日本脑炎病毒E Ⅲ结构域蛋白的表达及多克隆抗体的制备与鉴定  

PROKARYOTIC EXPRESSION OF ENVELOPE PROTEIN DOMAIN Ⅲ OF JAPANESE ENCEPHALITIS VIRUS AND PREPARATION OF POLYCLONAL ANTIBODIES

作　　者：沈强[1,2] 魏建超[2] 史子学[2] 常维山[1] 赵凡凡[2] 蒋春英[2] 王水明[3] 马志永[2] 

机构地区：[1]山东农业大学,泰安271018 [2]中国农业科学院上海兽医研究所,上海200241 [3]南京出入境检验检疫局,南京211136

出　　处：《中国动物传染病学报》2013年第5期7-12,共6页Chinese Journal of Animal Infectious Diseases

基　　金：国家国际科技合作项目(2010DFB33920);国家质检总局科技计划项目资助(2012KJ)

摘　　要：使用原核表达系统融合表达乙型脑炎病毒EⅢ结构域蛋白,经过纯化后多次免疫BALB/c小鼠,制备了鼠抗JEV E蛋白的多克隆抗体。Western blot结果表明该抗体与重组蛋白能够发生明显反应,与浓缩的JEV反应显出1条分子量大小约为53kDa的目的条带(JEV E蛋白);间接免疫荧光(indirect immunofluorescence,IFA)显示,多克隆抗体与BHK-21细胞内的JEV作用,在细胞质内产生绿色荧光,而对照组无特异性荧光;空斑减少中和试验测定多抗血清的中和抗体效价(PRNT50)为1:160。以上结果暗示制备的多克隆抗体能与重组蛋白和JEV E蛋白都能发生良好反应,并且该多抗有较高水平的JEV中和抗体效价。In this study, E protein domain III （EDIII） of JEV was expressed in E. coli and purified with a His column. BALB/e mice were immunized with EDIII and polyclonal antibodies to E protein were prepared. The reactivity of the prepared antibodies was characterized in Western blotting and IFA. Furthermore, the polyclonal antibodies at dilution of 1：160 blocked the entry of JEV into BHK-21 cells in plaque reduction neutralization test. These results demonstrated that the polyclonal antibodies of JEV EDIII possessed good antigenic specificity and could be used for development of a diagnostic method of JEV infection.

关 键 词：乙型脑炎病毒 E蛋白 原核表达 多克隆抗体 中和抗体 

分 类 号：S852.659.6[农业科学—基础兽医学]