1:25 dilution). Fourteen of 35 (40%) cats sampled from horse farms had circulating S. neurona agglutinating antibodies. Twenty-seven of the 275 (10%) cats from the spay/neuter clinic also had detectable S. neurona antibodies. Overall, 115 of 123 (93%) horses tested positive for anti-S. neurona antibodies, with each farm having greater than a 75% exposure rate among sampled horses. Twenty-one opossums were trapped on seven of the nine farms. Eleven opossums had Sarcocystis sp. sporocysts, six of them were identified as S. neurona sporocysts based on bioassays in gamma-interferon gene knockout mice with each opossum representing a different farm. Demonstration of S. neurona agglutinating antibodies in domestic and feral cats corroborates previous research demonstrating feral cats to be naturally infected, and also suggests that cats can be frequently infected with S. neurona and serve as one of several natural intermediate hosts for S. neurona.

Enzyme-linked immunosorbent assay and polymerase chain reaction performance using Mexican and Guatemalan discrete typing unit I strains of Trypanosoma cruzi.

PubMed

Ballinas-Verdugo, Martha; Reyes, Pedro Antonio; Mejia-Dominguez, Ana; López, Ruth; Matta, Vivian; Monteón, Victor M

2011-12-01

Thirteen Trypanosoma cruzi isolates from different geographic regions of Mexico and Guatemala belonging to discrete typing unit (DTU) I and a reference CL-Brener (DTU VI) strain were used to perform enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR). A panel of 57 Mexican serum samples of patients with chronic chagasic cardiopathy and asymptomatic infected subjects (blood bank donors) were used in this study. DNA from the above 14 T. cruzi strains were extracted and analyzed by PCR using different sets of primers designed from minicircle and satellite T. cruzi DNA. The chronic chagasic cardiopathy serum samples were easily recognized with ELISA regardless of the source of antigenic extract used, even with the CL-Brener TcVI, but positive serum samples from blood bank donors in some cases were not recognized by some Mexican antigenic extracts. On the other hand, PCR showed an excellent performance despite the set of primers used, since all Mexican and Guatemalan T. cruzi strains were correctly amplified. In general terms, Mexican, Guatemalan, and CL-Brener T. cruzi strains are equally good sources of antigen when using the ELISA test to detect Mexican serum samples. However, there are some strains with poor performance. The DTU I strains are easily detected using either kinetoplast or satellite DNA target designed from DTU VI strains.

Detection of Francisella tularensis-Specific Antibodies in Patients with Tularemia by a Novel Competitive Enzyme-Linked Immunosorbent Assay

PubMed Central

Sharma, Neekun; Hotta, Akitoyo; Yamamoto, Yoshie; Fujita, Osamu; Uda, Akihiko; Morikawa, Shigeru; Yamada, Akio

2013-01-01

A novel competitive enzyme-linked immunosorbent assay (cELISA) was developed and evaluated for detection of antibodies against Francisella tularensis in humans. The assay is based on the ability of serum antibodies to inhibit the binding of monoclonal antibodies (MAbs) directed against F. tularensis lipopolysaccharide antigens. The assay was evaluated using serum samples of tularemia patients, inactivated F. tularensis-immunized rabbits, and F. tularensis-infected mice. Antibodies against F. tularensis were successfully detected in serum samples of tularemia patients as well as the immunized and infected animals. The cELISA method was compared to indirect ELISA (iELISA) and the commonly used microagglutination test (MA) using serum samples of 19 tularemia patients and 50 healthy individuals. The sensitivity and specificity of cELISA were 93.9 and 96.1%, respectively, in comparison to the iELISA. MA was less sensitive than cELISA with a sensitivity and specificity of only 81.8 and 98.0%, respectively. A high degree of correlation (R2 = 0.8226) was observed between cELISA and iELISA results. The novel cELISA developed in this study appears to be highly sensitive and specific for serodiagnosis of human tularemia. The potential of the MAb-based cELISA to be used in both human and animal samples emphasizes its usefulness for serological survey of tularemia among multiple animal species. PMID:23114700

Rapid detection of EBOLA VP40 in microchip immunofiltration assay

NASA Astrophysics Data System (ADS)

Miethe, Peter; Gary, Dominik; Hlawatsch, Nadine; Gad, Anne-Marie

2015-05-01

In the spring of 2014, the Ebola virus (EBOV) strain Zaire caused a dramatic outbreak in several regions of West Africa. The RT-PCR and antigen capture diagnostic proved to be effective for detecting EBOV in blood and serum. In this paper, we present data of a rapid antigen capture test for the detection of VP40. The test was performed in a microfluidic chip for immunofiltration analysis. The chip integrates all necessary assay components. The analytical sensitivity of the rapid test was 8 ng/ml for recombinant VP40. In serum and whole blood samples spiked with virus culture material, the detection limit was 2.2 x 102 PFU/ml. The performance data of the rapid test (15 min) are comparable to that of the VP40 laboratory ELISA.

Sensitivity and specificity of a nested polymerase chain reaction for detection of lentivirus infection in lions (Panthera leo).

PubMed

Adams, Hayley; van Vuuren, Moritz; Kania, Stephen; Bosman, Anna-Mari; Keet, Dewald; New, John; Kennedy, Melissa

2010-12-01

Feline immunodeficiency virus (FIV) is a lentivirus in the Retroviridae family that causes lifelong infection in domestic cats. The lentivirus of African lions (Panthera leo), referred to as FIVple, is endemic in certain lion populations in eastern and southern Africa. Lentivirus infection leads to immunologic dysfunction and immunosuppressive disease in domestic cats; however, little is known about the pathogenic effects of infection in lions, nor about the epidemiologic impact on free-ranging and captive populations. Whole blood and serum samples were collected opportunistically from free-ranging lions in Kruger National Park, Republic of South Africa (RSA). Whole blood and serum samples were also collected from captive wild lions in the RSA. A nested polymerase chain reaction (PCR) assay for detection of FIV was performed on all whole blood samples. In addition, serum samples were tested for cross-reactive antibodies to domestic feline lentivirus antigens and puma lentivirus synthetic envelope peptide antigen. The PCR assay successfully amplified the lion lentivirus from African lions. The relative sensitivity and relative specificity were 79% and 100%, respectively, and the positive and negative predictive values were 100% and 67%, respectively. This research represents the first study to compare genetic material with antibody-based methods of lentivirus detection on lions in RSA. Using PCR as an additional diagnostic test for FIV in lions will increase screening sensitivity and will allow viral characterization among circulating isolates and monitoring of changes in the viral epidemiology within geographic regions and populations over time.

Monitoring multiple myeloma patients treated with daratumumab: teasing out monoclonal antibody interference.

PubMed

McCudden, Christopher; Axel, Amy E; Slaets, Dominique; Dejoie, Thomas; Clemens, Pamela L; Frans, Sandy; Bald, Jaime; Plesner, Torben; Jacobs, Joannes F M; van de Donk, Niels W C J; Moreau, Philippe; Schecter, Jordan M; Ahmadi, Tahamtan; Sasser, A Kate

2016-06-01

Monoclonal antibodies are promising anti-myeloma treatments. As immunoglobulins, monoclonal antibodies have the potential to be identified by serum protein electrophoresis (SPE) and immunofixation electrophoresis (IFE). Therapeutic antibody interference with standard clinical SPE and IFE can confound the use of these tests for response assessment in clinical trials and disease monitoring. To discriminate between endogenous myeloma protein and daratumumab, a daratumumab-specific immunofixation electrophoresis reflex assay (DIRA) was developed using a mouse anti-daratumumab antibody. To evaluate whether anti-daratumumab bound to and shifted the migration pattern of daratumumab, it was spiked into daratumumab-containing serum and resolved by IFE/SPE. The presence (DIRA positive) or absence (DIRA negative) of residual M-protein in daratumumab-treated patient samples was evaluated using predetermined assessment criteria. DIRA was evaluated for specificity, limit of sensitivity, and reproducibility. In all of the tested samples, DIRA distinguished between daratumumab and residual M-protein in commercial serum samples spiked with daratumumab and in daratumumab-treated patient samples. The DIRA limit of sensitivity was 0.2 g/L daratumumab, using spiking experiments. Results from DIRA were reproducible over multiple days, operators, and assays. The anti-daratumumab antibody was highly specific for daratumumab and did not shift endogenous M-protein. As the treatment of myeloma evolves to incorporate novel monoclonal antibodies, additional solutions will be needed for clinical monitoring of patient responses to therapeutic regimens. In the interim, assays such as DIRA can inform clinical outcomes by distinguishing daratumumab from endogenous M-protein by IFE.

ironPhone: Mobile device-coupled point-of-care diagnostics for assessment of iron status by quantification of serum ferritin.

PubMed

Srinivasan, Balaji; O'Dell, Dakota; Finkelstein, Julia L; Lee, Seoho; Erickson, David; Mehta, Saurabh

2018-01-15

Iron deficiency (ID) is an urgent public health problem that has devastating effects on maternal and child health. However, due to poor access and affordability, screening and diagnosis for ID is often limited to proxy hemoglobin measurements alone. Here, we report the development and validation of ironPhone, a mobile-device coupled portable diagnostics for quantification of serum ferritin concentrations, an iron status biomarker, within a few minutes, from a drop of fingerprick blood. The ironPhone diagnostic platform comprises of a smartphone accessory, an app, and a disposable lateral flow immunoassay test strip to quantify serum ferritin. For initial validation in the lab, we optimized and evaluated the performance of ironPhone with known ferritin concentrations in spiked buffer and serum samples. Following lab validation, we performed a human validation by collecting fingerprick whole blood samples from 20 participants to assess iron status using ironPhone and compared the results with the laboratory standard IMMULITE 2000 analyzer. Findings from the ironPhone for the buffer and spiked serum samples provided a calibration curve with R 2 values of 0.97 (n=27) and 0.93 (n=12), respectively. On comparison with the laboratory standard IMMULITE analyzer in whole blood samples, a correlation of 0.92 (P<0.0001) was observed with a sensitivity of over 90% for predicting ID (ferritin<15.0µg/L) via the ironPhone, demonstrating its promise for iron status assessment at the point-of-care. Copyright © 2017 Elsevier B.V. All rights reserved.

Reference measurement procedure for total glycerides by isotope dilution GC-MS.

PubMed

Edwards, Selvin H; Stribling, Shelton L; Pyatt, Susan D; Kimberly, Mary M

2012-04-01

The CDC's Lipid Standardization Program established the chromotropic acid (CA) reference measurement procedure (RMP) as the accuracy base for standardization and metrological traceability for triglyceride testing. The CA RMP has several disadvantages, including lack of ruggedness. It uses obsolete instrumentation and hazardous reagents. To overcome these problems the CDC developed an isotope dilution GC-MS (ID-GC-MS) RMP for total glycerides in serum. We diluted serum samples with Tris-HCl buffer solution and spiked 200-μL aliquots with [(13)C(3)]-glycerol. These samples were incubated and hydrolyzed under basic conditions. The samples were dried, derivatized with acetic anhydride and pyridine, extracted with ethyl acetate, and analyzed by ID-GC-MS. Linearity, imprecision, and accuracy were evaluated by analyzing calibrator solutions, 10 serum pools, and a standard reference material (SRM 1951b). The calibration response was linear for the range of calibrator concentrations examined (0-1.24 mmol/L) with a slope and intercept of 0.717 (95% CI, 0.7123-0.7225) and 0.3122 (95% CI, 0.3096-0.3140), respectively. The limit of detection was 14.8 μmol/L. The mean %CV for the sample set (serum pools and SRM) was 1.2%. The mean %bias from NIST isotope dilution MS values for SRM 1951b was 0.7%. This ID-GC-MS RMP has the specificity and ruggedness to accurately quantify total glycerides in the serum pools used in the CDC's Lipid Standardization Program and demonstrates sufficiently acceptable agreement with the NIST primary RMP for total glyceride measurement.

Simple and fast analysis of tetrabromobisphenol A, hexabromocyclododecane isomers, and polybrominated diphenyl ethers in serum using solid-phase extraction or QuEChERS extraction followed by tandem mass spectrometry coupled to HPLC and GC.

PubMed

Li, Jian; Chen, Tian; Wang, Yuwei; Shi, Zhixiong; Zhou, Xianqing; Sun, Zhiwei; Wang, Dejun; Wu, Yongning

2017-02-01

Two simplified sample preparation procedures for simultaneous extraction and clean-up of tetrabromobisphenol A, α-, β-, and γ-hexabromocyclododecane and polybrominated diphenyl ethers in human serum were developed and validated. The first procedure was based on solid-phase extraction. Sample extraction, purification, and lipid removal were carried out directly on an Oasis HLB cartridge. The second procedure was a quick, easy, cheap, effective, rugged, and safe-based approach using octadecyl-modified silica particles as a sorbent. After sample extraction and cleanup, tetrabromobisphenol A/hexabromocyclododecane was separated from polybrominated diphenyl ethers by using a Si-based cartridge. Tetrabromobisphenol A and hexabromocyclododecane were then detected by high-performance liquid chromatography coupled to tandem mass spectrometry, while polybrominated diphenyl ethers were detected by gas chromatography coupled to tandem mass spectrometry. The results of the spike recovery test using fetal bovine serum showed that the average recoveries of the analytes ranged from 87.3 to 115.3% with relative standard deviations equal to or lower than 13.4 %. Limits of detection of the analytes were in the range of 0.4-19 pg/mL except for decabromodiphenyl ether. The developed method was successfully applied to routine analysis of human serum samples from occupational workers and the general population. Extremely high serum polybrominated diphenyl ethers levels up to 3.32 × 10 4 ng/g lipid weight were found in occupational workers. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Second trimester serum cortisol and preterm birth: an analysis by timing and subtype.

PubMed

Bandoli, Gretchen; Jelliffe-Pawlowski, Laura L; Feuer, Sky K; Liang, Liang; Oltman, Scott P; Paynter, Randi; Ross, Kharah M; Schetter, Christine Dunkel; Ryckman, Kelli K; Chambers, Christina D

2018-05-24

We hypothesized second trimester serum cortisol would be higher in spontaneous preterm births compared to provider-initiated (previously termed 'medically indicated') preterm births. We used a nested case-control design with a sample of 993 women with live births. Cortisol was measured from serum samples collected as part of routine prenatal screening. We tested whether mean-adjusted cortisol fold-change differed by gestational age at delivery or preterm birth subtype using multivariable linear regression. An inverse association between cortisol and gestational age category (trend p = 0.09) was observed. Among deliveries prior to 37 weeks, the mean-adjusted cortisol fold-change values were highest for preterm premature rupture of the membranes (1.10), followed by premature labor (1.03) and provider-initiated preterm birth (1.01), although they did not differ statistically. Cortisol continues to be of interest as a marker of future preterm birth. Augmentation with additional biomarkers should be explored.

Can urine dipstick predict an elevated serum creatinine?

PubMed

Shah, Kaushal; Kilian, Barbara; Hsieh, Wei-Jen; Kyrillou, Emily; Hedge, Vishal; Newman, David H

2010-06-01

Chart review studies have suggested that point-of-care urine dipstick testing may accurately predict an elevation in serum creatinine (Cr). We aimed to prospectively evaluate the test characteristics of proteinuria/hematuria in predicting elevated serum Cr. A prospective, observational study was conducted between March 2007 and June 2008 at 2 affiliated, urban hospitals with an annual emergency department census of 150,000. Patients undergoing laboratory urinalysis, point-of-care urine dipstick, and a serum chemistry panel were enrolled. Trained research assistants collected data on consecutive patients 18 hours per day using preformatted data forms and entry into an anonymized Access (Microsoft, Seattle, Wash) database. Demographic baseline variables including age, sex, chief complaint, vital signs, and source of sample (catheter vs "clean catch") were also collected. An elevated Cr level was defined as greater than 1.3 based on the laboratory reference range. Standard statistical methods were used to calculate diagnostic test operating characteristics of proteinuria or hematuria as a predictor of elevated serum Cr. Five thousand four hundred sixteen subjects were enrolled with 28.3% male and a mean age of 50.2 years. Elevated serum Cr greater than 1.3 mg/dL was found in 13.9% (755/5416) of subjects. The sensitivity of either proteinuria or hematuria for elevated Cr was 82.5% (95% confidence interval [CI], 80%-85%) and specificity was 34.4% (95% CI, 33%-36%). Positive predictive value was 16.9% (95% CI, 16%-18%) and negative predictive value was 92.4% (95% CI, 91-94%). The likelihood ratio for a positive test was 1.3 (95% CI, 1.1-1.5), and the likelihood ratio for a negative test was 0.5 (95% CI, 0.3-0.8). Although negative predictive value was high, the presence of proteinuria/hematuria was only moderately predictive of elevated serum Cr level. Copyright (c) 2010 Elsevier Inc. All rights reserved.

The use of serum free light chain dimerization patterns assist in the diagnosis of AL amyloidosis.

