表达绿色荧光蛋白的重组痘苗病毒的初步应用

目的:制备表达绿色荧光蛋白的重组痘苗病毒,并初步探讨其应用。方法:构建制备表达绿色荧光蛋白的重组痘苗病毒RVJ11LacZ-I1LGFP;分别利用药物昔多福韦与抗痘苗病毒高效价免疫血清,建立基于该病毒的荧光生成抑制实验及荧光减数中和实验。结果:荧光生成抑制实验与传统的噬斑生成抑制实验相比,结果一致,但判读更直接快速;重组痘苗病毒RVJ11LacZ-I1LGFP亦可用于体外快速高通量评价正痘病毒疫苗的中和能力。结论:利用表达绿色荧光蛋白的重组痘苗病毒建立了直接简便快速高通量的抗痘病毒药物筛选及体外中和评价技术。

Sensitive Fluorescence-Based Anti-Poxvirus Assay and Microneutral izing Assay In Vitro Using a Modified Vaccinia Virus(Tiantan)with GFP Expression

Objective: To prepare recombinant vaccinia virus(Tiantan) expressing green fluorescent protein(GFP),and to explore its application.Methods: Recombinant vaccinia virus expressing GFP was constructed and named as rRVJ11LacZ-I1LGFP.Using anti-poxvirus inhibitor(cidofovir),we developed fluorescence-based inhibi tion assay(FBIA) based on rRVJ11LacZ-I1LGFP and compared with a traditional plaque inhibition assay(PIA).Meantime,using a vaccinia immune serum,we also developed an fluorescence reduction neutralization test(FRNT) based on rRVJ11LacZ-I1LGFP.Results: The FBIA can be used for screening and efficacy evaluation of orthopox virus inhibitor in vitro,the result was similar with PIA and designed to replace the PIA.We also developed an in novative FRNT for neutralizing capacity evaluation of vaccine against poxvirus.The readout of both fluores cence-based assays were simple,rapid and sensitive,and potential for automated high-throughput.Conclusion: FBIA and FRNT assay was developed using a modified vaccinia virus(Tiantan) with GFP expression.