Enzyme-linked Immunosorbent Assay Screening Test Research Articles

Urinary ethyl glucuronide (uEtG) as a marker for alcohol consumption in liver transplant candidates: a real-world cohort.

In order to reduce alcohol relapse after liver transplantation (LT), the German national guidelines for waiting-list maintenance and organ allocation demand a minimum 6-month period of alcohol abstinence pre-LT, confirmed by measuring urinary ethyl glucuronide (uEtG). Between January 2015 and June 2016, uEtG was measured at least once in 339 cirrhotic patients with an indication for LT at the University Medical Center Mainz. uEtG was measured with an enzyme-linked immunosorbent assay (ELISA) screening test (cutoff value: 500 µg/L). For uEtG values ≥ 500 µg/L, liquid chromatography-mass spectrometry (LC-MS/MS) was performed as a confirmatory assay. Data were collected prospectively in a transplant database. Of the 339 potential liver transplant candidates, uEtG was negative in 86.4 %. Most patients were male (64.3 %), with an average age of 56.42 ± 10.1 years. In the multivariate analysis, mean corpuscular volume (p = 0.001), urinary creatinine (p = 0.001), gamma-glutamyl transferase (p = 0.001), and hemoglobin (p = 0.003) were significantly associated with a positive uEtG test result. The sensitivity of the ELISA screening test was 100 % for uEtG values > 2000 µg/L, as confirmed by LC-MS/MS. uEtG is an effective parameter to reveal alcohol consumption by patients on the waiting list for LT. The sensitivity of the ELISA is excellent for uEtG values > 2000 µg/L, for which LC-MS/MS confirmation could be omitted.

Seroepidemiology of human T-cell lymphotropic virus type-I in blood donors of Northeastern Iran, Sabzevar.

Background and Objectives:Human T-cell lymphotropic virus type-I (HTLV-I) infection is considered as a public health challenge in endemic areas. The virus is associated with severe diseases, such as adult T-cell leukemia/lymphoma, and HTLV-I-associated myelopathy/tropical spastic paraparesis. One of the major routes of the HTLV-I transmission includes blood transfusion. Sabzevar is located in the endemic region of HTLV-I infection. The aim of the present study was to determine the seroprevalence of HTLV-I infection in the blood donors in Sabzevar.Materials and Methods:A total of 35,067 blood donors in Sabzevar from March 2009 to April 2012 who were screened with HTLV-I on the enzyme-linked immunosorbent assay screening test were included in this survey. Reactive samples that confirmed by western blot were considered to be seropositive cases. The required data were obtained from blood donors’ database of blood transfusion service.Results:The overall prevalence of HTLV-1 based on the positive result of western blot test was 0.14%. The seropositive donors aged 17–59 years with a mean age of 38.10 ± 11.82. The prevalence rates of HTLV-I infection in 3 years of study were 0.19%, 0.14%, and 0.09%, respectively. A significant relation between age, sex, educational level, and history of blood donation was observed with seropositivity of HTLV-I.Conclusion:The improvement of donor selection and laboratory screening caused a decline in the prevalence of infection in blood donors. Given the lower prevalence of infection in regular donors with lower age and higher educational level, more efforts should be done to attract blood donors from these populations.

Development of a new immunoassay for the detection of ethyl glucuronide (EtG) in meconium: validation with authentic specimens analyzed using LC-MS/MS. Preliminary results