PubMed

Gatt, Moshe E; Kaplan, Batia; Yogev, Dean; Slyusarevsky, Elana; Pogrebijski, Galina; Golderman, Sizilia; Kukuy, Olga; Livneh, Avi

2018-05-16

The discrimination between benign and malignant forms of plasma cell dyscrasia (PCD) is often difficult. Free light chain monomer-dimer pattern analysis (FLC-MDPA) may assist in solving this dilemma and distinguish between AL amyloidosis and benign PCD. Serum samples of patients with AL amyloidosis and benign PCD were analysed in a blinded manner. Quantitative Western blotting was performed to estimate dimerization and clonality indices, and thereby determine the source of the tested samples, as derived either from benign or malignant PCD. The findings obtained by the FLC-MDPA were compared with the actual diagnosis. Of 37 samples from patients with active AL amyloidosis, 34 (91·9%) fulfilled dimerization criteria for diagnosis of AL amyloidosis. Of the 45 samples from patients with benign PCD, 10 (21·2%) tested falsely positive or gave an inconclusive result. Thus, the sensitivity of the analysis was 92·5% with a remarkable negative predictive value of 91·9%. In addition, of 20 patients who were in complete or very good partial remission, only one tested positive. By multivariate analysis, FLC-MDPA was the best independent marker predicting AL amyloidosis (odds ratio of 84). The FLC-MDPA offers a highly effective tool in the diagnostic assessment of patients with PCD. © 2018 John Wiley & Sons Ltd.

[The lysate and recombinant antigens in ELISA-test-systems for diagnostic of herpes simplex].

PubMed

Ganova, L A; Kovtoniuk, G V; Korshun, L N; Kiseleva, E K; Tereshchenko, M I; Vudmaska, M I; Moĭsa, L N; Shevchuk, V A; Spivak, N Ia

2014-08-01

The lysate and recombinant antigens of various production included informula of ELISA-test-systems were analyzed. The ELISA-test-systems are used for detection of IgG to Herpes simplex virus type I and II. For testing the panel of serums PTH 201 (BBI Inc.) were used. The samples of this panel contain antibodies to Herpes simplex virus type I and II in mixed titers. The 69 serums of donors were used too (17 samples had IgG to Herpes simplex virus type I, 23 samples to Herpes simplex virus type II and 29 samples had no antibodies to Herpes simplex virus). The diagnostic capacity of mixture of recombinant antigens gG1 Herpes simplex virus type I and gG2 Herpes simplex virus type II (The research-and-production complex "DiaprofMed") was comparable with mixture of lysate antigen Herpes simplex virus type I and II (Membrane) EIE Antigen ("Virion Ltd."). In the test-systems for differentiation of IgG to Herpes simplex virus type I the recombinant antigen gG1 Herpes simplex virus type I proved to be comparable with commercial analogue Herpes simplex virus-1 gG1M ("Viral Therapeutics Inc."'). At the same time, capacity to detect IgG to Herpes simplex virus type II in recombinant protein gG2 Herpes simplex virus type II is significantly higher than in its analogue Herpes simplex virus-2 gG2c ("Viral Therapeutics Inc.").

Comparison of a fluorometric assay kit with high-performance liquid chromatography for the assessment of serum retinol concentration.

PubMed

Elom, Aglago Kouassivi; Imane, El Menchawy; Kaoutar, Benjeddou; Khalid, El Kari; Asmaa, El Hamdouchi; Mehdi, Azlaf; Noureddine, El Haloui; Hassan, Aguenaou

2015-06-01

Although high-performance liquid chromatography (HPLC) is the commonly used method for the analysis of retinol in biological samples, simple and rapid test kits are available. This study compared a rapid test kit (ICHECK Fluoro®) to HPLC for the assessment of serum retinol concentrations. For the analysis by HPLC, sample preparation included standard deproteinization and extraction phases. The analysis by ICHECK was performed by injecting serum into IEX reagent vials (n=89) and mixing manually for separation. After precipitation of the proteins, the vial was introduced into the chamber of the ICHECK Fluoro and analysed at 0 min (ICHECK0min) and 15 min later (ICHECK15min). Bland and Altman approach was applied to test the agreement between HPLC and ICHECK. Mean HPLC, ICHECK0min and ICHECK15min values were 421.2±106.0 µg/L, 423.1±118.3 µg/L and 413.2±107.6 µg/L, respectively. Retinol concentrations significantly decreased in the IEX solution over time (p<0.001). No significant proportional bias was observed between HPLC and ICHECK0min (r-0.038, p=0.73) and ICHECK15min (r=-0.024, p=0.82). Fixed biases (HPLC minus ICHECK) for ICHECK0min and ICHECK15min were respectively -1.9±23.1 µg/l (p=0.45) and 8.0±22.7 µg/l (p=0.002). ICHECK Fluoro may offer a reliable mean for assessing serum retinol for measurements performed with no significant time delay.

Evaluation of a commercial ELISA kit for detection of antibodies against Toxoplasma gondii in serum, plasma and meat juice from experimentally and naturally infected sheep

PubMed Central

2013-01-01

Background Toxoplasmosis is one of the most common food borne zoonoses worldwide, and can be a serious life-threatening disease in the congenitally infected fetus and in immunosupressed patients. Among food animals, sheep along with goats and pigs possess the highest incidence of T. gondii cysts in meat, and play a major role as a source of human infection. Methods In this study, a new commercial ELISA kit (PrioCHECK® Toxoplasma Ab SR, Prionics Schlieren-Zurich, Switzerland) for the detection of anti-T. gondii antibodies in serum, plasma and meat juice of sheep, was evaluated by comparing it with the indirect fluorescent antibody test (IFAT), indirect haemagglutination test (IHA) and real-time PCR, on samples from experimentally inoculated and naturally exposed sheep. Results The commercial ELISA detected the infection status in 50% and 100% of sheep orally inoculated with 10,000 T. gondii oocysts (n = 6), from two or three weeks post infection (wpi), respectively, both on serum and plasma samples. Meat juice from all experimentally inoculated sheep collected at slaughter (12 wpi) showed positive ELISA values. In naturally exposed sheep (n = 396), the ELISA showed a very good agreement with IFAT (kappa = 0.91-1.0) and IHA (kappa = 0.96-1.0) performed on serum; and a positive correlation was observed between ELISA values and IFAT titers. By a Receiver Operating Characteristics (ROC) curve analysis, the commercial ELISA had relative sensitivities between 93.33% and 100%, and relative specificities between 96.87% and 100% respect to IFAT and IHA, depending on the considered cut-off value and animal groups tested. Furthermore, the ELISA correctly recognized all animals reacting positive in real-time PCR. The ELISA results on meat juice agreed with those on serum samples in all experimentally inoculated animals, and in 94 out of 96 (97.9%) naturally exposed sheep, when meat juice was tested at a 1:10 dilution. Conclusion The commercial ELISA kit evaluated in this study could represent a valuable tool to improve the surveillance and reporting system for T. gondii in sheep populations at the farm level or for diagnosis at the slaughterhouse, contributing to the control of this widespread zoonosis. PMID:23561035

Quantification of myo-inositol, 1,5-anhydro-D-sorbitol, and D-chiro-inositol using high-performance liquid chromatography with electrochemical detection in very small volume clinical samples

PubMed Central

Schimpf, Karen J.; Meek, Claudia C.; Leff, Richard D.; Phelps, Dale L.; Schmitz, Daniel J.; Cordle, Christopher T.

2015-01-01

Inositol is a six-carbon sugar alcohol and is one of nine biologically significant isomers of hexahydroxycyclohexane. Myo-inositol is the primary biologically active form and is present in higher concentrations in the fetus and newborn than in adults. It is currently being examined for the prevention of retinopathy of prematurity in newborn preterm infants. A robust method for quantifying myo-inositol (MI), D-chiro-inositol (DCI) and 1,5-anhydro-D-sorbitol (ADS) in very small-volume (25 μL) urine, blood serum and/or plasma samples was developed. Using a multiple-column, multiple mobile phase liquid chromatographic system with electrochemical detection, the method was validated with respect to (a) selectivity, (b) accuracy/recovery, (c) precision/reproducibility, (d) sensitivity, (e) stability and (f) ruggedness. The standard curve was linear and ranged from 0.5 to 30 mg/L for each of the three analytes. Above-mentioned performance measures were within acceptable limits described in the Food and Drug Administration’s Guidance for Industry: Bioanalytical Method Validation. The method was validated using blood serum and plasma collected using four common anticoagulants, and also by quantifying the accuracy and sensitivity of MI measured in simulated urine samples recovered from preterm infant diaper systems. The method performs satisfactorily measuring the three most common inositol isomers on 25 μL clinical samples of serum, plasma milk, and/or urine. Similar performance is seen testing larger volume samples of infant formulas and infant formula ingredients. MI, ADS and DCI may be accurately tested in urine samples collected from five different preterm infant diapers if the urine volume is greater than 2–5 mL. PMID:26010453

Detection of antibodies against classical swine fever virus in fecal samples from wild boar.

PubMed

Seo, Sang won; Sunwoo, Sun young; Hyun, Bang hoon; Lyoo, Young S

2012-12-28

Classical swine fever (CSF) is a contagious viral disease that affects pigs. Wild boars can play an important epidemiological role in CSF outbreaks. In the past decades, studies conducted in many countries have reported that the CSF virus (CSFV) may persist in wild boar populations. The existence of CSFV in the free-ranging wild boar populations was indirectly confirmed by determining the prevalence of antibodies against CSFV in the serum of hunted wild boars. However, analyzing sero-prevalence in hunted wild boars to study the risk of CSF outbreaks is difficult due to insufficient number of samples, limitation of hunting area and biased age distribution of hunted wild boars. To improve this survey method, we collected feces of wild boars from their habitat and tested them using CSFV antibody enzyme-linked immunosorbent assay (ELISA) and CSF virus neutralization (VN) test. In this study, ELISA was found to be highly sensitive for detecting antibodies against CSFV in fecal samples. Most of doubtful or positive results obtained in CSFV ELISA were confirmed by VN tests. Despite the high coincidence rate of antibody-positive samples between CSFV ELISA and VN test, the possibility of false positive reaction should be considered. In the regional distribution, a fact that antibody-positive fecal and serum samples were found in geographically close area was shown. Hence, presence of antibodies in fecal samples may provide vital information regarding the risk of CSF outbreaks in wild boar groups in geographical proximity. Copyright © 2012 Elsevier B.V. All rights reserved.

Molecular characterization of chicken infectious anemia virus from contaminated live-virus vaccines.

PubMed

Li, Yang; Hu, Yan; Cui, Shuai; Fu, Jiayuan; Wang, Yixin; Cui, Zhizhong; Fang, Lichun; Chang, Shuang; Zhao, Peng

2017-05-01

The aim of this study was to investigate possible causes of the pervasiveness of chicken infectious anemia virus (CIAV) infection in chickens in recent years in China. A total of 14 batches of live-virus vaccines were examined using PCR to detect CIAV contamination, of which only 2 samples (a Newcastle disease vaccine and a fowl pox vaccine) tested positive for CIAV. These Newcastle and fowl pox vaccines were then inoculated into 1-day-old specific-pathogen-free chickens. Serum samples were collected from chickens infected with the PCR-positive vaccines, and these tested positive for CIAV-specific antibodies as tested using ELISA. In addition, DNA samples isolated from the serum samples also tested positive by PCR. The results indicated that the samples were contaminated with CIAV and identified 2 exogenous CIAV strains, designated CIAV-N22 and CIAV-F10, in the respective samples. The full genome sequences of these novel CIAV strains were sequenced and analyzed. Phylogenetic tree analysis indicated that the CIAV-F10 strain might represent a recombinant viral strain arising from the parental CIAV strains JQ690762 and KJ728816. Overall, the results suggested that vaccination with CIAV-contaminated vaccines contributed to the prevalence and spread of CIAV infection in chickens. Furthermore, the CIAV contaminant was likely subsequently transmitted to commercial chickens through congenital transmission. Our findings therefore highlight the need for more extensive screening of live-virus vaccines for poultry in China to reduce the threat of contamination with exogenous viruses. © 2016 Poultry Science Association Inc.

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

PubMed

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

2014-07-01

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

Porcine reproductive and respiratory syndrome virus (PRRSV) surveillance using pre-weaning oral fluid samples detects circulation of wild-type PRRSV.

PubMed

Kittawornrat, Apisit; Panyasing, Yaowalak; Goodell, Christa; Wang, Chong; Gauger, Phillip; Harmon, Karen; Rauh, Rolf; Desfresne, Luc; Levis, Ian; Zimmerman, Jeffrey

2014-01-31

Oral fluid samples collected from litters of piglets (n=600) one day prior to weaning were evaluated as a method to surveil for porcine reproductive and respiratory syndrome virus (PRRSV) infections in four sow herds of approximately 12,500 sow each. Serum samples from the litters' dam (n=600) were included for comparison. All four herds were endemically infected with PRRSV and all sows had been vaccinated ≥ 2 times with PRRSV modified-live virus vaccines. After all specimens had been collected, samples were randomized and assayed by PRRSV real-time reverse transcription polymerase chain reaction (RT-qPCR) and four PRRSV antibody ELISA assays (IgM, IgA, IgG, and Commercial Kit). All sow serum samples were negative by PRRSV RT-qPCR, but 9 of 600 oral fluid samples tested positive at two laboratories. Open reading frame 5 (ORF5) sequencing of 2 of the 9 positive oral fluid samples identified wild-type viruses as the source of the infection. A comparison of antibody responses in RT-qPCR positive vs. negative oral fluid samples showed significantly higher IgG S/P ratios in RT-qPCR-positive oral fluid samples (mean S/P 3.46 vs. 2.36; p=0.02). Likewise, sow serum samples from RT-qPCR-positive litter oral fluid samples showed significantly higher serum IgG (mean S/P 1.73 vs. 0.98; p<0.001) and Commercial Kit (mean S/P 1.97 vs. 0.98; p<0.001) S/P ratios. Overall, the study showed that pre-weaning litter oral fluid samples could provide an efficient and sensitive approach to surveil for PRRSV in infected, vaccinated, or presumed-negative pig breeding herds. Copyright © 2014 Elsevier B.V. All rights reserved.

Competitive and blocking enzyme-linked immunoassay for detection of fetal bovine serum antibodies to bovine viral diarrhea virus.

PubMed

Katz, J B; Hanson, S K

1987-02-01

A competitive blocking enzyme-linked immunoassay (CELIA) was developed to detect bovine viral diarrhea virus (BVDV) antibodies in undiluted fetal bovine serum (FBS). The CELIA was based on competition of serum BVDV antibodies with biotin-labelled anti-BVDV immunoglobulins (Ig) for a limited quantity of solid-phase BVDV antigen. Antigen preparation was simple, FBS could be tested undiluted, and detergent-containing washes were unnecessary. A series of dilutions of postnatal bovine BVDV antiserum prepared in FBS and a set of 147 undiluted abbatoir FBS samples were tested by both CELIA and serum neutralization tests (SNT). CELIA results on both sets of specimens correlated positively with SNT titers (r = 0.99 and r = 0.85). Relative to the SNT, CELIA sensitivity was 100%; specificity was 76%. CELIA detected a level of BVDV antibody below the 1:2-titer threshold detectable with the SNT. Advantages, limitations, and theoretical differences between the CELIA and SNT are discussed. A similar comparison of CELIA with non-competitive enzyme-linked immunoassay approaches to BVDV serodiagnosis is made. It is concluded that the CELIA is valuable in selecting only BVDV-seronegative FBS for use in virologic cell culture media.

Association between serum uric acid levels and obesity among university students (China).

PubMed

Duan, Ying; Liang, Wei; Zhu, Lijun; Zhang, Ting; Wang, Linghong; Nie, Zhognhua; Chen, Yan; He, Lianping; Jin, Yuelong; Yao, Yingshui

2015-06-01

To evaluate the association between serum uric acid and obesity among university students who participated in routine health screening in 2013. In this cross-sectional study, 3529 subjects were analyzed. Obesity categories were classified by BMI levels references in China. And serum uric acid levels were classified by serum uric acid quartiles. Two-sample T-test and Wilcoxon Rank sum test were used to compare age, biochemical and anthropometric parameters of subjects of two genders. Rank correlation used to analyze relationship between serum uric acid and obesity. There were 1285 males (mean age, 19.8 ± 1.3 years) and 2244 females (mean age, 19.9 ± 1.3 years) in this study. Association between 2nd serum uric acid quartile and normal in male are significant and coefficient was 0.519. The 3rd serum uric acid quartile and normal in female was associated significantly (r = 0.173, p = 0.010). And associations between overweight and 3rd and 4th serum uric acid quartiles in female were significant (r = 0.128, p = 0.038 in 1st quartile and r = 0.282, p = 0.004 in 4th quartile). The 4th serum uric acid quartile and Obesity in two gender groups were significantly associated (r = 0.291, p = 0.000 in male and r = 0.484, p = 0.001 in female). High serum uric acid was positively associated with obesity in overweight and obesity group. However, the association was weak between two variables because serum uric acid influenced obesity with other related factors together. Copyright AULA MEDICA EDICIONES 2014. Published by AULA MEDICA. All rights reserved.

Evaluation of serum sialic acid, fucose levels and their ratio in oral squamous cell carcinoma.

