Development and assessment of a novel real-time PCR assay for quantitation of HBV DNA

• Published in: Journal of virological methods 2002-05, Vol.103 (2), p.201-212
• Main Authors: Paraskevis, D., Haida, C., Tassopoulos, N., Raptopoulou, M., Tsantoulas, D., Papachristou, H., Sypsa, V., Hatzakis, A.
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; DNA, Viral - analysis; HBV; Hepatitis B - virology; Hepatitis B virus; Hepatitis B virus - genetics; Hepatitis B virus - isolation & purification; Human viral diseases; Humans; Infectious diseases; Medical sciences; Polymerase Chain Reaction - methods; Quantitation; Real-time PCR; Reproducibility of Results; Reverse Transcriptase Polymerase Chain Reaction; Viral diseases; Viral hepatitis
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/S0166-0934(02)00033-2

HBV DNA quantitation is used extensively for the monitoring of treatment of hepatitis B virus (HBV) infection. The aim of this study was to develop a highly sensitive and reproducible real-time PCR (RTD-PCR) assay for the quantitation of HBV DNA using the LightCycler system. The performance of this assay was assessed by analyzing serial dilutions of HBV genomic DNA of known concentration and the lower limit of detection was found to be 1 DNA copy/reaction. By using serial dilutions of plasmid standard, RTD-PCR was determined to quantify HBV DNA in a 10–log 10 dynamic range. RTD-PCR was found to be more sensitive than the commercially available tests such as the Quantiplex™ HBV DNA and the AMPLICOR HBV MONITOR™ assays. The median coefficient of variation of interexperimental variability was 3.2%. The HBV DNA values obtained with RTD-PCR were highly correlated with assays available commercially. These findings suggest that our RTD-PCR assay combines high sensitivity and reproducibility for HBV DNA quantitation in an incomparable high dynamic range of quantitation.