脑炎心肌炎病毒3C蛋白酶抗体的制备和检测  

Preparation and detection of antibody to encephalomyocarditis virus 3C protease

作　　者：宋芹芹[1] 李公启[1] 韩俊[1] Song Qinqin;Li Gongqi;Han Jun(State Key Laboratory of Infectious Disease Prevention and Control,Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases,National Institute for Viral Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China)

机构地区：[1]中国疾病预防控制中心病毒病预防控制所传染病预防控制国家重点实验室传染病诊断与治疗创新协作中心,北京102206

出　　处：《中华实验和临床病毒学杂志》2021年第4期463-466,共4页Chinese Journal of Experimental and Clinical Virology

基　　金：传染病重大专项(2018ZX10102001)。

摘　　要：目的:制备脑炎心肌炎病毒(Encephalomyocarditis virus,EMCV)的3C蛋白酶抗体并检测抗体特异性反应位点。方法:构建原核表达载体pQE30-3C,将蛋白进行体外原核表达,免疫纯种BALB/c小鼠,制备单克隆抗体,通过Western blot检测制备的抗体与3C蛋白结合的特异性,通过免疫荧光和Western blot检测抗体与EMCV病毒中表达的3C蛋白酶结合的特异性。原核表达具有不同抗原性的EMCV 3C的3个不同片段,并通过Western blot检测单克隆抗体与EMCV 3C蛋白酶不同片段的反应特异性。结果:单克隆抗体与原核表达的3C蛋白酶反应良好,通过Western blot可以检测到病毒中表达的3C蛋白,通过免疫荧光也可以检测到EMCV病毒中的3C蛋白。通过与3C片段的反应发现,单克隆抗体与C端反应,即抗体针对的是C端处的抗原决定簇。结论:成功制备EMCV 3C蛋白酶的抗体,能与原核表达的3C蛋白和病毒3C蛋白反应,且针对的是C端的抗原决定簇。Objective To prepare encephalomyocarditis virus(EMCV)3C protease antibody and detect response sites of these antibody.Methods Prokaryotic expression vector pGEX-2T-3C was constructed and expressed.Then BALB/c mice was immunized with 3C protein and antibodies were obtained.The specificity of the antibody binding to 3C protein was detected by immunofluorescence and Western blotting.Three different fragments of EMCV 3C were expressed and detected with monoclonal antibodies through Western blotting.Results The antibody reacted specifically with 3C proteins of prokaryotic expression,and bound well with 3C protein of BHK cells infected with EMCV virus through Western blotting and immunofluorescence.The reaction between fragments of 3C protein and the antibody revealed that the antibody could react with the C-terminal of 3C protein.Conclusions The prepared monoclonal antibody recognized C terminal of EMCV 3C protein.

关 键 词：脑炎心肌炎病毒 3C蛋白酶 单克隆抗体 

分 类 号：S85[农业科学—兽医学]