山羊关节炎-脑炎病毒P28蛋白的表达与间接ELISA方法的建立  

Establishment of the indirect ELISA for detection of antibody against caprine arthritis encephalitis virus

作　　者：刘慧敏[1,2] 相文华[2] 李一经[1] 

机构地区：[1]东北农业大学动物医学院,黑龙江哈尔滨150030 [2]中国农业科学院哈尔滨兽医研究所哈尔滨维科生物技术开发公司,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2014年第7期555-558,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：黑龙江省自然科学基金(ZJN0702-02)

摘　　要：为建立检测山羊关节炎-脑炎病毒(CAEV)血清抗体的间接ELISA方法,本研究将原核表达并纯化的CAEV P28蛋白作为检测抗原,采用方阵法确定包被抗原和待检血清的最佳工作浓度,经过对反应条件的优化建立了CAEV间接ELISA抗体检测方法。该方法仅对CAEV血清检测为阳性,对口蹄疫、山羊痘和蓝舌病病毒的阳性血清检测结果均为阴性;敏感性试验表明标准阳性血清1∶3 200倍稀释时,检测结果仍为阳性。该方法的批内和批间重复试验的变异系数分别小于7%和10%。对人工感染山羊血清进行检测,该ELISA方法最早可以在第2周检出抗体,琼脂凝集免疫扩散试验(AGID)最早可以在第4周检出抗体;与AGID试验方法相比,ELISA检出率提高27.2%。结果表明,本研究所建立的间接ELISA方法具有良好的特异性、敏感性和可重复性,为该病的诊断和监测提供了有效的技术手段。Caprine arthritis encephalitis （CAE） is a chronic debilitating disease of goats caused by a lentivirus responsible for economic losses as a result of a drop in milk production and weight loss. To establish the serum diagnosis method of CAE, an indirect ELISA was developed for the detection of CAE virus （CAEV） antibody with the P28 protein of CAEV as coating antigen expressed in E.coli. Under the optimized reaction conditions, the assay was able to specifically detect the positive serum of CAEV with the detection limit of 1：3,200 for the CAEV standard positive serum, but no cross-reactions were found against the positive sera of other virus, including foot and mouth disease, bluetongue virus and goat pox virus. The coefficients of variation for intra- and inter-assay were lower than 7% and 10%, respectively. The ELISA was capable to detect positively for the experimentally infected goats as early at 14 days post inoculation, comparing with 28 days detected by agar gel immunodiffusion test. The established ELISA is a rapid, sensitive and specific for detecting antibodies against CAEV and has a potential to complement for diagnosis of CAE and for serologic surveillance of CAEV infection.

关 键 词：山羊关节炎-脑炎病毒 p28基因 间接ELISA 

分 类 号：S852.65[农业科学—基础兽医学]