以原核表达的非洲猪瘟病毒pK205R蛋白为包被抗原的间接ELISA抗体检测方法的建立  

Development of an indirect ELISA for detection of antibody against African swine fever virus(ASFV) with prokaryotic expressed ASFV pK205R protein as the coating antigen

作　　者：邬旭龙[1] 彭彬[1] 姜睿姣 肖璐[1] 王印[1,2] 姚学萍[1,2] 杨泽晓[1,2] 张鹏飞[1] 张博[1] 

机构地区：[1]四川农业大学动物医学院,四川成都611130 [2]动物疫病与人类健康四川省重点实验室,四川成都611130

出　　处：《中国预防兽医学报》2016年第10期800-803,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金："十二五"国家科技支撑计划(2013BAD12B04)

摘　　要：为建立快速检测非洲猪瘟病毒(ASFV)的血清学方法,本研究原核表达ASFV pK205R重组蛋白,并以其为包被抗原,通过反应条件优化,建立了一种快速的ASFV间接ELISA检测方法。结果显示,原核表达的ASFV pK205R蛋白约为44 ku,western blot证实表达蛋白具有良好的反应原性;以其作为包被抗原建立的ASFV抗体间接ELISA方法仅对ASFV血清检测为阳性,与猪瘟病毒、猪伪狂犬病毒、猪繁殖与呼吸综合症病毒、副猪嗜血杆菌及猪大肠杆菌阳性血清均无交叉反应,具有良好的特异性;该方法检测灵敏度为1∶2 560;批内和批间重复性试验的变异系数均小于10%;利用该方法对临床样品检测的结果与国外商品化试剂盒检测结果符合率为100%。本研究建立的ASFV抗体间接ELISA方法为防止该病传入我国以及ASFV的实时监测提供技术储备。To develop a rapid and sensitive assay for the detection of antibody against African swine fever virus （ASFV）, an indirect ELISA for ASFV antibody detection was established with the prokaryotic expressed ASFV pK205R protein as the coating antigen. Under the optimized reacting conditions, the indirect ELISA was specific for the detection of ASF positive serum, and had no cross-reaction with positive sera to classical swine fever virus, pseudorabies virus, porcine reproductive and respiratory syndrome virus, Haemophilus parasuis and Escherichia coli. The limit of the detection reached 2,560 times dilution using standard positive serum, and inter-and intra-assay coefficient of variation （CV） were both less than 10%. The coincidence rate between the established assay and import ASFV commercial kit was 100%. The established indirect ELISA detection for ASF was sensitive and efficient, it provided technique for prevention and real-time monitoring of ASFV infection.

关 键 词：原核表达 K205R 免疫印迹 间接ELISA 

分 类 号：S852.65[农业科学—基础兽医学]