A new PCR-based in vitro neutralization assay of hepatitis B virus / 中华实验和临床病毒学杂志

ABSTRACT

<p><b>OBJECTIVE</b>To establish a PCR-based neutralization assay of hepatitis B virus (HBV), which may be applied for detecting neutralizing antibodies against HBV and used as an in vitro model to screen new HBV vaccines.</p><p><b>METHODS</b>Immune serum was mixed with HBV stock. The mixture was incubated and then inoculated onto Hep G2 cell monolayers. After adsorption, washing and incubation, HBV DNA was extracted from the cells and detected by PCR. The neutralization effect was determined based on the PCR results.</p><p><b>RESULTS</b>Two HBV stocks suitable for the neutralization assay were selected from 18 serum samples collected from patients with hepatitis B. The neutralization assay was optimized in the conditions of using 10 infectious doses of the HBV stock and incubating the cell culture for 24 hours prior to PCR detection. Four immune sera obtained from mice immunized with commercial HBV vaccine and 2 serum specimens from mice immunized with 2 new HBV vaccine candidates definitely blocked the in vitro HBV adsorption. However, 4 sera obtained from normal mice and 2 sera from mice immunized with 2 hepatitis E virus vaccine candidates did not show any neutralizing activity.</p><p><b>CONCLUSION</b>The established new PCR-based in vitro HBV neutralization assay is a simple, rapid and economic assay. It may be used as a model for primary evaluation for HBV vaccine candidates prior to primate assay.</p>