A simple and rapid method for the quantitation of secreted hepatitis B virions in cell culture models.

Cell culture models for hepatitis B virus (HBV) remain the mainstay for screening and testing the efficacy of anti‑hepatitis B virus agents. Gradient‑based ultracentrifugation followed by Southern Blotting is used for hepatitis B virion estimation in cell culture; this method has several limitations. We report the development of an assay using a commercially available HBsAg‑ELISA plate for immunocapture followed by real‑time PCR for quantification of hepatitis B virions in cell cultures. This assay is rapid, highly sensitive (50 copies/reaction) and highly specific for virion‑associated DNA. In addition, the assay requires only 20 μL of supernatant, allowing scaling down of transfections.

Significance of the hepatitis C virus (HCV) core antigen as an alternative plasma marker of active HCV infection.

PURPOSE: To evaluate the role of core antigen (Ortho trak-C assay) as a marker of active HCV infection in comparison to HCV RNA as detected by reverse transcription polymerase chain reaction (RT-PCR). METHODS: This evaluation was carried out during January 2000 to December 2003 in HCV infected individuals who were treatment naomicronve or were on anti-viral therapy. Additionally, sequential plasma samples from patients on clinical follow-up were included in this study. A total of 167 samples from 61 patients were tested by trak-C and RT-PCR. HCV RNA detection was achieved by a RT-PCR. Trak-C assay results were also compared in a limited proportion of these samples with known HCV viral load and genotype. RESULTS: Of 167 samples tested, 56.9% were RNA positive and 43.1% were RNA negative while 50.3% were trak-C positive and 49.7% were trak-C negative, yielding a sensitivity of 85.3% and a specificity of 95.8% for the trak-C assay (Kappa co-efficient = 0.8). The concentration of HCVcAg and HCV RNA showed significant correlation (n=38, r=0.334, P =0.04). The trak-C assay detected the most prevalent HCV genotypes in India without significant difference (P =0.335). The difference between mean absorbance values of HCV RNA positive samples compared to HCV RNA negative samples in the trak-C assay was highly significant (P < 0.000). Qualitative results of trak-C assay and RT-PCR were comparable in 93% of follow-up samples. CONCLUSIONS: Trak-C assay can be recommended for confirmation of HCV infection and follow-up in laboratories with resource-poor facilities.