PubMed

Chinnannavar, Sangamesh Ningappa; Ashok, Lingappa; Vidya, Kodige Chandrashekhar; Setty, Sunil Mysore Kantharaja; Narasimha, Guru Eraiah; Garg, Ranjana

2015-01-01

Detection of cancer at the early stage is of utmost importance to decrease the morbidity and mortality of the disease. Apart from the conventional biopsy, minimally invasive methods like serum evaluation are used for screening large populations. Thus, this study aimed to estimate serum levels of sialic acid and fucose and their ratio in oral cancer patients and in healthy control group to evaluate their role in diagnosis. Serum samples were collected from 52 healthy controls (group I) and 52 squamous cell carcinoma patients (group II). Estimation of serum levels of sialic acid and fucose and their ratio was performed. This was correlated histopathologically with the grades of carcinoma. Statistical analysis was done by using analysis of variance (ANOVA) test and unpaired "t" test. Results showed that serum levels of sialic acid and fucose were significantly higher in oral cancer patients compared to normal healthy controls (P < 0.001). The sialic acid to fucose ratio was significantly lower in cancer patients than in normal controls (P < 0.01). However, comparison with histological grading, habits, gender, and age group did not show any significant result. The mean serum sialic acid and fucose levels showed an increasing trend from controls to malignant group and their corresponding ratio showed decreasing trend from controls to malignant group. The ratio of sialic acid to fucose can be a useful diagnostic aid for oral cancer patients.

Eating a healthy lunch improves serum alanine aminotransferase activity.

PubMed

Iwamoto, Masako; Yagi, Kaori; Yazumi, Kayoko; Komine, Airi; Shirouchi, Bungo; Sato, Masao

2013-09-14

Nutritional guidance and diet control play important roles in the treatment of obesity and non-alcoholic fatty liver. However, in Japan, nutritional guidance is difficult to provide in practice. Therefore, we evaluated the effects of providing the 'once-a-day' intervention of a healthy lunch on various metabolic parameters. For a 1-month preparatory period, 10 subjects generally consumed the lunches that were provided by the worksite cafeteria. This was followed by a 1-week washout period, after which, the subjects consumed healthy, low-calorie, well-balanced lunches for a 1-month test period. After the preparatory and test periods, blood samples were obtained from all subjects. The serum levels of indices relevant to metabolic syndrome and fatty liver were measured. Serum alanine aminotransferase activity significantly decreased by 20.3% after the healthy intervention. However, the indices of metabolic syndrome did not significantly change. Analysis of the relationship between serum alanine aminotransferase activity and nutrient content indicated that the improvement of serum alanine aminotransferase status was due to the higher vegetable content and lower animal-source protein of the meals provided. In summary, the 'once-a-day' intervention of providing a healthy lunch improved serum alanine aminotransferase status. A diet high in vegetables and low in animal-based protein is important in maintaining a healthy condition.

Serologic evidence of influenza A(H1N1)pdm09 virus in northern sea otters

USGS Publications Warehouse

Li, Zhu-Nan; Ip, Hon S.; Frost, Jessica F.; White, C. LeAnn; Murray, Michael J.; Carney, Paul J.; Sun, Xiang-Jie; Stevens, James; Levine, Min Z.; Katz, Jacqueline M.

2014-01-01

Sporadic epizootics of pneumonia among marine mammals have been associated with multiple animal-origin influenza A virus subtypes (1–6); seals are the only known nonhuman host for influenza B viruses (7). Recently, we reported serologic evidence of influenza A virus infection in free-ranging northern sea otters (Enhydra lutris kenyoni) captured off the coast of Washington, USA, in August 2011 (8). To investigate further which influenza A virus subtype infected these otters, we tested serum samples from these otters by ELISA for antibody-binding activity against 12 recombinant hemagglutinins (rHAs) from 7 influenza A hemagglutinin (HA) subtypes and 2 lineages of influenza B virus (Technical Appendix Table 1). Estimated ages for the otters were 2–19 years (Technical Appendix Table 2); we also tested archived serum samples from sea otters of similar ages collected from a study conducted during 2001–2002 along the Washington coast (9).

Detection of Candida albicans Sap2 in cancer patient serum samples by an indirect competitive enzyme-linked immunosorbent assay for the diagnosis of candidiasis.

PubMed

Wang, Yicun; Gao, Xiang; Zhi Gang, J U; Liu, Jingyuan; Dong, Shuai; Wang, Li

2013-01-01

The secreted aspartyl proteinases 2 (Sap2) of Candida albicans (C. albicans) is a potential marker of candididasis. It is a virulence factor associated with adherence and tissue invasion. In order to detect Sap2 in clinical sera, we developed an indirect competitive enzyme-linked immunosorbent assay (ELISA). Polyclonal antibodies were produced for Sap2 by injecting Sap2 into a New Zealand White inbred rabbit. They could be used at a dilution exceeding 1:1200 in an indirect ELISA, and detected Sap2 concentration up to 1 ng/mL. Of the 286 cancer serum samples tested, 16.8% were found as candidiasis. The test was simple and economical to perform and had a level of sensitivity for detection of low-titer positive sera; thus, it may be proven to be of value in epidemiological studies on candidiasis.

[Value of positive auto controls in the gel centrifugation method].

PubMed

Eichler, H; Kretschmer, V

1994-01-01

We studied 97 samples of patients being positive in the autocontrol of the indirect antiglobulin test (IAT) in the gel system (DiaMed). In 83.2%, retesting with monospecific anti-IgG serum gave also positive results, due to a specific phenomenon caused, for example, by drug-specific antibodies (AB), warm auto-AB or allo-AB. In contrast, only 52.9% of the samples retested by the standard tube technique with polyspecific antiglobulin serum reacted positive. Only in 6 patients slightly increased cold agglutinins could be detected. None of the investigated patients showed any clinical or laboratory signs of hemolysis except one with pernicious anemia. We conclude that positive results of the autocontrol in the gel IAT should be confirmed by an additional DAT in the tube technique. If this second test shows a negative result, transfusions can take place without any restrictions.

Comparative evaluation of recombinant LigB protein and heat-killed antigen-based latex agglutination test with microscopic agglutination test for diagnosis of bovine leptospirosis.

PubMed

Nagalingam, Mohandoss; Thirumalesh, Sushma Rahim Assadi; Kalleshamurthy, Triveni; Niharika, Nakkala; Balamurugan, Vinayagamurthy; Shome, Rajeswari; Sengupta, Pinaki Prasad; Shome, Bibek Ranjan; Prabhudas, Krishnamsetty; Rahman, Habibur

2015-10-01

This study aimed to develop latex agglutination test (LAT) using recombinant leptospiral immunoglobulin-like protein (LigB) (rLigB) antigen and compare its diagnostic efficacy with LAT using conventional heat-killed leptospiral antigen and microscopic agglutination test (MAT) in diagnosing bovine leptospirosis. The PCR-amplified 1053-bp ligB gene sequences from Leptospira borgpetersenii Hardjo serovar were cloned in pET 32 (a) vector at EcoRI and NotI sites and expressed in BL21 E. coli cells as fusion protein with thioredoxin (-57 kDa) and characterized by SDS-PAGE and immunoblot. Out of 390 serum samples [cattle (n = 214), buffaloes (n = 176)] subjected to MAT, 115 samples showed reciprocal titre≥100 up to 1600 against one or more serovars. For recombinant LigB protein/antigen-based LAT, agglutination was observed in the positive sample, while no agglutination was observed in the negative sample. Similarly, heat-killed leptospiral antigen was prepared from and used in LAT for comparison with MAT. A two-sided contingency table was used for analysis of LAT using both the antigens separately against MAT for 390 serum samples. The sensitivity, specificity and positive and negative predictive values of recombinant LigB LAT were found to be 75.65, 91.27, 78.38 and 89.96 %, respectively, and that of heat-killed antigen-based LAT were 72.17, 89.82, 74.77 and 88.53 %, respectively, in comparison with MAT. This developed test will be an alternative/complementary to the existing battery of diagnostic assays/tests for specific detection of pathogenic Leptospira infection in bovine population.

Human brucellosis among pyrexia of unknown origin cases and occupationally exposed individuals in Goa Region, India.

PubMed

Pathak, Ajay D; Dubal, Zunjar B; Doijad, Swapnil; Raorane, Abhay; Rodrigues, Savio; Naik, Rajeshwar; Naik-Gaonkar, Shraddha; Kalorey, Dewanand R; Kurkure, Nitin V; Naik, Rajesh; Barbuddhe, Sukhadeo B

2014-01-01

Brucellosis is a widespread zoonotic infection. This disease is endemic in many parts of Asia, including India. Brucellosis is a major cause of pyrexia of unknown origin (PUO). Persons exposed to infected animals or contaminated animal products are at high risk. Seropositivity among animal handlers, veterinarians and dairy workers has been documented in India. Thus, the present study was aimed to determine prevalence of brucellosis among PUO cases and occupationally exposed individuals. In this study, serum samples (n=282) from cases of pyrexia of unknown origin (PUO) (n=243), and occupationally exposed individuals (n=39) were collected and tested for brucellosis by Rose Bengal plate test (RBPT), serum agglutination test (SAT), indirect ELISA, IgG and IgM ELISA. Blood culture for isolation of Brucella was performed for 10 serologically positive patients using BACTEC 9050 automated blood culture system. Biochemical tests and PCR techniques were used for confirmation of the isolates. Of the samples tested, 4.25%, 3.54%, 6.02% and 4.96% samples were positive by RBPT, SAT, indirect ELISA and IgG ELISA, respectively. None of the sample was positive for IgM ELISA. Of the 10 blood samples cultured bacteriologically, one Brucella isolate was recovered. The isolate was confirmed as Brucella abortus. Amplification of the bcsp31 and IS711 genes was also observed. Seropositivity for brucellosis was observed among PUO cases, animal handlers and dairy workers in Goa, India. The serological tests showed variable results. One Brucella isolate was obtained by performing blood culture. Confirmation of the case was done rapidly using molecular tools. General awareness about clinical symptoms should be increased which will improve proper diagnosis within short time frame.

The prevalence of low serum zinc and copper levels and dietary habits associated with serum zinc and copper in 12- to 36-month-old children from low-income families at risk for iron deficiency.

PubMed

Schneider, Julie M; Fujii, Mary L; Lamp, Catherine L; Lönnerdal, Bo; Zidenberg-Cherr, Sheri

2007-11-01

Iron and zinc share common food sources, and children at risk of iron deficiency may also develop zinc deficiency. We determined the prevalence of zinc and copper deficiency and examined factors associated with serum zinc and copper in young children from low-income families at risk of iron deficiency. A cross-sectional study design was used to assess serum zinc and copper, along with an interview-assisted survey to assess factors associated with serum zinc and copper in a convenience sample. Participants were 435 children aged 12 to 36 months recruited from select clinics of the Special Supplemental Nutrition Program for Women, Infants, and Children in Contra Costa and Tulare Counties, California. Frequencies were used to report prevalence. Multiple linear regressions were conducted to examine factors associated with serum zinc and copper, controlling for age, sex, and ethnicity. The prevalence of low serum zinc level (<70 microg/dL [<10.7 micromol/L]) was 42.8%, and low serum copper level (<90 microg/dL [<14.2 micromol/L]) was <1%. Mean+/-standard deviation of serum copper was 150+/-22 microg/dL (23.6+/-3.5 micromol/L) and 140+/-24 microg/dL (22.1+/-3.8 micromol/L) for anemic and non-anemic children, respectively (t test, P=0.026). In multiple linear regression consumption of sweetened beverages was negatively associated with serum zinc level, and consumption of >15 g/day meat was positively associated with serum zinc level, whereas current consumption of breast milk and >15 g/day beans were positively associated with serum copper level. The prevalence of low serum zinc concentration in the sample was high, and warrants further investigation amongst vulnerable populations.

Cross sectional, comparative study of serum erythropoietin, transferrin receptor, ferritin levels and other hematological indices in normal pregnancies and iron deficiency anemia during pregnancy.

PubMed

Sharma, Jai B; Bumma, Sirisha D; Saxena, Renu; Kumar, Sunesh; Roy, Kallol K; Singh, Neeta; Vanamail, P

2016-08-01

To test the correlation of the serum erythropoietin levels, serum transferrrin receptor levels and serum ferritin levels along with other hematological parameters in normal pregnant and anemic pregnant patients. In a prospective study, 120 pregnant women were recruited between 18 and 36 weeks of gestation; 53 normal pregnant patients, 67 anemic pregnant patients, in which, 17 had mild, 30 had moderate anemia, 20 had severe anemia. A blood sample was taken. The various hematological parameters, hemoglobin (Hb), mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), total iron binding capacity (TIBC), serum ferritin, percentage saturation of iron, serum erythropoietin (SEPO) levels, serum transferrin receptors (STfRS) were performed. For statistics, Student's 't' test, Pearson's Chi test, Mann Whitney test and Bartlett test were used as per data. MCV was significantly reduced in anemic pregnancies as compared to non-anemic pregnancies (80.2±9.6 vs 94.12±9.8fl, p=0.001), MCHC was also reduced in them (30.2±3.38% vs 34.2±2.33%, p=0.176), TIBC was significantly increased in anemic pregnancies (343.31±28.54% vs 322.88±23.84%, p=0.001), serum ferritin was significantly reduced (24.9±10.48μg/L vs 31.03±9.98μg/L, p=0.001), percentage saturation of iron was also reduced (53.85±13.21% vs 62.04±15.79%, p=0.0024), serum erythropoietin levels were significantly higher in anemic women (26.24±26.61mU/ml vs 18.12±19.08mU/ml, p=0.064). The levels were significantly higher in severe anemia (46.5±46.8mU/ml than in moderate anemia 27.4±28.1mU/ml and mild anemia 22.8±22.8mU/ml. Serum transferrin receptors were significantly higher in anemic pregnancies than in non-anemic pregnancies (1.40±0.0802μg/ml vs 1.08±0.641μg/ml, p=0.019) with rise being higher in severe anemia (2.28±0.986μg/ml) than in moderate (1.4±0.816μg/ml) and mild anemia (1.16±0.702μg/ml). Various hematological parameters especially sTfR, serum erythropoietin, serum ferritin and sTfR/log ferritin levels correlate with the severity of anemia. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Studies on equine recurrent uveitis. II: The role of infection with Leptospira interrogans serovar pomona.

PubMed

Halliwell, R E; Brim, T A; Hines, M T; Wolf, D; White, F H

1985-10-01

An enzyme linked immunosorbent assay was developed for the detection of immunoglobulin class specific antibodies to Leptospira interrogans serovar pomona in the serum and aqueous humor of horses. Serum antibody was also assayed by microscopic agglutination tests. Although higher levels of antibody were found in sera from horses with signs of uveitis, the association was not statistically significant. Antibodies to pomona were detected in the aqueous of 12 eyes from the 101 horses sampled at a slaughterhouse, and in most instances, a comparison of the aqueous/serum antibody level with that of the total aqueous/serum IgG level indicated intraocular antibody synthesis. Antibodies were also found in 4 aqueous (or vitreous) samples out of 9 obtained from horses with clinically documented uveitis and the above comparison again indicated intraocular antibody synthesis. The data point to an important role for pomona as an etiology of equine recurrent uveitis but also emphasize that the initiating cause for this disease is often obscure in that association with leptospirosis cannot be shown in many instances.

Analysis of some chosen elements of cerebrospinal fluid and serum in amyotrophic lateral sclerosis patients by total reflection X-ray fluorescence

NASA Astrophysics Data System (ADS)

Ostachowicz, B.; Lankosz, M.; Tomik, B.; Adamek, D.; Wobrauschek, P.; Streli, C.; Kregsamer, P.

2006-11-01

Trace elements play an important role in the human central nervous system. Significant variations of the concentration of trace elements in body fluids may occur in neurodegenerative diseases. In the present work an investigation of the elemental composition of the serum, and the cerebrospinal fluid in amyotrophic lateral sclerosis patients and a control group was performed. For the analysis of the body fluids Total reflection X-ray Fluorescence (TXRF) spectrometry was used. The samples were taken during routine diagnostic procedures. Na, Mg, Cl, K, Ca, Cu, Zn, and Br were determined in both fluids. In order to validate the results of analysis a serum standard reference material was measured. A t-test was applied to check if the mean concentrations of the elements are different for ALS and the control group. For the serum samples higher values for Br were found in the ALS group, for the cerebrospinal fluid lower values of Na, Mg and Zn as well as higher Ca values were found in the ALS group compared to the control group.

Evaluation of bilirubin interference and accuracy of six creatinine assays compared with isotope dilution-liquid chromatography mass spectrometry.

PubMed

Nah, Hyunjin; Lee, Sang-Guk; Lee, Kyeong-Seob; Won, Jae-Hee; Kim, Hyun Ok; Kim, Jeong-Ho

2016-02-01

The aim of this study was to estimate bilirubin interference and accuracy of six routine methods for measuring creatinine compared with isotope dilution-liquid chromatography mass spectrometry (ID-LC/MS). A total of 40 clinical serum samples from 31 patients with serum total bilirubin concentration >68.4μmol/L were collected. Serum creatinine was measured using two enzymatic reagents and four Jaffe reagents as well as ID-LC/MS. Correlations between bilirubin concentration and percent difference in creatinine compared with ID-LC/MS were analyzed to investigate bilirubin interference. Bias estimations between the six reagents and ID-LC/MS were performed. Recovery tests using National Institute of Standards and Technology (NIST) Standard Reference Material (SRM) 967a were also performed. Both the enzymatic methods showed no bilirubin interference. However, three of the four Jaffe methods demonstrated significant bilirubin concentration-dependent interference in samples with creatinine levels <53μmol/L, and two of them showed significant bilirubin interference in samples with creatinine levels ranging from 53.0 to 97.2μmol/L. Comparison of these methods with ID-LC/MS using patients' samples with elevated bilirubin revealed that the tested methods failed to achieve the bias goal at especially low levels of creatinine. In addition, recovery test using NIST SRM 967a showed that bias in one Jaffe method and two enzymatic methods did not achieve the bias goal at either low or high level of creatinine, indicating they had calibration bias. One enzymatic method failed to achieve all the bias goals in both comparison experiment and recovery test. It is important to understand that both bilirubin interference and calibration traceability to ID-LC/MS should be considered to improve the accuracy of creatinine measurement. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Peste des petits ruminants infection in domestic ruminants in Sudan.

