Asia1型口蹄疫病毒单克隆抗体的制备及双抗体夹心ELISA方法的建立  

Preparation of monoclonal antibody against Foot-and-mouth disease virus serotype Asia1 and establishment of double-antibody sandwich ELISA

作　　者：梁伟峰[1,2] 周国辉[2] 刘文铭 李超斯[1,2] 刘云[1] 于力[2] 

机构地区：[1]东北农业大学动物医学学院,黑龙江哈尔滨150030 [2]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/牛羊病创新团队,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2016年第3期226-229,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金青年项目(31101801)

摘　　要：为建立Asia1型口蹄疫(FMD)快速病原学诊断方法,本研究采用纯化的Asia1型口蹄疫病毒(FMDV)免疫BALB/c小鼠,取免疫小鼠脾细胞与SP2/0骨髓瘤细胞进行融合并筛选到一株能够稳定分泌抗Asia l型FMDV单克隆抗体(MAb)2E10。经间接免疫荧光试验特异性鉴定,2E10为FMDV型特异型MAb。其抗体亚类为Ig G1/资,杂交瘤细胞上清和腹水抗体效价分别为1颐256和1颐104,相对亲和力常数为2.5 mol/L。在此基础上,以本实验室前期制备的MAb 2D8为包被抗体,以HRP标记的MAb 2E10为检测抗体建立了检测Asia1型FMDV抗原的双抗体夹心ELISA(DAS-ELISA)方法。该方法针对FMDV细胞毒的最低检出量为103TCID50,对O型FMDV、A型FMDV、牛肠道病毒、牛呼肠孤病毒及牛传染性鼻气管炎病毒检测结果均为阴性。本研究建立的Asia1型FMDV DAS-ELISA方法为Asia1型FMD病原学诊断试剂盒的研发奠定了基础。To establish a rapid assay for serotype Asial Foot-and-mouth disease virus （FMDV） detection, a hybridoma cell line （2El0） secreting monoclonal antibody （MAb） against the Asial FMDV was prepared by fusion of the SP2/0 myloma cells with spleen cells of BALB/c mice immunized by purified FMDV. The MAb 2El0 was IgG1 isotype and contained kappa light chain, which was specific to serotype Asial FMDV with affinity index （RAI） of 2.5 mol/L. A double-antibody sandwich ELISA （DAS-ELISA） for detection of FMDV serotype Asial was developed with the MAb 2D8 prepered previously in this Lab as capture antibody and the MAb 2El0 conjugated with horseradish peroxidase （HRP） as detector antibody. This assay was specific for serotype Asial FMDV detection with a detection limit of 103 TCIDs0 FMDV, while no cross reactions were found for other related viruses such as serotype O FMDV, A FMDV, bovine enteroviruses, reovious, infectious bovine rhinotracheitis virus. The study would facilitate further development of the pathogen diagnosis kit for serotype Asial FMDV detection.

关 键 词：ASIA1型口蹄疫病毒 单克隆抗体 双抗体夹心ELISA 

分 类 号：S852.65[农业科学—基础兽医学]