诺如病毒衣壳蛋白单克隆抗体的制备及鉴定  

Preparation and characterization of monoclonal antibodies against Norovirus capsid protein

作　　者：郭丽[1] 周红莉[1] 江涛[2] 王敏[1] 计融[2] 王健伟[1] 洪涛[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所,北京100052 [2]中国疾病预防控制中心营养与食品安全所

出　　处：《中国人兽共患病学报》2006年第5期414-418,共5页Chinese Journal of Zoonoses

基　　金：国家"十五"科技攻关计划资助项目(项目编号:2003BA712A03-04;2001BA804A22)

摘　　要：目的制备诺如病毒衣壳蛋白特异性单克隆抗体,为诺如病毒的快速检测及致病机制的研究提供实验材料。方法用分别表达GGII1、GGII3、GGII4和GGII7型诺如病毒衣壳蛋白的重组腺病毒联合免疫BALB/c小鼠,取其脾细胞与小鼠骨髓瘤细胞融合,获得分泌高滴度针对衣壳蛋白的杂交瘤细胞株,测定单抗免疫球蛋白亚类及单抗效价,用间接ELISA、间接免疫荧光和Western-blot检测单克隆细胞株的特异性。结果通过细胞融合和克隆化,筛选出4株分泌抗衣壳蛋白的杂交瘤细胞株V1E、V1E9、V1D6和V6D6。间接ELISA、免疫荧光和Western-blot检测结果表明,V1E和V1E9可以与GGII4型诺如病毒衣壳蛋白发生特异性反应,V1D6和V6D6可以同时与GGII1、GGII3、GGII4和GGII7型诺如病毒衣壳蛋白发生特异性反应。结论获得了诺如病毒特异性单克隆抗体,为制备免疫诊断试剂盒及致病机制的研究奠定了基础。To prepare the monoelonal antibodies against Norovirus capsid protein in order to develope a rapid assay for Norovirus and to investigate the pathogenesis of virus. BALB/c mice were separately immunized with combination of the recombinant adenoviruses expressing GGⅡ1, GGⅡ3, GGⅡ4 and GGⅡ7 Norovirus capsid protein respectively. Hybridoma cell lines secreting monoclonal antibodies （McAbs） against Norovirus capsid protein were screened after the fusion of mouse splenic cells with SP2/0 cells. The Ig subtypes and titer of the McAbs were assayed with ELISA and the specificity of the MeAbs obtained was examined by indirect ELISA, indirect fluorescence and Western-blot assay. After cell fusion and three rounds of subcloning, four hybridoma cell lines secreting monoclonal antibodies specifically against Norovirus capsid protein were obtained, and respectively named as V1E, V1E9, V1D6 and V6D6. Indirect ELISA, indirect fluorescence and Western-blot assay showed that the McAbs V1E3 and V1E9 could specifically recognize GGⅡ4 Norovirus capsid protein, while V1D6 and V6D6 specifically with GGⅡ1, GGⅡ3, GGⅡ4 and GGⅡ7 Norovirus capsid protein. It is concluded that the specific anti-Norovirus capsid protein McAbs were developed and these MeAbs may be useful in the development of detection assay and basic research of Norovirus.

关 键 词：诺如病毒 衣壳蛋白 单克隆抗体 

分 类 号：R373.2[医药卫生—病原生物学]