PubMed

Intisar, K S; Ali, Y H; Haj, M A; Sahar, M A T; Shaza, M M; Baraa, A M; Ishag, O M; Nouri, Y M; Taha, K M; Nada, E M; Ahmed, A M; Khalafalla, A I; Libeau, G; Diallo, A

2017-04-01

The existence of peste des petits ruminants (PPR) in domestic ruminants and camels in Sudan during 2008-2012 was investigated. Lung tissues and serum samples were randomly collected from sheep, goats, cattle, and camels at different areas of Sudan. A total of 12,384 serum samples were collected from clinically healthy 7413 sheep, 1988 camels, 1501 cattle, 1459 goats, and 23 gazelles at different areas in the Sudan. They were examined for PPR antibodies using competitive ELISA (cELISA). The overall detected seroprevalence of PPR in tested sera was 49.4%; seroprevalence values within species were 67.1, 48.2, 25.8, 2.1, and 21.7% in sheep, goat, cattle, camels, and gazelles, respectively. The highest seroprevalence (68.1%) was observed in sera collected from Darfur states, then the central states (54.3%). A total of 1276 lung tissue samples (623 sheep, 324 cattle, 220 camels, and 109 goats) were collected. The majority of lung samples were collected from clinically healthy animals that showed lesions on PM in slaughterhouses (95%) and during PPR outbreaks; samples were tested for PPR antigen using immunocapture ELISA (IcELISA). PPR antigen was detected in 233 out of the 1276 tested samples (18.3%). Positive results were observed in samples collected from clinically healthy and diseased animals. The observed prevalence values in each species were 33.6, 21.1, 15.4, and 12.3% in camel, goat, sheep, and cattle, respectively. PPR antigen was detected in samples from different areas; however, the highest prevalence (63.9%) was found in samples collected from the eastern states, then Khartoum state (28%). Trials for virus isolation were done in different cell cultures. Out of 30 IcELISA-positive samples inoculated in primary bovine and ovine kidney cells, Vero cells, the PPR virus was successfully isolated from 15 (eight sheep, five camels, and two goats) samples in the three cell culture types. Using RT-PCR, PPRV nucleic acid was detected in all 25 IcELISA-positive tested samples.

Celiac disease in non-clinical populations of Japan.

PubMed

Fukunaga, Mai; Ishimura, Norihisa; Fukuyama, Chika; Izumi, Daisuke; Ishikawa, Nahoko; Araki, Asuka; Oka, Akihiko; Mishiro, Tomoko; Ishihara, Shunji; Maruyama, Riruke; Adachi, Kyoichi; Kinoshita, Yoshikazu

2018-02-01

Celiac disease is a chronic autoimmune enteropathy caused by gluten ingestion. While its prevalence in Western countries is reported to be as high as 1%, the prevalence has not been evaluated in a large-scale study of a Japanese population. The aim of our study was to clarify the possible presence of celiac disease in a Japanese non-clinical population as well as in patients showing symptoms suggestive of the disease. Serum samples were collected from 2008 non-clinical adults and 47 patients with chronic unexplained abdominal symptoms between April 2014 and June 2016. The anti-tissue transglutaminase (TTG) immunoglobulin A antibody titer was determined as a screening test for celiac disease in all subjects, and individuals with a value of >2 U/mL subsequently underwent testing for the presence of serum endomysial IgA antibody (EMA) as confirmation. Those testing positive for EMA or with a high concentration (>10 U/mL) of TTG were further investigated by histopathological examinations of duodenal mucosal biopsy specimens and HLA typing tests. Of the 2008 non-clinical adults from whom serum samples were collected, 161 tested positive for TTG, and all tested negative for EMA. Four subjects who had a high TTG titer were invited to undergo confirmatory testing, and the histopathological results confirmed the presence of celiac disease in only a single case (0.05%). Of the 47 symptomatic patients, one (2.1%) was found to have a high TTG titer and was diagnosed with celiac disease based on duodenal histopathological findings. The presence of celiac disease in a non-clinical Japanese population was low at 0.05% and was rarely found in patients with unexplained chronic abdominal symptoms.

Relationship between Serum Iron Profile and Blood Groups among the Voluntary Blood Donors of Bangladesh.

PubMed

Hoque, M M; Adnan, S D; Karim, S; Al-Mamun, M A; Faruki, M A; Islam, K; Nandy, S

2016-04-01

Blood donation results in a substantial iron loss and subsequent mobilization from body stores. Chronic iron deficiency is a well-recognized complication of regular blood donation. The present study conducted to compare the level of serum ferritin, serum iron, total iron binding capacity (TIBC) and percentage transferrin saturation in different ABO and Rhesus type blood groups among the voluntary blood donors of Bangladesh. The present prospective study included 100 healthy voluntary donors attending at Department of Blood Transfusion, Dhaka Medical College, Dhaka between the periods of July 2013 to Jun 2014. From each donor 10mL venous blood sample was taken and divided into heparinized and non-heparinized tubes for determination of hemoglobin (Hb), hematocrit (Hct), serum iron (SI), total iron binding capacity (TIBC) and serum ferritin by standard laboratory methods. Percentage of transferrin saturation (TS) calculated from serum iron and TIBC. Data were analyzed with SPSS (version 16) software and comparisons between groups were made using student's t-test and one way ANOVA. In the present study mean±SD of age of the respondents was 27.2±6.5 years with a range of 18 to 49 years and 81.0% were male and 19.0% were female. Among the donors 18.0% had blood group A, 35.0% had blood group B, 14.0% had blood group AB and 33.0% had blood group O. Among the donors 91.0% had rhesus positive and 9.0% had rhesus negative. Donors with blood group O had lowest haemoglobin, serum iron and transferring saturation levels. Donors with blood group A had highest TIBC level. Donors with blood group B had lowest serum ferritin level. An independent samples 't' test showed statistically significant difference in serum ferritin and percentage transferrin saturation between blood group AB and blood group O and in percentage transferrin saturation between blood group B and blood group O. One way ANOVA showed that there is no significant difference in haemoglobin, serum iron, serum ferritin and percentage transferring saturation in different ABO and Rh blood grouping categories. Blood donors with blood group O had lowest haemoglobin, serum iron and transferring saturation levels and donors with blood group A had highest TIBC level. Blood donors with blood group B had lowest serum ferritin level. The understanding of the different blood groups ability to retain iron in their system can give an insight into their ability to handle the disease iron deficiency anaemia.

The Antimicrobial Peptide Lysozyme Is Induced after Multiple Trauma

PubMed Central

Klüter, Tim; Fitschen-Oestern, Stefanie; Lippross, Sebastian; Weuster, Matthias; Pufe, Thomas; Tohidnezhad, Mersedeh; Beyer, Andreas; Seekamp, Andreas; Varoga, Deike

2014-01-01

The antimicrobial peptide lysozyme is an important factor of innate immunity and exerts high potential of antibacterial activity. In the present study we evaluated the lysozyme expression in serum of multiple injured patients and subsequently analyzed their possible sources and signaling pathways. Expression of lysozyme was examined in blood samples of multiple trauma patients from the day of trauma until 14 days after trauma by ELISA. To investigate major sources of lysozyme, its expression and regulation in serum samples, different blood cells, and tissue samples were analysed by ELISA and real-time PCR. Neutrophils and hepatocytes were stimulated with cytokines and supernatant of Staphylococcus aureus. The present study demonstrates the induction and release of lysozyme in serum of multiple injured patients. The highest lysozyme expression of all tested cells and tissues was detected in neutrophils. Stimulation with trauma-related factors such as interleukin-6 and S. aureus induced lysozyme expression. Liver tissue samples of patients without trauma show little lysozyme expression compared to neutrophils. After stimulation with bacterial fragments, lysozyme expression of hepatocytes is upregulated significantly. Toll-like receptor 2, a classic receptor of Gram-positive bacterial protein, was detected as a possible target for lysozyme induction. PMID:25258475

Evaluation of immunochromatographic test (ICT) strips for the serological detection of Babesia bovis and Babesia bigemina infection in cattle from Western Java, Indonesia.

PubMed

Guswanto, Azirwan; Allamanda, Puttik; Mariamah, Euis Siti; Munkjargal, Tserendorf; Tuvshintulga, Bumduuren; Takemae, Hitoshi; Sivakumar, Thillaiampalam; AbouLaila, Mahmoud; Terkawi, Mohamad Alaa; Ichikawa-Seki, Madoka; Nishikawa, Yoshifumi; Yokoyama, Naoaki; Igarashi, Ikuo

2017-05-30

Three types of immunochromatographic test (ICT) strips were prepared for the detection of an antibody response against spherical body protein 4 (SBP-4) of Babesia bovis (bovICT), C-terminal-truncated rhoptry-associated protein 1 (rRAP1/CT17) of B. bigemina (bigICT), and the combination of both proteins (dual-ICT). The evaluation of their performance was conducted using a confirmed positive and negative serum panel for B. bovis and B. bigemina. Together with ELISA, the ICT strips were applied to determine the seroprevalence of bovine babesiosis in Western Java, Indonesia. Among 991 serum samples, 28.4%, 25.3%, and 24.5% of cattle were detected to be seropositive to B. bovis infection using ELISA, bovICT, and dual-ICT, respectively. B. bigemina seropositive was detected in 27.1%, 24.2%, and 22.8% of samples using ELISA, bigICT, and dual-ICT, respectively. The comparison of ICT strips and ELISA results using field serum samples showed good agreement with Kappa values >0.7 between all methods The application of ICT strips is preferable in the field situations where rapid diagnosis is required. Furthermore, the data showed the current seroprevalence of bovine babesiosis in Western Java, Indonesia, and efficient control strategies are needed to reduce economic losses due to the disease. Copyright © 2017 Elsevier B.V. All rights reserved.

Reciprocal contribution of clinical studies and the HP10 antigen ELISA for the diagnosis of extraparenchymal neurocysticercosis.

PubMed

Parkhouse, R Michael E; Carpio, Arturo; Campoverde, Alfredo; Sastre, Patricia; Rojas, Glenda; Cortez, María Milagros

2018-02-01

To evaluate diagnosis of active neurocysticercosis, paired cerebral spinal fluid (CSF) and serum samples from 24 neurocysticercosis (NCC) patients and 17 control neurological patients were assayed in the HP10 Taenia antigen (Ag) ELISA. The CSF samples were also tested with an HP10 Lateral Flow Assay (LFA). The HP10 Ag was detected by ELISA in the CSF of 5/5 patients with Definitive extraparenchymal NCC, and in 4/5 of the corresponding sera. In the Definitive parenchymal group, on the other hand, the HP10 Ag was absent in 2/3 CSF (with a very low value in the one positive sample) and all the corresponding serum samples. Samples of CSF from 4/7 patients in the Probable parenchymal group, were also significantly HP10 Ag positive, suggesting the presence of extraparenchymal cysts not identified by the imaging studies. With the possible exception of one patient, the corresponding serum samples of the Probable parenchymal NCC group, were all HP10 Ag negative. Samples of CSF from 9 NCC patients diagnosed with Mixed parenchymal and extraparenchymal NCC were all significantly HP10 Ag positive, confirming the presence of extraparenchymal cysts, with only 7/9 of the corresponding serum samples being HP10 positive. Thus detection of the HP10 Ag indicates extraparenchymal and not parenchymal cyst localization and is more sensitive with CSF than serum. Three neurological patients clinically diagnosed as subarachnoid cyst, hydrocephalus and tuberculoma, respectively, were clearly positive for HP10 Ag. Of these, two were confirmed as NCC by subsequent imaging; the third died prior to further examination. Thus, a total of 8 patients had their clinical diagnosis questioned. Finally, there was good agreement between the HP10 Ag ELISA and LFA with CSF samples giving an optical density ≥0.4 in the ELISA assay. In conclusion, the HP10 Ag assay should provide a valuable and reciprocal tool in the clinical diagnosis and follow up of extraparenchymal NCC. Copyright © 2017 Elsevier B.V. All rights reserved.

An Immunoassay to Rapidly Measure Acetaminophen Protein Adducts Accurately Identifies Patients With Acute Liver Injury or Failure.

PubMed

Roberts, Dean W; Lee, William M; Hinson, Jack A; Bai, Shasha; Swearingen, Christopher J; Stravitz, R Todd; Reuben, Adrian; Letzig, Lynda; Simpson, Pippa M; Rule, Jody; Fontana, Robert J; Ganger, Daniel; Reddy, K Rajender; Liou, Iris; Fix, Oren; James, Laura P

2017-04-01

A rapid and reliable point-of-care assay to detect acetaminophen protein adducts in the serum of patients with acute liver injury could improve diagnosis and management. AcetaSTAT is a competitive immunoassay used to measure acetaminophen protein adducts formed by toxic metabolites in serum samples from patients. We compared the accuracy of AcetaSTAT vs high-pressure liquid chromatography with electrochemical detection (HPLC-EC; a sensitive and specific quantitative analytic assay) to detect acetaminophen protein adducts. We collected serum samples from 19 healthy individuals (no liver injury, no recent acetaminophen use), 29 patients without acetaminophen-associated acute liver injury, and 33 patients with acetaminophen-associated acute liver injury participating in the Acute Liver Failure Study Group registry. Each serum sample was analyzed by AcetaSTAT (reported as test band amplitude) and HPLC-EC (the reference standard). We also collected data on patient age, sex, weight, level of alanine aminotransferase on test day and peak values, concentration of acetaminophen, diagnoses (by site investigator and causality review committee), and outcome after 21 days. Differences between groups were analyzed using the Fisher exact test for categoric variables and the Kruskal-Wallis test or rank-sum test for continuous variables. AcetaSTAT discriminated between patients with and without acetaminophen-associated acute liver injury; the median AcetaSTAT test band amplitude for patients with acetaminophen-associated acute liver injury was 584 (range, 222-1027) vs 3678 (range, 394-8289) for those without (P < .001). AcetaSTAT identified patients with acetaminophen-associated acute liver injury with 100% sensitivity, 86.2% specificity, a positive predictive value of 89.2%, and a negative predictive value of 100%. Results from AcetaSTAT were positive in 4 subjects who received a causality review committee diagnosis of non-acetaminophen-associated acute liver injury; HPLC-EC and biochemical profiles were consistent with acetaminophen-associated acute liver injury in 3 of these cases. The competitive immunoassay AcetaSTAT shows a high degree of concordance with HPLC-EC results in identifying patients with acetaminophen-associated acute liver injury. This rapid and simple assay could increase early detection of this disorder and aid clinical management. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

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

PubMed

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

2011-04-01

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

High serum selenium levels are associated with impaired fasting glucose and elevated fasting serum glucose in Linyi, China.

PubMed

Li, Zhe; Li, Xia; Ju, Wen; Wu, Guanrui; Yang, Xiaomei; Fu, Xiaofeng; Gao, Xibao

2018-01-01

The relationship between selenium level and impaired fasting glucose or elevated fasting serum glucose remains controversial. This study aimed to evaluate these associations in China. This observational population study adopted a cluster sampling approach to enroll participants. Baseline information on selenium categories was tested using one-way analysis of variance and Kruskal-Wallis equality-of-populations rank tests. Multivariable logistic regression was used to investigate the association between serum selenium level and impaired fasting glucose or elevated fasting serum glucose. The mean serum selenium concentration was 121.5μg/L which in a relatively high baseline Se status. Differences were observed among individuals with normal, impaired fasting glucose and elevated fasting serum glucose levels in their basic information, physical examination results and laboratory findings. After adjusting for their basic information, physical examination results and laboratory findings, compared with the low-selenium group, the high-selenium groups (124.9-143.9 and above 143.9μg/L) had ORs for elevated fasting serum glucose of 2.31 (1.37-3.90) and 2.67 (1.59-4.48), respectively (both P<0.05). A sex-difference was observed, and a significant association between selenium levels and impaired fasting glucose was observed for males but not for females. The findings of this observational study suggest that relatively high selenium levels might be positively associated with elevated fasting serum glucose and relatively high selenium levels might be positively associated with impaired fasting glucose in men. Copyright © 2017 Elsevier GmbH. All rights reserved.

Optimization and proficiency testing of a pseudovirus-based assay for detection of HIV-1 neutralizing antibody in China.

PubMed

Nie, Jianhui; Wang, Wenbo; Wen, Zhiheng; Song, Aijing; Hong, Kunxue; Lu, Shan; Zhong, Ping; Xu, Jianqing; Kong, Wei; Li, Jingyun; Shang, Hong; Ling, Hong; Ruan, Li; Wang, Youchun

2012-11-01

Among the neutralizing antibody evaluation assays, the single-cycle pseudovirus infection assay is high-throughput and can provide rapid, sensitive and reproducible measurements after a single cycle of infection. Cell counts, pseudovirus inoculation levels, amount of diethylaminoethyl-dextran (DEAE-dextran), and the nonspecific effects of serum and plasma were tested to identify the optimal conditions for a neutralizing antibody assay based on pseudoviruses. Optimal conditions for cell counts, pseudovirus inoculation, and amount of DEAE-dextran were 1 × 10(4)cells/well, 200TCID(50)/well, and 15 μg/ml, respectively. Compared with serum samples, high-concentration anticoagulants reduced the relative light unit (RLU) value. The RLU value increased sharply initially but then decreased slowly with dilution of the plasma sample. Test kits containing 10 HIV-1 CRF07/08_BC pseudovirus strains and 10 plasma samples from individuals infected with HIV-1 CRF07/08_BC were assembled into two packages and distributed to nine laboratories with a standard operating procedure included. For the 10 laboratories that evaluated the test, 17 of 44 (37%) laboratory pairs were considered equivalent. A statistical qualification rule was developed based on the testing results from 5 experienced laboratories, where a laboratory qualified if at least 83% of values lied within the acceptable range. Copyright © 2012 Elsevier B.V. All rights reserved.

