Original Article
Evaluation of a novel fully automated real-time PCR assay for hepatitis B virus DNA quantification
Xiaoying Chen, Lu Long, Qiong Li, Jian Gao, Tianming Li, Xiangbo Huang, Lei He, Xiangmei Chen, Min Li, Tao Shen
Published 2015-10-11
Cite as Chin J Lab Med, 2015, 38(10): 696-700. DOI: 10.3760/cma.j.issn.1009-9158.2015.10.011
Abstract
ObjectiveTo evaluate the analytical performance of a novel HBV DNA assay based on automated DNA extraction and real-time fluorescence quantitative PCR.
MethodsAnalytic verification studies. Accuracy and lower limit of detection were assessed by determining a panel of HBV standard plasma of WHO. HBV standard plasma (genotype A, B, C and D) at 6 different concentrations were measured 18 times to evaluate precision and reproducibility. Pseudo-viral particles at high HBV DNA concentration were serially diluted to assess linear range. One hundred and forty-four clinical specimens were quantified for HBV DNA so as to evaluate the correlation between the new test and COBAS® system.
ResultsQuantification of HBV standard plasma showed acceptable accuracy, with each deviation between observed and expected values within ±0.35 lg IU/ml (–0.17–0.32 lg IU/ml). Intra-assay coefficients of variation (CV) for genotype A, B, C and D were 3.87%–6.32%, 0.45%–14.68%, 0.16%–8.36% and 0.64%–13.01% respectively, and the inter-assay CV were 5.67%–9.69%, 1.28%–15.68%, 0.36%–9.05% and 1.69%–13.65%, separately. Linearity assessment exhibited an excellent dynamic range of linear quantification from 20 to 1.0×1010 IU/ml (r=0.998, P<0.001). And the satisfactory results obtained at 3 levels of HBV DNA concentration (10, 20, 50 IU/ml, respectively) confirmed the claimed lower limit of detection with 5/5 detectable rate at 20 IU/ml. Furthermore, good correspondence was observed between the new HBV DNA assay and the COBAS® system with 100% (144/144) qualitative coincidence and significant correlation based on 104 positive data (r=0.984, P<0.000 1).
ConclusionsThe novel fully-automated real-time PCR assay displayed good analytical and clinical performance for highly sensitive detection of HBV DNA. It was well suited for monitoring antiviral responses as well as drug resistance according to current clinical practice guidelines for the management of chronic HBV infection.(Chin J Lab Med, 2015, 38: 696-700)
Key words:
Hepatitis B virus; DNA viruses; Real-time polymerase chain reaction; Reagent kits, diagnostic
Contributor Information
Xiaoying Chen
Department of Clinical Laboratory, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127, China
Lu Long
Department of Microbiology &
Infectious Disease Center of Peking University, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
Qiong Li
Jian Gao
Tianming Li
Xiangbo Huang
Lei He
Xiangmei Chen
Min Li
Tao Shen