Ethyl glucuronide (EtG) measurement in neonatal meconium has emerged as a reliable marker to objectively assess prenatal exposure to maternal ethanol complementary to fatty acid ethyl ester (FAEEs) measurement. The detection of EtG in meconium is currently a lengthy, difficult and expensive process using liquid chromatography tandem mass spectrometry (LC-MS/MS) as the analytical procedure. An enzyme-linked immunosorbent assay (ELISA) for the identification of EtG in meconium was developed, validated and applied to authentic meconium specimens from newborns collected in Europe. The ELISA procedure was calibrated using 0.45, 0.9, 1.35 and 1.8 nmol/g (100, 200 300 and 400 ng/g) standards. Meconium (0.25 g) was mixed thoroughly, with extraction buffer (pH 7.3; 0.5 mL). The tube was capped, sonicated, centrifuged and the supernatant was decanted. An aliquot of the extract (50 μL) was placed in the well of the microplate followed by enzyme conjugate (150 μL). The plate was incubated for 1 h, washed with deionized water, dried and substrate (200 μL) was added. After 30 min incubation, stop solution was added and the plate was read at 450 nm and 650 nm. Samples were also analyzed for EtG and FAEEs by validated LC-MS/MS assays. Using an EtG cut-off of 0.9 nmol/g for both ELISA screening test and confirmatory LC-MS/MS, immunoassay sensitivity was 100%; specificity 78%; positive-predictive value (PPV) 29% and negative-predictive value (NPV) 100%. The assay is proposed as a preliminary screening test for the meconium of newborns suspected of being born to mothers drinking alcohol during pregnancy.

Plasma Exchange for Heparin-Induced Thrombocytopenia: Is There Enough Evidence?

Plasma Exchange for Heparin-Induced Thrombocytopenia: Is There Enough Evidence?

Lidocaine in the horse: its pharmacological effects and their relationship to analytical findings.

Lidocaine is a local anaesthetic agent that is widely used in equine medicine. It is also an Association of Racing Commissioners International (ARCI) Class 2 foreign substance that may cause regulators to impose substantial penalties if residues are identified in post race urine samples. Therefore, an analytical/pharmacological database was developed for this drug. Using our abaxial sesamoid local anaesthetic model, the highest no-effect dose (HNED) for the local anaesthetic effect of lidocaine was determined to be 4 mg. Using enzyme-linked immunosorbent assay (ELISA) screening, administration of the HNED of lidocaine to eight horses yielded peak serum and urine concentrations of apparent lidocaine of 0.84 ng/mL at 30 min and 72.8 ng/mL at 60 min after injection, respectively. These concentrations of apparent lidocaine are readily detectable by routine ELISA screening tests (LIDOCAINE ELISA, Neogen, Lexington, KY). ELISA screening does not specifically identify lidocaine or its metabolites, which include 3-hydroxylidocaine, dimethylaniline, 4-hydroxydimethylaniline, monoethylglycinexylidine, 3-hydroxymonoethylglycinexylidine, and glycinexylidine. As 3-hydroxylidocaine is the major metabolite recovered from equine urine, it was synthesized, purified and characterized, and a quantitative mass spectrometric method was developed for 3-hydroxylidocaine as recovered from horse urine. Following subcutaneous (s.c.) injection of the HNED of lidocaine, the concentration of 3-hydroxylidocaine recovered from urine reached a peak of about 315 ng/mL at 1 h after administration. The mean pH of the 1 h post dosing urine samples was 7. 7, and there was no apparent effect of pH on the amount of 3-hydroxylidocaine recovered. Within the context of these experiments, the data suggests that recovery of less than 315 ng/mL of 3-hydroxylidocaine from a post race urine sample is unlikely to be associated with a recent local anaesthetic effect of lidocaine. Therefore these data may be of assistance to industry professionals in evaluating the significance of small concentrations of lidocaine or its metabolites in postrace urine samples. It should be noted that the quantitative data are based on analytical methods developed specifically for this study, and that methods used by other laboratories may yield different recoveries of urine 3-hydroxylidocaine.

Prevalence of human T cell lymphotropic virus type II in American Indian populations of the southwestern United States.