Salivary diagnosis of measles: a study of notified cases in the United Kingdom, 1991-3.

PubMed Central

Brown, D. W.; Ramsay, M. E.; Richards, A. F.; Miller, E.

1994-01-01

OBJECTIVES--To validate a method for salivary diagnosis of measles and to assess the diagnostic accuracy of notified cases of measles. DESIGN--Blood and saliva samples were collected within 90 days of onset of symptoms from patients clinically diagnosed as having measles and tested for specific IgM by antibody capture radioimmunoassay. SETTING--17 districts in England and one in southern Ireland during August 1991 to February 1993. SUBJECTS--236 children and adults with measles notified by a general practitioner. RESULTS--Specific IgM was detected in serum in only 85 (36%) of the 236 cases. In cases associated with outbreaks and tested within six weeks of onset, 53/57 (93%) of samples were IgM positive, thereby confirming the sensitivity of serum IgM detection as a marker of recent infection. The serological confirmation rate was lower in cases with a documented history of vaccination (13/87; 15%) than in those without (70/149; 47%) and varied with age, being lowest in patients under a year, of whom only 4/36 (11%) were confirmed. Measles specific IgM was detected in 71/77 (92%) of adequate saliva samples collected from patients with serum positive for IgM. In cases where measles was not confirmed, 6/101 had rubella specific IgM and 5/132 had human parvovirus B19 specific IgM detected in serum. CONCLUSIONS--The existing national surveillance system for measles, which relies on clinically diagnosed cases, lacks the precision required for effective disease control. Saliva is a valid alternative to serum for IgM detection, and salivary diagnosis could play a major role in achieving measles elimination. Rubella and parvovirus B19 seem to be responsible for a minority of incorrectly diagnosed cases of measles in the United Kingdom and other infectious causes of measles-like illness need to be sought. PMID:8167513

Effectiveness of chamomile tea on glycemic control and serum lipid profile in patients with type 2 diabetes.

PubMed

Rafraf, M; Zemestani, M; Asghari-Jafarabadi, M

2015-02-01

This study aimed at assessing the effects of chamomile tea consumption on glycemic control and serum lipid profile in patients with type 2 diabetes mellitus (T2DM). This single-blind randomized controlled clinical trial was conducted on 64 individuals with T2DM (males and females) aged between 30 and 60 years. The intervention group (n = 32) consumed chamomile tea (3 g/150 mL hot water) three times per day immediately after meals for 8 weeks. The control group (n = 32) followed a water regimen for the same intervention period. Fasting blood samples, anthropometric measurements, and 3-day, 24-h dietary recalls were collected at the baseline and at the end of the trial. Data were analyzed by independent t test, paired t test, Pearson correlation test, and analysis of covariance. Chamomile tea significantly decreased concentration of HbA1C (p = 0.03), serum insulin levels (p < 0.001), homeostatic model assessment for insulin resistance (p < 0.001), total cholesterol (p = 0.001), triglyceride (p < 0.001), and low-density lipoprotein cholesterol (p = 0.05) compared with control group. No significant changes were shown in serum high-density lipoprotein cholesterol levels in both groups. Chamomile tea has some beneficial effects on glycemic control and serum lipid profile in T2DM patients.

Selective precipitation reaction: a novel diagnostic test for tissue pathology in Atlantic salmon, Salmo salar, infected with salmonid alphavirus (SAV3).

PubMed

Braceland, M; Tinsley, J; Cockerill, D; Bickerdike, R; McLoughlin, M F; Eckersall, P D

2017-08-01

While investigating biomarkers for infection with salmonid alphavirus (SAV), the cause of pancreas disease (PD), a selective precipitation reaction (SPR) has been discovered in serum which could be an on-farm qualitative test and an in-laboratory quantitative assay for health assessments in aquaculture. Mixing serum from Atlantic salmon, Salmo salar, with SAV infection with a sodium acetate buffer caused a visible precipitation which does not occur with serum from healthy salmon. Proteomic examination of the precipitate has revealed that the components are a mix of muscle proteins, for example enolase and aldolase, along with serum protein such as serotransferrin and complement C9. The assay has been optimized for molarity, pH, temperature and wavelength so that the precipitation can be measured as the change in optical density at 340 nm (Δ 340 ). Application of the SPR assay to serum samples from a cohabitation trial of SAV infection in salmon showed that the Δ 340 in infected fish rose from undetectable to a maximum at 6 weeks post-infection correlating with histopathological score of pancreas, heart and muscle damage. This test may have a valuable role to play in the diagnostic evaluation of stock health in salmon. © 2016 The Authors. Journal of Fish Diseases Published by John Wiley & Sons Ltd.

Reproducibility of biomarkers in induced sputum and in serum from chronic smokers.

PubMed

Zuiker, Rob G J A; Kamerling, Ingrid M C; Morelli, Nicoletta; Calderon, Cesar; Boot, J Diderik; de Kam, Marieke; Diamant, Zuzana; Burggraaf, Jacobus; Cohen, Adam F

2015-08-01

Soluble inflammatory markers obtained from non-invasive airway sampling such as induced sputum may be useful biomarkers for targeted pharmaceutical interventions. However, before these soluble markers can be used as potential targets, their variability and reproducibility need to be established in distinct study populations. This study aimed to assess the reproducibility of biomarkers obtained from induced sputum and serum in chronic smokers and non-smokers. Sputum and serum samples were obtained from 16 healthy non-smokers and 16 asymptomatic chronic smokers (for both groups: 8M/8F, 30-52 years, FEV1 ≥80% pred.; ≥10 pack years for the smokers) on 2 separate visits 4-10 days apart. Soluble markers in serum and sputum were analysed by ELISA. The differences between smokers vs non-smokers were analysed with a t-test and variability was assessed on log-transformed data by a mixed model ANOVA. Analysable sputum samples could be obtained from all 32 subjects. In both study populations neutrophils and macrophages were the predominant cell types. Serum Pulmonary Surfactant Associated Protein D had favourable reproducibility criteria for reliability ratio (0.99), intra-subject coefficient of variation (11.2%) and the Bland Altman limits of agreement. Furthermore, chronic smokers, compared to non-smokers, had significantly higher sputum concentrations of IL-8 (1094.6 pg/mL vs 460.8 pg/mL, p = 0.006)), and higher serum concentrations of Pulmonary Surfactant Associated Protein D (110.9 pg/mL vs 64.7 pg/mL, p = 0.019), and lower concentrations of Serum Amyloid A (1352.4 pg/mL vs 2297.5 pg/mL, p = 0.022). Serum Pulmonary Surfactant Associated Protein D proved to be a biomarker that fulfilled the criteria for reproducibility in both study groups. Copyright © 2015 Elsevier Ltd. All rights reserved.

Prevalence of pernicious anemia in patients with macrocytic anemia and low serum B12

PubMed Central

AA, Abdulmanea; AH, Alsaeed; AP, Shaik; FH, AlGahtani

2014-01-01

Objective: The current research evaluated the prevalence of pernicious anemia (PA) in patients with macrocytic anemia (high MCV) and low serum B12 in Riyadh. Methods: Blood testing was done in 77 patients (males: 45.5%, females: 54.5%) with macrocytic anemia; 84 patients; (males: 23.8%, females: 76.2%) with low serum B12 and 30 healthy subjects. Complete blood count, differential count, folic acid, vitamin B12, intrinsic factor, gastric parietal cell antibodies and holotranscobalamin II were assessed. Results: A total of five subjects from 161 patients had PA; three of these patients had macrocyticanemia (3.90%) and two patients had low serum B12 (2.38%). Significant differences (p<0.05) in some hematological, immunological, biochemical parameters were found in subjects with macrocytic anemia and low serum B12 compared to controls. Conclusions: Pernicious anemia in patients with macrocytic anemia and low serum B12 was for the selected sample size can be assumed to be uncommon in Riyadh, Saudi Arabia. PMID:25674111

Investigation of a mumps outbreak among university students with two measles-mumps-rubella (MMR) vaccinations, Virginia, September-December 2006.

PubMed

Rota, J S; Turner, J C; Yost-Daljev, M K; Freeman, M; Toney, D M; Meisel, E; Williams, N; Sowers, S B; Lowe, L; Rota, P A; Nicolai, L A; Peake, L; Bellini, W J

2009-10-01

Following the clinical diagnosis of the first case of mumps on September 22, 2006 at the University of Virginia (UVA), 52 suspected cases were identified through active surveillance for mumps by the end of December 2006. Samples were collected from 47 students who presented with parotitis despite a documented history of two doses of measles, mumps, and rubella (MMR) vaccine. Six of 47 serum samples (13%) were positive for mumps IgM, and 46/47 specimens were positive for mumps IgG. Endpoint titration of acute phase serum samples from laboratory-confirmed cases did not provide evidence that elevated serum IgG is a consistent marker for infection among cases due to secondary vaccine failure. Buccal swab samples from 39 of the 47 students were tested by real-time reverse transcription-polymerase chain reaction (RT-PCR) and/or viral culture. Mumps virus or mumps RNA was detected in 12 of 39 buccal samples (31%). Genetic analysis of the virus from the outbreak at UVA indicated that the outbreak was not linked to the large mumps outbreak in the Midwestern US that occurred earlier in 2006. Our findings support the use of viral detection to improve laboratory diagnosis of mumps among persons who have received two doses of MMR.

A Human Serum-Based Enzyme-Free Continuous Glucose Monitoring Technique Using a Needle-Type Bio-Layer Interference Sensor

PubMed Central

Seo, Dongmin; Paek, Sung-Ho; Oh, Sangwoo; Seo, Sungkyu; Paek, Se-Hwan

2016-01-01

The incidence of diabetes is continually increasing, and by 2030, it is expected to have increased by 69% and 20% in underdeveloped and developed countries, respectively. Therefore, glucose sensors are likely to remain in high demand in medical device markets. For the current study, we developed a needle-type bio-layer interference (BLI) sensor that can continuously monitor glucose levels. Using dialysis procedures, we were able to obtain hypoglycemic samples from commercial human serum. These dialysis-derived samples, alongside samples of normal human serum were used to evaluate the utility of the sensor for the detection of the clinical interest range of glucose concentrations (70–200 mg/dL), revealing high system performance for a wide glycemic state range (45–500 mg/dL). Reversibility and reproducibility were also tested over a range of time spans. Combined with existing BLI system technology, this sensor holds great promise for use as a wearable online continuous glucose monitoring system for patients in a hospital setting. PMID:27669267

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

PubMed

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

2015-03-01

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

Micro-bead injection spectroscopy for label-free automated determination of immunoglobulin G in human serum.

PubMed

Ramos, Inês I; Magalhães, Luís M; Barreiros, Luisa; Reis, Salette; Lima, José L F C; Segundo, Marcela A

2018-01-01

Immunoglobulin G (IgG) represents the major fraction of antibodies in healthy adult human serum, and deviations from physiological levels are a generic marker of disease corresponding to different pathologies. Therefore, screening methods for IgG evaluation are a valuable aid to diagnostics. The present work proposes a rapid, automatic, and miniaturized method based on UV-vis micro-bead injection spectroscopy (μ-BIS) for the real-time determination of human serum IgG with label-free detection. Relying on attachment of IgG in rec-protein G immobilized in Sepharose 4B, a bioaffinity column is automatically assembled, where IgG is selectively retained and determined by on-column optical density measurement. A "dilution-and-shoot" approach (50 to 200 times) was implemented without further sample treatment because interferences were flushed out of the column upon sample loading, with minimization of carryover and cross-contamination by automatically discarding the sorbent (0.2 mg) after each determination. No interference from human serum albumin at 60 mg mL -1 in undiluted sample was found. The method allowed IgG determination in the range 100-300 μg mL -1 (corresponding to 5.0-60 mg mL -1 in undiluted samples), with a detection limit of 33 μg mL -1 (1.7 mg mL -1 for samples, dilution factor of 50). RSD values were < 9.4 and < 11.7%, for intra and inter-assay precision, respectively, while recovery values for human serum spiked with IgG at high pathological levels were 97.8-101.4%. Comparison to commercial ELISA kit showed no significant difference for tested samples (n = 8). Moreover, time-to-result decreased from several hours to < 5 min and analysis cost decreased 10 times, showing the potential of the proposed approach as a point-of-care method. Graphical abstract Micro-Bead Injection Spectroscopy method for real time, automated and label-free determination of total serum human Immunoglobulin G (IgG). The method was designed for Lab-on-Valve (LOV) platforms using a miniaturised protein G bioaffinity separative approach. IgG are separated from serum matrix components upon quantification with low non-specific binding in less than 5 min.

Frequency distribution of hepatitis C virus genotypes in different geographical regions of Pakistan and their possible routes of transmission.

PubMed

Idrees, Muhammad; Riazuddin, Sheikh

2008-05-23

Information regarding hepatitis C virus genotypes and subtypes circulating in Pakistan and various risk factors for their transmission are not known well. The specific objective of this study was to find out the frequency of various HCV genotypes present in well-characterized Pakistani HCV isolates and their possible routes of transmission. A total of 3351 serum samples were tested by type-specific genotyping assay. Out of 3351 HCV RNA positive patients, 2039 were males and 1312 were females. As regard as genotyped samples, 2165 belonged to Punjab region, 823 belonged to N.W.F.P., 239 to Sindh and 124 patients were from Balochistan. Out of the total 3351 tested serum samples, type-specific PCR fragments were observed in 3150 (94.00%) serum samples. The distribution of genotypes of the typeable samples as determined by this assay, was as follows: 1664 (49.05%) genotype 3a; 592 (17.66%) genotype 3b; 280 (8.35%) genotype 1a; 252 (7.52%) genotype 2a; 101 (3.01%) genotype 1b; 50 (1.49%) with genotype 4; 25 (0.75%) with 3c; 27 (0.80%) genotype 2b; 6 (0.18%) with subtype 5a; 5 (0.15%) genotype 1c; 4 (0.12%) with subtype 6a; 3 (0.09%) genotype 2c; and 161 (4.80%) patients were infected with mixed infection. Two hundred and one (5.99%) serum samples were found untypeable by the present genotyping system. More than 86% and 72% patients with genotypes 3a and 3b respectively had received multiple injections in past. For genotypes 1a and 1b the route of transmission was major/minor surgery along with unknown reasons. Majority of the cases with type 2a, 2b and indeterminate genotypes were sporadic. Mixed infections were common in thalassaemic patients. The most common HCV genotype in Pakistan is type 3a. Regional difference in genotypes was observed only in Balochistan province of Pakistan. More than 70% of the cases were acquired in hospitals through reuse of needles/syringes and major/minor surgery that is very common in this country.

Antineuronal antibodies in idiopathic achalasia and gastro-oesophageal reflux disease

PubMed Central

Moses, P L; Ellis, L M; Anees, M R; Ho, W; Rothstein, R I; Meddings, J B; Sharkey, K A; Mawe, G M

2003-01-01

Background and aims: The precise aetiology of achalasia is unknown although autoimmunity has been implicated and is supported by several studies. We screened sera from patients with achalasia or gastro-oesophageal reflux disease (GORD) to test for circulating antimyenteric neuronal antibodies. Methods: Serum was obtained from 45 individuals with achalasia, 16 with GORD, and 22 normal controls. Serum was used in immunohistochemistry to label whole mount preparations of ileum and oesophagus of the guinea pig and mouse. Also, sections of superior cervical and dorsal root ganglia, and spinal cord were examined. Results: Positive immunostaining of the myenteric plexus was detected in significantly more achalasia and GORD samples than control samples (achalasia, p<0.001; GORD, p<0.01), and immunoreactivity was significantly more intense with achalasia and GORD serum samples than controls (achalasia, p<0.01; GORD, p<0.05). There was no correlation between intensity of immunoreactivity and duration of achalasia symptoms. In most cases, achalasia and GORD sera stained all ileal submucosal and myenteric neurones, and oesophageal neurones. Immunostaining was not species specific; however, immunostaining was largely specific for enteric neurones. Western blot analysis failed to reveal specific myenteric neuronal proteins that were labelled by antibodies in achalasia or GORD serum. Conclusions: These data suggest that antineuronal antibodies are generated in response to tissue damage or some other secondary phenomenon in achalasia and GORD. We conclude that antineuronal antibodies found in the serum of patients with achalasia represent an epiphenomenon and not a causative factor. PMID:12692044

Evaluation of recombinant porin (rOmp2a) protein as a potential antigen candidate for serodiagnosis of Human Brucellosis.

