Stable hepatitis C virus RNA detection by RT-PCR during four days storage

BACKGROUND: Suboptimal specimen processing and storage conditions of samples which contain hepatitis C virus (HCV) RNA may result in a decline of HCV RNA concentration or false-negative results in the detection of HCV RNA in serum. We evaluated the stability of HCV RNA in serum and clotted blood samples stored at room temperature or at 4°C for 4 days with the aim of optimizing the standard procedures of processing and storage of samples. METHODS: Blood from five HCV RNA positive patients was collected in tubes with and without separator gel, centrifuged 1 or 6 hours after collection. Samples were then left 6, 24, 48, 72 or 96 h at room temperature (21.5 – 25.4°C) or at 4°C before determining their HCV RNA level using the COBAS AMPLICOR HCV MONITOR Test, vs 2.0 (Roche Diagnostic Systems). RESULTS: The logarithm of the HCV RNA level measurements remained within a 0.3 value of the means for 4 days at both temperatures (room temperature or 4°C). CONCLUSIONS: We conclude that blood samples may be collected and aliquoted within 6 h of collection and can be stored at 4°C for 72 hours as proposed by the manufacturer without significant differences in measured HCV RNA level. Our results indicate that lapses in this scheme may still yield reliable results

Evaluation of a Cytomegalovirus Glycoprotein B Recombinant Enzyme Immunoassay To Discriminate between a Recent and a Past Infection

Fetal damage following cytomegalovirus (CMV) intrauterine infection is mostly linked to primary infection. To differentiate primary infection from nonprimary infection, immunoglobulin M (IgM) tests are not reliable enough, and measurement of the IgG avidity appears to be the method that is the most widely used at present. In the present study the performance of the Vidas (bioMérieux) avidity assay was compared with that of a new enzyme immunoassay based on the use of a recombinant CMV glycoprotein B protein (Biotest)

A latex agglutination test for the detection of canine parvovirus and corresponding antibodies.

Latex particles coated with rat monoclonal antibodies directed against a canine parvovirus are agglutinated by the virus antigens. In this study, the reaction was quantitated by a technique which had been described as immunoassay by particles counting. The method is as sensitive as a radioimmunoassay with the same antibody, but has the advantages of simplicity and safety. It allows the detection of parvoviral antigens to 4 ng/ml. Incubation of these antigens with specific antibodies resulted in inhibition of the agglutination reaction. By this procedure the presence of antibodies against canine parvovirus can be detected to a concentration of 150 ng/ml. The system can be easily automated, thus increasing reproducibility. This latex agglutination technique can be performed as a slide test; and although this method is 4-5 times less sensitive, it could be useful in field studies, for the detection of the viral antigens and the corresponding antibodies

An anti-HIV-1 gag protein rat monoclonal antibody library.

A set of 18 rat monoclonal antibodies (MAbs) directed against human immunodeficiency virus type 1 (HIV-1) gag proteins was derived from 4 independent fusion protocols. The epitopes recognized were delineated using a random fragment expression library representing the whole HIV-1IIIB genome. This panel of rat MAbs was used to analyze the antigenicities of the HIV-1 CAp24 major core protein and the HIV-1 NCp7 nucleocapsid protein. As a result, a limited set of antigenic domains as defined, 3 on CAp24 between amino acids (aa) 195 and 268, 323 and 329, 329 and 352, and one on NCp7 (aa 382-392). Only 4 mouse anti-CAp24 MAbs appeared to recognize the COOH-terminal domain (aa 329-352) defined by the majority of our MAbs. The rat anti-CAp24 (Q1B10) and the rat anti-NCp7 (I5B11) MAbs described here, defined two newly described epitopes, aa 323-329 and aa 382-392

Human herpes virus 8 infection in kidney transplant patients in Belgium.

Kaposi sarcoma (KS) may arise as a complication of kidney transplantation. In the Saint Luc Teaching Hospital in Brussels, patients of both Belgian and foreign origin are treated. The prevalence of human herpes virus 8 (HHV-8) infection differs in different geographical settings. We wanted to estimate the background infection rate and the risk of infection in our transplant population: a first step towards evaluating the necessity of HHV-8 screening. Serum samples were taken from 210 organ donors over a period of 7 years (30 per year) and from 200 kidney recipients from whom two sera were tested, one pre-transplant and the second 6-12 months post-transplant. All sera were screened for HHV-8 by an enzyme-linked immunosorbant assay using recombinant ORF 65 and ORF 73 antigens and an immunofluorescence assay for the latent antigen. Reactive samples were confirmed by western blotting. Seven donors (3.3%) were positive for HHV-8 antibodies. Of 198 pre-transplant sera available for evaluation, 15 were positive (7.6%). Post-transplantation 18/199 (9%) were positive: four (2.1% of negatives) had a documented seroconversion and one lost the antibodies. No patients developed KS. A substantial number of kidney transplant patients already had antibodies to HHV-8 at the time of transplantation. A further 2.1% of seronegative patients had seroconversion, which could have been acquired through the transplanted organ (3.3% of donors were positive) or through transfusion