We investigated the seroprevalence of human T cell lymphotropic virus type II (HTLV-II) using a screening enzyme-linked immunosorbent assay (ELISA) and immunoblot confirmation among the predominant tribes (Pueblo and Athapaskan) served by two large Indian Health Service facilities in New Mexico. Among persons being treated for sexually transmitted diseases, eight (3.2%) of 250 were seropositive for HTLV II, compared with eight (2.1%) of 385 women attending prenatal clinics. In a survey of unselected patients at one of the facilities, 15 (3.4%) of 446 were seropositive. Of 31 seropositive subjects, 25 were infected with HTLV-II and six infections could not be typed. Sera from nine (29%) of the 31 infected subjects had absorbance values less than the manufacturer's cutoff in the ELISA. Both Pueblo and Athapaskan groups had similar overall seroprevalences, but women tended to have a slightly higher seroprevalence than men, and seroprevalence tended to increase with age. These data show that HTLV-II infection is present among diverse groups of American Indians in the southwestern United States. Present ELISA screening tests, such as those used in this study, lack sensitivity to HTLV-II infection unless a reduced absorbance cutoff is used.

Prevalence of antibodies to hepatitis C virus in a Dutch group of haemodialysis patients related to risk factors

In January 1992, 122 chronic haemodialysis patients (>9 months dialysis) from the University Hospital Utrecht and its outpatient dialysis facility were tested for the presence of anti-hepatitis C virus (HCV) antibodies. The objective was to identify risk factors for HCV infection in chronic haemodialysis patients in an attempt to explain the high prevalence of anti-HCV antibodies found among such haemodialysis patients. A second generation enzyme linked immuno-sorbent assay (EIA) was used as a screening test. Results were confirmed by a recombinant immunoblot assay and by the polymerase chain reaction (PCR). Four (3·3%) of 122 patients reacted positively in the EIA screening test as well as in the immunoblot assay; three of these were positive using PCR. None of the patients with anti-HCV antibodies had received blood products other than blood from transfusions, none had markers for a hepatitis B virus (HBV) infection or admitted intravenous drug abuse. A total number of 2395 units of blood, unscreened for HCV, had been administered to our dialysis group, an average of 19·6 ( sd 44·7) units per patient. The seroprevalence of anti-HCV antibodies among blood donors in Utrecht was 0·03%. Patients with antibodies to HCV had been on dialysis longer than those dialysis patients without HCV antibodies (odds ratio 1·8, 95% CI 0·99-3·29). We conclude that the risk for HCV infection for this dialysis group can only partially be attributed to unscreened blood transfusions. Haemodialysis itself may play a role in transmission of HCV.

Incidence of Human Immunodeficiency Virus Seroconversion in US Navy and Marine Corps Personnel, 1986 Through 1988

From 1986 through 1988, the US Navy and US Marine Corps administered 1,956,631 enzyme-linked immunosorbent assay screening tests for antibodies to the human immunodeficiency virus to 1,070,511 active-duty enlisted and officer personnel. This study identified all persons who had an initial test with negative results. This population was then followed up and those who later tested positive for human immunodeficiency virus were identified as seroconverters. There were 582 seroconversions identified from a total of 987,479 person-years at risk. The age adjusted seroconversion rate and 95% Poisson confidence intervals for navy personnel was 0.69 per 1000 person-years (95% confidence interval, 0.63 to 0.76). Age-adjusted rates in men were 5.0 times those of women. Age-adjusted rates in blacks were 3.7 times those of whites. The age-adjusted seroconversion rate in Marine Corps personnel was 0.28 per 1000 person-years (95% confidence interval, 0.22 to 0.36). Similar demographic patterns were present in the Marine Corps and the US Navy. This study is one of the first reports of incidence of human immunodeficiency virus seroconversion by demographic characteristics in a large, young, and apparently healthy population.

Pea lectin as a universal cell binding agent for ELISA (screening) tests

A cell binding assay (CBA) was developed in which plant lectins are used as binding agents between microtiter plates and human cells. After binding, the cells were fixed by mild glutaraldehyde treatment. Their antigenic activity was investigated by enzyme-linked immunosorbent assay (ELISA) with several monoclonal antibodies. The binding method resulted in cell layers that remained firmly attached to the plates during the washing and incubation procedures of the ELISA. A comparative phenotype analysis, performed by indirect membrane fluorescence, showed that cells bound by this method, do not lose their antigenic activity. This binding assay can be used as a rapid, large scale screening test for monoclonal antibodies to membrane antigens of malignant and normal cells.