PubMed

Pathak, Prachi; Kumar, Ashu; Thavaselvam, Duraipandian

2017-07-11

Brucellosis is an important zoonotic disease caused by different Brucella species and human brucellosis is commonly prevalent in different states of India. Among various Brucella species, B. melitensis is most pathogenic to human and included as category B biothreat which can cause infection through aerosol, cut, wounds in skin and contact with infected animals. The diagnosis of human brucellosis is very important for proper treatment and management of disease as there is no vaccine available for human use. The present study was designed to clone, express and purify immunodominant recombinant omp2a (rOmp2a) porin protein of B. melitensis and to evaluate this new antigen candidate for specific serodiagnosis of human brucellosis by highly sensitive iELISA (indirect enzyme linked immunosorbent assay). Omp2a gene of B. melitensis 16 M strain was cloned and expressed in pET-SUMO expression system. The recombinant protein was purified under denaturing conditions using 8 M urea. The purified recombinant protein was confirmed by western blotting by reacting with anti-HIS antibody. The sero-reactivity of the recombinant protein was also checked by reacting with antisera of experimentally infected mice with B. melitensis 16 M at different time points. Serodiagnostic potential of recombinant porin antigen was tested against 185 clinical serum samples collected from regions endemic to brucellosis in southern part of India by iELISA. The samples were grouped into five groups. Group 1 contained cultured confirmed positive serum samples of brucellosis (n = 15), group 2 contained sera samples from positive cases of brucellosis previously tested by conventional methods of RBPT (n = 28) and STAT (n = 26), group 3 contained sera samples negative by RBPT(n = 36) and STAT (n = 32), group 4 contained sera samples of other febrile illness and PUO case (n = 35) and group 5 contained confirmed negative sera samples from healthy donors (n = 23). The rOmp2a was found to be immunoreactive by iELISA and western blotting. The test showed a sensitivity of 93.75% and specificity of 95.83% when tested against 185 serum samples. For determination of statistical significance between experimental groups and control groups, Student's t test was performed on the data. Omp2a emerges as a potential antigen candidate for serodiagnosis of human brucellosis.

Changes in erythropoietin levels during space flight or space flight simulation

NASA Technical Reports Server (NTRS)

Dunn, C. D. R.; Hen, J. P.

1980-01-01

Two hundred and seventy samples from 24 subjects involved in 3 bedrest studies and from 3 subjects involved in Spacelab Mission Development Test 3 were assayed for erythropoietin (Ep), in an in vitro fetal mouse liver cell assay, and for ferritin using a commercially available immunoradiometric assay kit. No trends or significant changes in serum Ep were observed. Serum ferritin concentrations tended to increases slightly during the 'missions', reflecting a redirection of iron from the suppressed erythron into iron stores.

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

PubMed

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

2014-11-04

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

Baseline Assessment of 25-Hydroxyvitamin D Reference Material and Proficiency Testing/External Quality Assurance Material Commutability: A Vitamin D Standardization Program Study.

PubMed

Phinney, Karen W; Sempos, Christopher T; Tai, Susan S-C; Camara, Johanna E; Wise, Stephen A; Eckfeldt, John H; Hoofnagle, Andrew N; Carter, Graham D; Jones, Julia; Myers, Gary L; Durazo-Arvizu, Ramon; Miller, W Greg; Bachmann, Lorin M; Young, Ian S; Pettit, Juanita; Caldwell, Grahame; Liu, Andrew; Brooks, Stephen P J; Sarafin, Kurtis; Thamm, Michael; Mensink, Gert B M; Busch, Markus; Rabenberg, Martina; Cashman, Kevin D; Kiely, Mairead; Galvin, Karen; Zhang, Joy Y; Kinsella, Michael; Oh, Kyungwon; Lee, Sun-Wha; Jung, Chae L; Cox, Lorna; Goldberg, Gail; Guberg, Kate; Meadows, Sarah; Prentice, Ann; Tian, Lu; Brannon, Patsy M; Lucas, Robyn M; Crump, Peter M; Cavalier, Etienne; Merkel, Joyce; Betz, Joseph M

2017-09-01

The Vitamin D Standardization Program (VDSP) coordinated a study in 2012 to assess the commutability of reference materials and proficiency testing/external quality assurance materials for total 25-hydroxyvitamin D [25(OH)D] in human serum, the primary indicator of vitamin D status. A set of 50 single-donor serum samples as well as 17 reference and proficiency testing/external quality assessment materials were analyzed by participating laboratories that used either immunoassay or LC-MS methods for total 25(OH)D. The commutability test materials included National Institute of Standards and Technology Standard Reference Material 972a Vitamin D Metabolites in Human Serum as well as materials from the College of American Pathologists and the Vitamin D External Quality Assessment Scheme. Study protocols and data analysis procedures were in accordance with Clinical and Laboratory Standards Institute guidelines. The majority of the test materials were found to be commutable with the methods used in this commutability study. These results provide guidance for laboratories needing to choose appropriate reference materials and select proficiency or external quality assessment programs and will serve as a foundation for additional VDSP studies.

Tocopherol in Elder Self-Neglect

NASA Technical Reports Server (NTRS)

Aung, K.; Burnett, J.; Dyer, C.; Smith, S. M.

2009-01-01

Although elder self-neglect is the most common form of elder mistreatment, its pathophysiology is not well understood. Alpha-tocopherol is a lipid soluble antioxidant required for the preservation of cell membranes. Since the association between tocopherol and cognitive impairment in older adults has been described, we explored the possibility of its role in elder self-neglect. OBJECTIVE: (1) To determine whether serum tocopherol levels are associated with elder self-neglect, and (2) to assess the association of serum tocopherol levels and cognitive function in elder self-neglect. METHODS: Serum tocopherol levels were measured in a cohort of 67 self-neglecting elders and 67 matched controls, recruited for the Consortium for Research in Elder Self-neglect of Texas. Pearson s correlation tests were performed to assess bivariate associations between serum tocopherol levels and cognitive function. RESULTS: Mean serum alpha-tocopherol levels were 10.8 +/- 4.7 ug/mL in self-neglect group and 13.0 +/- 4.9 ug/mL in control group (p = 0.006, unpaired student s t-test). None of the participants from either group had alpha-tocopherol level lower than the reference range. Mean serum gamma-tocopherol levels were 2.0 +/- 1.0 ug/mL in self-neglect group and 2.0 +/- 1.1 in control group (p=0.83). Proportion of the elders with gamma-tocopherol level lower than the reference range were 4.5% (3/66) in self-neglect group and 10.4% (7/67) in control group (p=0.32, Fisher s Exact Test). Among the self-neglecting elders, no association was found between serum alpha-tocopherol levels and the Mini-Mental State Examination (MMSE) or the Wolf-Klein Clock Drawing Test (CDT) scores (r =-0.42, p=0.75 for MMSE; r=0.08, p=0.54 for CDT). No association was found between serum gamma-tocopherol levels and the MMSE or the CDT (r=-0.12, p=0.35 for MMSE; r=0.05, p=0.68 for CDT). CONCLUSION: In our sample, neither alpha-tocopherol nor gamma-tocopherol appears to have a role in pathophysiology of elder self-neglect. Nevertheless, significantly lower serum alpha-tocopherol levels in the self neglect group merits further research.

Active B12: a rapid, automated assay for holotranscobalamin on the Abbott AxSYM analyzer.

PubMed

Brady, Jeff; Wilson, Lesley; McGregor, Lynda; Valente, Edward; Orning, Lars

2008-03-01

Conventional tests for vitamin B(12) deficiency measure total serum vitamin B12, whereas only that portion of vitamin B12 carried by transcobalamin (holotranscobalamin) is metabolically active. Measurement of holotranscobalamin (holoTC) may be more diagnostically accurate for detecting B(12) deficiency that requires therapy. We developed an automated assay for holoTC that can be used on the Abbott AxSYM immunoassay analyzer. AxSYM Active B12 is a 2-step sandwich microparticle enzyme immunoassay. In step 1, a holoTC-specific antibody immobilized onto latex microparticles captures holoTC in samples of serum or plasma. In step 2, the captured holoTC is detected with a conjugate of alkaline phosphatase and antiTC antibody. Neither apoTC nor haptocorrin exhibited detectable cross-reactivity. The detection limit was < or = 0.1 pmol/L. Within-run and total imprecision (CV ranges) were 3.4%-5.1% and 6.3%-8.5%, respectively. Assay CVs were < 20% from at least 3 pmol/L to 107 pmol/L. With diluted serum samples, measured concentrations were 104%-114% of the expected values in the working range of the assay. No interference from bilirubin, hemoglobin, triglycerides, erythrocytes, rheumatoid factor, or total protein was detected at expected (abnormal) concentrations. A comparison of the AxSYM Active B12 assay with a commercial RIA for holoTC yielded the regression equation: AxSYM = 0.98RIA + 4.7 pmol/L (S(y x), 11.4 pmol/L; n = 204). Assay throughput was 45 tests/h. A 95% reference interval of 19-134 pmol/L holoTC was established with samples from 292 healthy individuals. The AxSYM Active B12 assay allows rapid, precise, sensitive, specific, and automated measurement of human holoTC in serum and plasma.

Sensitivity and specificity of a real-time reverse transcriptase polymerase chain reaction detecting feline coronavirus mutations in effusion and serum/plasma of cats to diagnose feline infectious peritonitis.

PubMed

Felten, Sandra; Leutenegger, Christian M; Balzer, Hans-Joerg; Pantchev, Nikola; Matiasek, Kaspar; Wess, Gerhard; Egberink, Herman; Hartmann, Katrin

2017-08-02

Feline coronavirus (FCoV) exists as two pathotypes, and FCoV spike gene mutations are considered responsible for the pathotypic switch in feline infectious peritonitis (FIP) pathogenesis. The aim of this study was to evaluate sensitivity and specificity of a real-time reverse transcriptase polymerase chain reaction (RT-PCR) specifically designed to detect FCoV spike gene mutations at two nucleotide positions. It was hypothesized that this test would correctly discriminate feline infectious peritonitis virus (FIPV) and feline enteric coronavirus (FECV). The study included 63 cats with signs consistent with FIP. FIP was confirmed in 38 cats. Twenty-five control cats were definitively diagnosed with a disease other than FIP. Effusion and/or serum/plasma samples were examined by real-time RT-PCR targeting the two FCoV spike gene fusion peptide mutations M1058 L and S1060A using an allelic discrimination approach. Sensitivity, specificity, negative and positive predictive values including 95% confidence intervals (95% CI) were calculated. FIPV was detected in the effusion of 25/59 cats, one of them being a control cat with chronic kidney disease. A mixed population of FIPV/FECV was detected in the effusion of 2/59 cats; all of them had FIP. RT-PCR was negative or the pathotype could not be determined in 34/59 effusion samples. In effusion, sensitivity was 68.6% (95% CI 50.7-83.2), specificity was 95.8% (95% CI 78.9-99.9). No serum/plasma samples were positive for FIPV. Although specificity of the test in effusions was high, one false positive result occurred. The use of serum/plasma cannot be recommended due to a low viral load in blood.

Significance of serum antibody test for toxocariasis in healthy healthcare examinees with eosinophilia in Seoul and Gyeongsangnam-do, Korea.

PubMed

Kim, Hong Seok; Jin, Yan; Choi, Min-Ho; Kim, Jae-Hwan; Lee, Young Ha; Yoon, Cheong Ha; Hwang, Eui-Hyuk; Kang, Hun; Ahn, Sang-Yong; Kim, Gi Jin; Hong, Sung-Tae

2014-12-01

There have been numerous reports on the relationship between eosinophilia and toxocariasis. The present study investigated seropositive rates of toxocariasis among healthy people with or without eosinophilia in urban and rural areas, and assessed risk factors for positive antibody test. A total of 610 healthy people, who visited health check-up (Medicheck®, Korea Association of Health Promotion), 310 from Seoul and 300 from Gyeongsangnam-do, were subjected for this study. Their serum samples were tested by ELISA with the crude antigen of Toxocara canis larvae. Cross-reactions with other tissue invading helminth antigens were also investigated. Total antibody positive rate of toxocariasis was 8.7% of the 610 subjects. When the subjects were grouped into 3 by their eosinophil counts, the antibody positive rates significantly differed by the groups; 5.9% (18/306) in the group<350/µL, 10.0% (11/110) in the group 350-500/µL, and 12.4% (24/194) in the group>500/µL (P=0.028). A total of 22 serum samples cross-reacted with other tissue-invading helminth antigens. A questionnaire analysis recognized drinking alcohol and smoking as significant risk factors of toxocariasis. In conclusion, toxocariasis antibody positive rate is correlated with eosinophil counts. It is recommended that healthy subjects with eosinophilia by routine health examination and risk factors undergo Toxocara serology by multiantigen ELISA to investigate etiology.

Improved quantification of a commercial enzyme-linked immunosorbent assay kit for measuring anti-MDA5 antibody.

PubMed

Gono, Takahisa; Okazaki, Yuka; Murakami, Akihiro; Kuwana, Masataka

2018-04-09

To compare the quantitative performance for measuring anti-MDA5 antibody titer of two enzyme-linked immunosorbent assay (ELISA) systems: an in-house ELISA and the commercial MESACUP TM anti-MDA5 test. Anti-MDA5 antibody titer was measured in sera from 70 patients with dermatomyositis using an in-house ELISA and the MESACUP TM anti-MDA5 test side-by-side. For the commercial ELISA kit, serum samples diluted 1:101 were used according to the manufacturer's protocol, but serial dilutions of sera were also examined to identify the optimal serum dilution for quantification. The anti-MDA5 antibody titers measured by the in-house and commercial ELISAs were positively correlated with each other (r = 0.53, p = .0001), but the antibody titer measured by the commercial ELISA was less sensitive to change after medical treatment, and 37 (80%) of 46 anti-MDA5-positive sera had antibody titer exceeding the quantification range specified by the manufacturer (≥150 index). Experiments using diluted serum samples revealed that diluting the sera 1:5050 improved the quantitative performance of the MESACUP TM anti-MDA5 test, including a better correlation with the in-house ELISA results and an increased sensitivity to change. We improved the ability of the commercial ELISA kit to quantify anti-MDA5 antibody titer by altering its protocol.

Sample preparation and liquid chromatography-tandem mass spectrometry for multiple steroids in mammalian and avian circulation.

PubMed

Koren, Lee; Ng, Ella S M; Soma, Kiran K; Wynne-Edwards, Katherine E

2012-01-01

Blood samples from wild mammals and birds are often limited in volume, allowing researchers to quantify only one or two steroids from a single sample by immunoassays. In addition, wildlife serum or plasma samples are often lipemic, necessitating stringent sample preparation. Here, we validated sample preparation for simultaneous liquid chromatography--tandem mass spectrometry (LC-MS/MS) quantitation of cortisol, corticosterone, 11-deoxycortisol, dehydroepiandrosterone (DHEA), 17β-estradiol, progesterone, 17α-hydroxyprogesterone and testosterone from diverse mammalian (7 species) and avian (5 species) samples. Using 100 µL of serum or plasma, we quantified (signal-to-noise (S/N) ratio ≥ 10) 4-7 steroids depending on the species and sample, without derivatization. Steroids were extracted from serum or plasma using automated solid-phase extraction where samples were loaded onto C18 columns, washed with water and hexane, and then eluted with ethyl acetate. Quantitation by LC-MS/MS was done in positive ion, multiple reaction-monitoring (MRM) mode with an atmospheric pressure chemical ionization (APCI) source and heated nebulizer (500°C). Deuterated steroids served as internal standards and run time was 15 minutes. Extraction recoveries were 87-101% for the 8 analytes, and all intra- and inter-run CVs were ≤ 8.25%. This quantitation method yields good recoveries with variable lipid-content samples, avoids antibody cross-reactivity issues, and delivers results for multiple steroids. Thus, this method can enrich datasets by providing simultaneous quantitation of multiple steroids, and allow researchers to reimagine the hypotheses that could be tested with their volume-limited, lipemic, wildlife samples.

Highly sensitive and multiplexed platforms for allergy diagnostics

NASA Astrophysics Data System (ADS)

Monroe, Margo R.

Allergy is a disorder of the immune system caused by an immune response to otherwise harmless environmental allergens. Currently 20% of the US population is allergic and 90% of pediatric patients and 60% of adult patients with asthma have allergies. These percentages have increased by 18.5% in the past decade, with predicted similar trends for the future. Here we design sensitive, multiplexed platforms to detect allergen-specific IgE using the Interferometric Reflectance Imaging Sensor (IRIS) for various clinical settings. A microarray platform for allergy diagnosis allows for testing of specific IgE sensitivity to a multitude of allergens, while requiring only small volumes of patient blood sample. However, conventional fluorescent microarray technology is limited by i) the variation of probe immobilization, which hinders the ability to make quantitative, assertive, and statistically relevant conclusions necessary in immunodiagnostics and ii) the use of fluorophore labels, which is not suitable for some clinical applications due to the tendency of fluorophores to stick to blood particulates and require daily calibration methods. This calibrated fluorescence enhancement (CaFE) method integrates the low magnification modality of IRIS with enhanced fluorescence sensing in order to directly correlate immobilized probe (major allergens) density to allergen-specific IgE in patient serum. However, this platform only operates in processed serum samples, which is not ideal for point of care testing. Thus, a high magnification modality of IRIS was adapted as an alternative allergy diagnostic platform to automatically discriminate and size single nanoparticles bound to specific IgE in unprocessed, characterized human blood and serum samples. These features make IRIS an ideal candidate for clinical and diagnostic applications, such a POC testing. The high magnification (nanoparticle counting) modality in conjunction with low magnification of IRIS in a combined instrument offers four significant advantages compared to existing sensing technologies: IRIS i) corrects for any variation in probe immobilization, ii) detects proteins from attomolar to nanomolar concentrations in unprocessed biological samples, iii) unambiguously discriminates nanoparticles tags on a robust and physically large sensor area, iv) detects protein targets with conjugated nanoparticle tags (~40nm diameter), which minimally affect assay kinetics compared to conventional microparticle tagging methods, and v) utilizes components that make the instrument inexpensive, robust, and portable. This platform was successfully validated on patient serum and whole blood samples with documented allergy profiles (ImmunoCAPRTM, ThermoFisher Scientific).

