Multiplex PCR normalization and parallel detection of HBV and HCV / 中华实验和临床病毒学杂志

ABSTRACT

<p><b>BACKGROUND</b>To design and establish a method of multiplex PCR normalization and parallel detection of HBV and HCV.</p><p><b>METHODS</b>Using two pairs of primers, one inner and the other outer, each having a 20 bp common sequence, the authors amplified target DNA for two rounds. All products would have this common sequence. Using this common sequence as primer the authors performed further amplification. Finally, multiplex PCR was normalized to a single PCR style and eliminated multiple factors disturbance. Four kinds of nucleic acid extraction method were compared. Multiplex PCR normalization was established and optimized by using orthogonal design by analysing 6 kinds of key factors. The method was evaluated by detecting 28 samples of HBV and HCV.</p><p><b>RESULTS</b>The sensitivity, specificity, diagnostic idex and efficiency for HBsAg, HCV antibody positive patients were 83.3%, 70.0%, 153.3%, 72.2% respectively. The sensitivity, specificity, idex and efficiency for HBsAg positive patients were 78.6%, 80.0%, 158.6%, and 79.2%, respectively. The sensitivity, specificity, idex and efficiency for HCV antibody positive patients were 75.0%, 90.0%, 165.0% and 83.3% respectively.</p><p><b>CONCLUSIONS</b>The multiplex PCR normalization method may have potential applicability in parallel amplification of multiple genes of pathogens.</p>