Serum IGF-1, IGFBP-3 and their ratio: Potential biochemical growth maturity indicators.

PubMed

Jain, Nimisha; Tripathi, Tulika; Gupta, S K; Rai, Priyank; Kanase, Anup; Kalra, Shilpa

2017-12-01

Determination of skeletal maturation and remaining growth potential is an essential part of treatment planning in orthodontics. The aim of our study was to determine the relationship between IGF-1 levels, IGFBP-3 levels with CVM staging to track the pre pubertal and pubertal growth spurts in female patients in North Indian population. This cross-sectional study was conducted on ninety female subjects in the age group of 8-20 years. Blood samples were collected and centrifuged and serum samples were then analysed by Human IGF-1 and IGFBP-3 enzyme-linked immunosorbent assay kits, specific for IGF-1 and IGFBP-3, respectively. CVM staging on lateral cephalometric radiograph was determined for all patients. Analysis of variance test followed by a post hoc test was used to compare mean IGF-1 and IGFBP-3 corresponding to six stages of cervical vertebrae maturation stages. Linear Pearson's correlations were performed to determine the trends of IGF-1, IGFBP-3, and its ratio relating to CVM stage. The kappa statistic was used to measure inter and intra examiner reliability. P value <0.05 was considered as statistically significant. Mean serum IGF-1 levels were found to be highest (403.3 ± 12.3 ng/ml) at CVMI3 stage of CVMI. The post-hoc test revealed a significant difference in IGF-1 levels between all stages of CVMI, thereby indicating a specific range of IGF-1 levels for a specific skeletal stage. Mean serum IGFBP-3 levels were found to be highest (5186.8 ± 1384.2 ng/ml) at CVMI4 stage of CVMI. The mean serum IGFBP-3 levels at CVMI4 were found to be significantly higher than the levels at all other CVMI stages except CVMI3 stage. IGF-1 and IGFBP-3 can serve as a potential biochemical indicator for assessment of skeletal maturity.

The Dutch Brucella abortus monitoring programme for cattle: the impact of false-positive serological reactions and comparison of serological tests.

PubMed

Emmerzaal, A; de Wit, J J; Dijkstra, Th; Bakker, D; van Zijderveld, F G

2002-02-01

The Dutch national Brucella abortus eradication programme for cattle started in 1959. Sporadic cases occurred yearly until 1995; the last infected herd was culled in 1996. In August 1999 the Netherlands was declared officially free of bovine brucellosis by the European Union. Before 1999, the programme to monitor the official Brucella-free status of bovine herds was primarily based on periodical testing of dairy herds with the milk ring test (MRT) and serological testing of all animals older than 1 year of age from non-dairy herds, using the micro-agglutination test (MAT) as screening test. In addition, serum samples of cattle that aborted were tested with the MAT. The high number of false positive reactions in both tests and the serum agglutination test (SAT) and complement fixation test (CFT) used for confirmation seemed to result in unnecessary blockade of herds, subsequent testing and slaughter of animals. For this reason, a validation study was performed in which three indirect enzyme-linked immunosorbent assays (ELISAs), the CFT and the SAT were compared using a panel of sera from brucellosis-free cattle, sera from experimentally infected cattle, and sera from cattle experimentally infected with bacteria which are known to induce cross-reactive antibodies (Pasteurella, Salmonella, Yersinia, and Escherichia). Moreover, four ELISAs and the MRT were compared using a panel of 1000 bulk milk samples from Brucella-free herds and 12 milk samples from Brucella abortus- infected cattle. It is concluded that the ELISA obtained from ID-Lelystad is the most suitable test to monitor the brucelosis free status of herds because it gives rise to fewer false-positive reactions than the SAT.

Comparing dried and liquid blood serum samples of depressed patients: An analysis by Raman and infrared spectroscopy methods.

PubMed

Depciuch, J; Parlinska-Wojtan, M

2018-02-20

Depression is a serious mental illness. To study the mechanism of depression and search for new, more effective therapies, animal models are often used. Unfortunately, none of the available models reflects all the symptoms of depression. Therefore researchers are looking for new tools to diagnose depression. Unfortunately, the nowadays-available depression diagnosis methods are only psychological tests. However, it is known, that the amount of phospholipids, proteins and lipids decreases during depression. Raman and FTIR (Fourier Transform Infra Red) spectroscopies provide information on the chemical compounds in the measured sample e.g. blood serum. These spectroscopic techniques may thus become reliable and accurate tools for evaluating changes in the amount of phospholipids and proteins in depression disease. In this study differences between dried and liquid blood serum samples of healthy and depressed individuals measured by Raman (range 0-3000cm -1 ) and FTIR (Fourier Transform Infrared) (range 900-3000cm -1 ) spectroscopy were evaluated. The resulting spectra and accurate analysis led to the conclusion that an appropriate measurement of the background and the elimination of peaks from water had the greatest impact on the reliability of the results. Furthermore, after detailed studies of FTIR and Raman spectra of dried and liquid blood serum samples, including a complete analysis of peaks after Kramers-Kröning (KK) transformation, it was found that the sample preparation did not affect the results obtained by Raman spectroscopy. In FTIR measurements only a minimal effect on peak intensity was observed. Copyright © 2017 Elsevier B.V. All rights reserved.

A modified haemagglutination inhibition test for rubella antibodies, using standardized, freeze-dried reagents. Report of a comparative multi-centre trial.

PubMed Central

van Weemen, B.; Kacaki, J.

1976-01-01

A modified haemagglutination inhibition test for rubella antibodies, using standardized freeze-dried reagents, was developed and compared with haemagglutination inhibition tests using fresh erythrocytes. This comparison was made in collaboration with six European laboratories. A total of 4205 serum samples were tested. The results show that: (1) Sensitivity and reliability of the modified test are good; (2) the modified test can be performed in polystyrene microtitration plates. PMID:789763

Serum 25-hydroxyvitamin D is associated with reduced verbal episodic memory in healthy, middle-aged and older adults.

PubMed

Lam, Virginie; Albrecht, Matthew A; Takechi, Ryusuke; Prasopsang, Prachya; Lee, Ya Ping; Foster, Jonathan K; Mamo, John C L

2016-06-01

There is increasing evidence supporting an association of higher serum vitamin D concentration with better cognitive performance in older individuals. However, to date, consideration of the putative association between vitamin D and cognition has been based principally on studies investigating clinical participant samples manifesting vitamin D deficiency, particularly in older people. Moreover, relationships between vitamin D and cognition are typically not considered in the context of counter-regulatory calcium-modulating hormones or calcium homeostasis. Serum vitamin D/bioactive (ionised) calcium/parathyroid hormone homeostasis was considered in the context of cognitive performance in healthy, middle-aged and older individuals. A cross-sectional sample of 179 participants between the ages of 47-84 years was recruited for this study (114 females, 65 males). Participants provided fasting blood samples for analysis of serum 25-hydroxyvitamin D levels, ionised calcium (iCa) and parathyroid hormone (PTH) and completed cognitive measures of verbal episodic learning and memory. Serum 25-hydroxyvitamin D concentrations were negatively associated (with and without covariates of age, gender, depression and NART scores, iCa, and PTH) with measures of verbal episodic learning and memory, in particular with trial 5 of the Rey Auditory Verbal Learning Test (RAVLT) and long-delay free recall on the RAVLT. Overall, the findings from this study suggest an association between higher vitamin D status and poorer performance on verbal episodic memory in middle-aged and older individuals with normal vitamin D-calcium-PTH homeostasis. Despite requiring replication in other participant samples, this is a potentially important finding as it indicates that it may not be beneficial from a cognitive perspective to provide vitamin D supplements in individuals with already adequate vitamin D status.

Detection of serum antibodies against measles, mumps and rubella after primary measles, mumps and rubella (MMR) vaccination in children.

PubMed

Rafiei Tabatabaei, Sedigheh; Esteghamati, Abdoul-Reza; Shiva, Farideh; Fallah, Fatemeh; Radmanesh, Raheleh; Abdinia, Babak; Shamshiri, Ahmad Reza; Khairkhah, Masoumeh; Shekari Ebrahimabad, Hamideh; Karimi, Abdollah

2013-01-01

In Iran, the measles, mumps and rubella vaccine (MMR) is administered in a two-dose protocol where the first dose is scheduled at 12 months of age. This study aims to determine the efficacy of the MMR vaccine by testing IgM and IgG antibody levels 4 - 7 weeks after primary vaccination. A single group cohort study was performed on healthy children, 12 - 15 months of age, who were vaccinated at health centers affiliated with Shahid Beheshti University of Medical Sciences in Tehran, from January to April 2009. Children with negative vaccination and/or clinical history for measles, mumps or rubella were administered the first dose of the MMR live attenuated vaccine. IgG and IgM antibodies were checked by enzyme linked immunoassay (ELISA) in serum samples 4 - 7 weeks after vaccination. A child was considered seropositive if antibody levels were higher than the assay cut-off level set by the ELISA kit. Samples from 240 children were checked for antibodies against measles and rubella. Measles serum IgM level was positive in 71.7% of samples and IgG in 75.8%. The rubella serum IgM level was positive in 71.7% of children and IgG in 73.8%. From 190 blood samples that were checked for mumps antibodies, serum IgM was positive in 68.9% and IgG in 95.3%. No significant relationship was found between seropositivity and age or gender. IgG and IgM antibody levels were below the assay cut-off levels against measles and rubella in approximately one-fourth of the children following primary MMR vaccination. A second dose was necessary to raise the level of protection against measles and rubella.

The SLE-key test serological signature: new insights into the course of lupus.

PubMed

Putterman, Chaim; Pisetsky, David S; Petri, Michelle; Caricchio, Roberto; Wu, Alan H B; Sanz, Ignacio; Oates, Jim C; Wallace, Steve; Sorek, Rachel; Gerwien, Robert; Safer, Pennina; Jakobi-Brook, Keren; Cohen, Irun R

2018-06-04

We previously described the multiplex autoantibody SLE-key Rule-Out test, which detects a signature of autoantibody reactivity that distinguishes healthy subjects from SLE patients with 94% sensitivity, 75% specificity and 93% negative predictive value; thus, an individual manifesting a positive Rule-Out test score is unlikely to have SLE (e.g. lupus is excluded). The objective of this current study was to evaluate the stability of the lupus-associated signature over time. We used banked serum samples from healthy subjects (n = 51) and lupus patients (n = 50 individual samples and n = 181 paired samples, for a total of n = 412 serum samples). The samples were drawn at different times after diagnosis to analyse the impact on the SLE-key Rule-Out test of time elapsed since diagnosis and any changes in disease activity (as reflected by the SLEDAI score). The SLE signature remains stable for the first 10 years after diagnosis; in this time frame, <10% of patients manifested a positive Rule-Out score and the SLE-key Rule-Out score was independent of the underlying disease activity as reflected by the SLEDAI score. After ⩾10 years, ∼30% of lupus subjects scored as SLE Ruled-Out; the proportion of patients manifesting this status was even greater in the subset of individuals with a SLEDAI score of 0. These findings raise the possibility that a significant number of SLE patients manifest a change in their serological signature over time, and that such a signature change may signify an evolution in the immunological features of their disease relevant to patient management.

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

PubMed Central

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

2015-01-01

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

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

PubMed

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

2015-01-01

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

Pharmacokinetics in lactating women: prediction of alprazolam transfer into milk.

PubMed Central

Oo, C Y; Kuhn, R J; Desai, N; Wright, C E; McNamara, P J

1995-01-01

1. Alprazolam, a triazolobenzodiazepine, is extensively prescribed for the treatment of anxiety disorders, which predominantly affect women of child-bearing age. The purpose of the present study was to assess the pharmacokinetics of alprazolam and its two hydroxylated metabolites: 4-hydroxy-alprazolam and alpha-hydroxy-alprazolam in lactating human volunteers and to test the predictability of four recently reported models for drug transfer into milk based on physicochemical properties. 2. Multiple milk and serum samples in eight lactating subjects were collected up to 36 h following single oral doses of 0.5 mg alprazolam; suckling of the infant was discontinued after drug administration. 4-Hydroxy-alprazolam was the predominant metabolite in serum samples while alpha-hydroxy-alprazolam was not detected. 3. The mean oral clearance of alprazolam was 1.15 +/- 0.32 ml min-1 kg-1. The time course of alprazolam in milk roughly paralleled the perspective plasma time profile (mean serum residence time = 16.42 +/- 4.69 h; mean milk residence time = 18.93 +/- 7.03 h). The mean terminal half-life in serum was 12.52 +/- 3.53 h. 4. Observed milk/serum concentration ratios were determined in vivo as AUCmilk/AUCserum (mean M/S(obs) = 0.36 +/- 0.11).(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8527284

Chorionic villus sampling for abnormal screening compared to historical indications: prevalence of abnormal karyotypes.

PubMed

Marshall, Nicole E; Fraley, Gwen; Feist, Cori; Burns, Michael J; Pereira, Leonardo

2012-08-01

To determine the prevalence of abnormal karyotype results in women undergoing chorionic villus sampling (CVS) for abnormal first trimester screening compared to CVS for historical indications (advanced maternal age (AMA) or prior aneuploidy). Retrospective cohort of all patients undergoing CVS at Oregon Health & Science University from January 2006 to June 2010. Patients were separated based on CVS indication: (1) positive ultrasound (U/S) or serum screening; or (2) AMA or prior aneuploidy with normal or no screening. Prevalence of abnormal karyotype results were compared between groups. Fetal karyotyping was successful in 500 of 506 CVS procedures performed. 203 CVS were performed for positive screening with 69 abnormal karyotypes (34.0%). 264 CVS were performed for historical indications with 11 abnormal karyotypes (4.2%). This difference was statistically significant (χ(2) 71.9, p < 0.001; OR 11.8 [95% CI 5.8, 24.6]). There were two age-related aneuplodies in AMA women without positive screening. 42 out of 44 AMA women diagnosed with aneuploidy (95.5%) had abnormal U/S and/or serum screening (35 U/S, 4 serum, 3 U/S and serum). Combined ultrasound and serum screening should be recommended to all women, including AMA women, prior to undergoing invasive testing to improve risk-based counseling and minimize morbidity.

Comparison of rapid diagnostic tests to detect Mycobacterium avium subsp. paratuberculosis disseminated infection in bovine liver.

PubMed

Zarei, Mehdi; Ghorbanpour, Masoud; Tajbakhsh, Samaneh; Mosavari, Nader

2017-08-01

Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne's disease, a chronic enteritis in cattle and other domestic and wild ruminants. The presence of MAP in tissues other than intestines and associated lymph nodes, such as meat and liver, is a potential public health concern. In the present study, the relationship between the results of rapid diagnostic tests of the Johne's disease, such as serum ELISA, rectal scraping PCR, and acid-fast staining, and the presence of MAP in liver was evaluated. Blood, liver, and rectal scraping samples were collected from 200 slaughtered cattle with unknown Johne's disease status. ELISA was performed to determine the MAP antibody activity in the serum. Acid-fast staining was performed on rectal scraping samples, and PCR was performed on rectal scraping and liver samples. PCR-positive liver samples were used for mycobacterial culture. Overall, the results of this study demonstrated that MAP can be detected and cultured from liver of slaughtered cattle and rapid diagnostic tests of Johne's disease have limited value in detecting cattle with MAP infection in liver. These findings show that the presence of MAP in liver tissue may occur in cows with negative results for rapid diagnostic tests and vice versa. Hence, liver might represent another possible risk of human exposure to MAP. Given concerns about a potential zoonotic role for MAP, these results show the necessity to find new methods for detecting cattle with MAP disseminated infection.

Comparison of three commercial IgG and IgM ELISA kits for the detection of tick-borne encephalitis virus antibodies.

PubMed

Ackermann-Gäumann, Rahel; Tritten, Marie-Lise; Hassan, Mona; Lienhard, Reto

2018-05-01

Tick-borne encephalitis (TBE) is endemic in many parts of Europe and Asia. The diagnosis of this disease is essentially based on the demonstration of specific antibodies. For reasons of simplicity, automatization and quick availability of test results, enzyme-linked immunosorbent assays (ELISAs) are the method of choice for serological diagnosis of TBE. Here, we evaluated three commercially available anti-TBEV IgG and IgM ELISAs using 251 serum samples: the SERION ELISA classic FSME Virus/TBE Virus IgG and IgM kit (Virion\\Serion), the RIDASCREEN ® FSME/TBE IgG and IgM kit (R-Biopharm), and the anti-FSME/TBE virus ELISA "Vienna" IgG/anti-FSME/TBE virus ELISA IgM kit (Euroimmun). In total, discrepant test results for IgG and/or IgM were observed for 37/251 (14.7 %) of tested samples; differences were statistically significant. Reference values defined by serum neutralization test (SNT, n = 25) or results provided by EQA organizers (n = 2) were established for a subset of samples. In relation to these values, false-positive results were observed mainly for Euroimmun Vienna IgG and RIDASCREEN IgG, whereas false-negative results were primarily observed for Virion\\Serion IgG and RIDASCREEN IgM kits. In routine diagnostics, specificity problems are of major relevance and may be addressed by analyzing the respective samples using SNT. Copyright © 2018 Elsevier GmbH. All rights reserved.

IL8 and IL16 levels indicate serum and plasma quality.

PubMed

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

2018-02-09

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

Reassessment of the Access Testosterone chemiluminescence assay and comparison with LC-MS method.

PubMed

Dittadi, Ruggero; Matteucci, Mara; Meneghetti, Elisa; Ndreu, Rudina

2018-03-01

To reassess the imprecision and Limit of Quantitation, to evaluate the cross-reaction with dehydroepiandrosterone-sulfate (DHEAS), the accuracy toward liquid chromatography-mass spectrometry (LC-MS) and the reference interval of the Access Testosterone method, performed by DxI immunoassay platform (Beckman Coulter). Imprecision was evaluated testing six pool samples assayed in 20 different run using two reagents lots. The cross-reaction with DHEAS was studied both by a displacement curve and by spiking DHEAS standard in two serum samples with known amount of testosterone. The comparison with LC-MS was evaluated by Passing-Bablock analysis in 21 routine serum samples and 19 control samples from an External Quality Assurance (EQA) scheme. The reference interval was verified by an indirect estimation on 2445 male and 2838 female outpatients. The imprecision study showed a coefficient of variation (CV) between 2.7% and 34.7% for serum pools from 16.3 and 0.27 nmol/L. The value of Limit of Quantitation at 20% CV was 0.53 nmol/L. The DHEAS showed a cross-reaction of 0.0074%. A comparison with LC-MS showed a trend toward a slight underestimation of immunoassay vs LC-MS (Passing-Bablock equations: DxI=-0.24+0.906 LCMS in serum samples and DxI=-0.299+0.981 LCMS in EQA samples). The verification of reference interval showed a 2.5th-97.5th percentile distribution of 6.6-24.3 nmol/L for male over 14 years and <0.5-2.78 nmol/L for female subjects, in accord with the reference intervals reported by the manufacturer. The Access Testosterone method could be considered an adequately reliable tool for the testosterone measurement. © 2017 Wiley Periodicals, Inc.

Evaluation of the chemiluminescent enzyme immunoassay system for the measurement of testosterone in the serum and whole blood of stallions

PubMed Central

TOISHI, Yuko; TSUNODA, Nobuo; NAGATA, Shun-ichi; KIRISAWA, Rikio; NAGAOKA, Kentaro; WATANABE, Gen; YANAGAWA, Yojiro; KATAGIRI, Seiji; TAYA, Kazuyoshi

2017-01-01

Testosterone (T) concentration is a useful indicator of reproductive function in male animals. However, T concentration is not usually measured in veterinary clinics, partly due to the unavailability of reliable and rapid assays for animal samples. In this study, a rapid chemiluminescent enzyme immunoassay system (CLEIA system) that was developed for the measurement of T concentration in humans use was validated for stallion blood samples. First, serum T concentrations were measured using the CLEIA system and compared with those measured by a fluoroimmunoassay that has been validated for use in stallions. The serum T concentrations measured by the two methods were highly correlated (r = 0.9865, n = 56). Second, to validate the use of whole blood as assay samples, T concentrations in whole blood and in the serum were measured by the CLEIA system. T concentrations in both samples were highly correlated (r = 0.9665, n = 64). Finally, to evaluate the practical value of the CLEIA system in clinical settings, T concentrations were measured in three stallions with reproductive abnormalities after the administration of human chorionic gonadotropin (hCG). Two stallions with small or absent testes in the scrotum showed an increase in T production in response to hCG administration and one stallion with seminoma did not. In conclusion, the CLEIA system was found to be a rapid and reliable tool for measuring T concentrations in stallions and may improve reproductive management in clinical settings and in breeding studs. PMID:29129877

Infectivity of severe acute respiratory syndrome during its incubation period.

PubMed

Zeng, Guang; Xie, Shu-Yun; Li, Qin; Ou, Jian-Ming

2009-12-01

To evaluate the infectivity of severe acute respiratory syndrome (SARS) during its incubation period by investigating chains of transmission and individuals isolated for medical observation with a view to providing scientific evidence for updating protocols of medical isolation. Individuals related with the two SARS chains of transmission in Beijing in 2003 and a group of individuals isolated for medical observation in Haidian district of Beijing during the SARS outbreak were selected as subjects of study. Contactors with SARS patients and those with symptom development following the contacts were investigated via questionnaire. Serum samples were collected from super transmitters and tested for SARS-CoV antibody by neutralization test and enzyme linked immunosorbent assay (ELISA). A total of 1112 contactors were investigated in three surveys. Of them, 669 had a history of close contact with symptomatic SARS patients, 101 developed symptoms with a rate of 15.1%, 363 had a history of close contact with patients in their incubation period, none of whom developed symptoms (0%). Serum samples were collected from 32 highly-exposed individuals, of whom 13 developing SARS symptoms after contact had serum samples positive for SARS-CoV antibody. Samples collected from the asymptomatic contactors were all negative for SARS-CoV antibody. SARS cases are infectious only during their symptomatic period and are non-infectious during the incubation period. Isolation for medical observation should be placed for individuals who are in close contact with symptomatic SARS patients. The results of our study are of decisive significance for the Ministry of Health to the definition of SARS close contactor.

Near-infrared Raman spectroscopy to detect anti-Toxoplasma gondii antibody in blood sera of domestic cats: quantitative analysis based on partial least-squares multivariate statistics

NASA Astrophysics Data System (ADS)

Duarte, Janaína; Pacheco, Marcos T. T.; Villaverde, Antonio Balbin; Machado, Rosangela Z.; Zângaro, Renato A.; Silveira, Landulfo

2010-07-01

Toxoplasmosis is an important zoonosis in public health because domestic cats are the main agents responsible for the transmission of this disease in Brazil. We investigate a method for diagnosing toxoplasmosis based on Raman spectroscopy. Dispersive near-infrared Raman spectra are used to quantify anti-Toxoplasma gondii (IgG) antibodies in blood sera from domestic cats. An 830-nm laser is used for sample excitation, and a dispersive spectrometer is used to detect the Raman scattering. A serological test is performed in all serum samples by the enzyme-linked immunosorbent assay (ELISA) for validation. Raman spectra are taken from 59 blood serum samples and a quantification model is implemented based on partial least squares (PLS) to quantify the sample's serology by Raman spectra compared to the results provided by the ELISA test. Based on the serological values provided by the Raman/PLS model, diagnostic parameters such as sensitivity, specificity, accuracy, positive prediction values, and negative prediction values are calculated to discriminate negative from positive samples, obtaining 100, 80, 90, 83.3, and 100%, respectively. Raman spectroscopy, associated with the PLS, is promising as a serological assay for toxoplasmosis, enabling fast and sensitive diagnosis.

Temporal variation in viral hemorrhagic septicemia virus antibodies in freshwater drum (Aplodinotus grunniens) indicates cyclic transmission in Lake Winnebago, Wisconsin

USGS Publications Warehouse

Wilson-Rothering, Anna; Marcquenski, Susan; Koenigs, Ryan P.; Bruch, Ronald; Kamke, Kendall; Isermann, Daniel A.; Thurman, Andrew; Toohey-Kurth, Kathy; Goldberg, Tony

2015-01-01

Viral hemorrhagic septicemia virus (VHSV) is an emerging pathogen that causes mass mortality in multiple fish species. In 2007, the Great Lakes freshwater strain, type IVb, caused a large die-off of freshwater drum (Aplodinotus grunniens) in Lake Winnebago, Wisconsin, USA. To evaluate the persistence and transmission of VHSV, freshwater drum from Lake Winnebago were tested for antibodies to the virus using recently developed virus neutralization (VN) and enzyme-linked immunosorbent (ELISA) assays. Samples were also tested by real-time reverse transcription-PCR (rRT-PCR) to detect viral RNA. Of 548 serum samples tested, 44 (8.03%) were positive by VN (titers ranging from 1:16 to 1:1,024) and 45 (8.21%) were positive by ELISA, including 7 fish positive by both assays. Antibody prevalence increased with age and was higher in one northwestern area of Lake Winnebago than in other areas. Of 3,864 tissues sampled from 551 fish, 1 spleen and 1 kidney sample from a single adult female fish collected in the spring of 2012 tested positive for VHSV by rRT-PCR, and serum from the same fish tested positive by VN and ELISA. These results suggest that VHSV persists and viral transmission may be active in Lake Winnebago even in years following outbreaks and that wild fish may survive VHSV infection and maintain detectable antibody titers while harboring viral RNA. Influxes of immunologically naive juvenile fish through recruitment may reduce herd immunity, allow VHSV to persist, and drive superannual cycles of transmission that may sporadically manifest as fish kills.

Temporal Variation in Viral Hemorrhagic Septicemia Virus Antibodies in Freshwater Drum (Aplodinotus grunniens) Indicates Cyclic Transmission in Lake Winnebago, Wisconsin

PubMed Central

Wilson-Rothering, Anna; Marcquenski, Susan; Koenigs, Ryan; Bruch, Ronald; Kamke, Kendall; Isermann, Daniel; Thurman, Andrew; Toohey-Kurth, Kathy

2015-01-01

Viral hemorrhagic septicemia virus (VHSV) is an emerging pathogen that causes mass mortality in multiple fish species. In 2007, the Great Lakes freshwater strain, type IVb, caused a large die-off of freshwater drum (Aplodinotus grunniens) in Lake Winnebago, Wisconsin, USA. To evaluate the persistence and transmission of VHSV, freshwater drum from Lake Winnebago were tested for antibodies to the virus using recently developed virus neutralization (VN) and enzyme-linked immunosorbent (ELISA) assays. Samples were also tested by real-time reverse transcription-PCR (rRT-PCR) to detect viral RNA. Of 548 serum samples tested, 44 (8.03%) were positive by VN (titers ranging from 1:16 to 1:1,024) and 45 (8.21%) were positive by ELISA, including 7 fish positive by both assays. Antibody prevalence increased with age and was higher in one northwestern area of Lake Winnebago than in other areas. Of 3,864 tissues sampled from 551 fish, 1 spleen and 1 kidney sample from a single adult female fish collected in the spring of 2012 tested positive for VHSV by rRT-PCR, and serum from the same fish tested positive by VN and ELISA. These results suggest that VHSV persists and viral transmission may be active in Lake Winnebago even in years following outbreaks and that wild fish may survive VHSV infection and maintain detectable antibody titers while harboring viral RNA. Influxes of immunologically naive juvenile fish through recruitment may reduce herd immunity, allow VHSV to persist, and drive superannual cycles of transmission that may sporadically manifest as fish kills. PMID:26135873

Does Carica papaya leaf-extract increase the platelet count? An experimental study in a murine model.

PubMed

Dharmarathna, Sinhalagoda Lekamlage Chandi Asoka; Wickramasinghe, Susiji; Waduge, Roshitha Nilmini; Rajapakse, Rajapakse Peramune Veddikkarage Jayanthe; Kularatne, Senanayake Abeysinghe Mudiyanselage

2013-09-01

To investigate the potential role of fresh Carica papaya (C. papaya) leaf extract on haematological and biochemical parameters and toxicological changes in a murine model. In total 36 mice were used for the trial. Fresh C. papaya leaf extract [0.2 mL (2 g)/mouse] was given only to the test group (18 mice). General behavior, clinical signs and feeding patterns were recorded. Blood and tissue samples were collected at intervals. Haematological parameters including platelet, red blood cell (RBC), white blood cell (WBC), packed cell volume (PCV), serum biochemistry including serum creatinine, serum glutamic-oxaloacetic transaminase (SGOT) and serum glutamic-pyruvic transaminase (SGPT) were determined. Organs for possible histopathological changes were examined. Neither group exhibited alteration of behavior or reduction in food and water intake. Similarly, no significant changes in SGOT, SGPT and serum creatinine levels were detected in the test group. Histopathological organ changes were not observed in either group of mice except in three liver samples of the test group which had a mild focal necrosis. The platelet count (11.33±0.35)×10⁵/µL (P=0.00004) and the RBC count (7.97±0.61)×10⁶/µL (P=0.00003) were significantly increased in the test group compared to that of the controls. However, WBC count and PCV (%) values were not changed significantly in the test group. The platelet count in the test group started to increase significantly from Day 3 (3.4±0.18×10⁵/µL), reaching almost a fourfold higher at Day 21 (11.3×10⁵/µL), while it was 3.8×10⁵/µL and 5.5×10⁵/µL at Day 3 and Day 21 respectively in the control. Likewise, the RBC count in the test group increased from 6×10⁶/µL to 9×10⁶/ µL at Day 21 while it remained near constant in the control group (6×10⁶/µL). Fresh C. papaya leaf extract significantly increased the platelet and RBC counts in the test group as compared to controls. Therefore, it is very important to identify those chemicals of C. papaya leaves as it can be recommended to be used as a medication to boost thrombopoiesis and erythropoiesis in humans and in animals in which these cell lineages have been compromised.

Does Carica papaya leaf-extract increase the platelet count? An experimental study in a murine model

PubMed Central

Dharmarathna, Sinhalagoda Lekamlage Chandi Asoka; Wickramasinghe, Susiji; Waduge, Roshitha Nilmini; Rajapakse, Rajapakse Peramune Veddikkarage Jayanthe; Kularatne, Senanayake Abeysinghe Mudiyanselage

2013-01-01

Objective To investigate the potential role of fresh Carica papaya (C. papaya) leaf extract on haematological and biochemical parameters and toxicological changes in a murine model. Methods In total 36 mice were used for the trial. Fresh C. papaya leaf extract [0.2 mL (2 g)/mouse] was given only to the test group (18 mice). General behavior, clinical signs and feeding patterns were recorded. Blood and tissue samples were collected at intervals. Haematological parameters including platelet, red blood cell (RBC), white blood cell (WBC), packed cell volume (PCV), serum biochemistry including serum creatinine, serum glutamic-oxaloacetic transaminase (SGOT) and serum glutamic-pyruvic transaminase (SGPT) were determined. Organs for possible histopathological changes were examined. Results Neither group exhibited alteration of behavior or reduction in food and water intake. Similarly, no significant changes in SGOT, SGPT and serum creatinine levels were detected in the test group. Histopathological organ changes were not observed in either group of mice except in three liver samples of the test group which had a mild focal necrosis. The platelet count (11.33±0.35)×105/µL (P=0.000 04) and the RBC count (7.97±0.61)×106/µL (P=0.000 03) were significantly increased in the test group compared to that of the controls. However, WBC count and PCV (%) values were not changed significantly in the test group. The platelet count in the test group started to increase significantly from Day 3 (3.4±0.18×105/µL), reaching almost a fourfold higher at Day 21 (11.3×105/µL), while it was 3.8×105/µL and 5.5×105/µL at Day 3 and Day 21 respectively in the control. Likewise, the RBC count in the test group increased from 6×106/µL to 9×106/ µL at Day 21 while it remained near constant in the control group (6×106/µL). Conclusions Fresh C. papaya leaf extract significantly increased the platelet and RBC counts in the test group as compared to controls. Therefore, it is very important to identify those chemicals of C. papaya leaves as it can be recommended to be used as a medication to boost thrombopoiesis and erythropoiesis in humans and in animals in which these cell lineages have been compromised. PMID:23998013

RHCE*ceCF encodes partial c and partial e but not CELO an antigen antithetical to Crawford

PubMed Central

Hipsky, Christine Halter; Lomas-Francis, Christine; Fuchisawa, Akiko; Reid, Marion E.; Moulds, Marilyn; Christensen, JoAnn; Nickle, Pam; Vege, Sunitha; Westhoff, Connie

2010-01-01

Background RH43 (Crawford) is encoded by RHCE*ce with nucleotide changes 48G>C, 697C>G, and 733C>G (RHCE*ceCF). We investigated the Rh antigen expression and antibody specificities in four patients with this allele. Methods Hemagglutination tests, DNA extraction, PCR-RFLP, AS-PCR, reticulocyte RNA isolation, RT-PCR cDNA analyses, cloning, and sequencing were performed by standard procedures. Results RBCs from two patients typed D+C−E−c+e+/−, hrS−/+W, hrB− and their serum was reactive (3+) with all RBC samples of common Rh phenotype tested, but non reactive with Rhnull or D- - RBCs (apparent alloanti-Rh17). At the RHCE locus, Patient 1 was homozygous for RHCE*ceCF, and Patient 2 inherited RHCE*ceCF in trans to a silenced RHCE*cE. Cross testing of serum and RBCs from these two samples showed mutual compatibility, indicating that both antibodies define the same novel high prevalence antigen on Rhce. Two additional patients, one whose serum contained alloanti-c but the RBCs typed C+c+, and one whose serum contained anti-e but the RBCs typed E+e+, also had RHCE*ceCF. RHCE*Ce was present in trans in the former and RHCE*cE in the latter patient. Conclusion We report that amino acid changes on RhceCF (Trp16Cys, Gln233Glu, and Leu245Val) alter the protein to the extent that c and e antigens are partial, and a high prevalence antigen, we have named CELO (provisional ISBT number 004058; RH58) is not expressed. CELO is antithetical to RH43 (Crawford). PMID:20609196