摘要
应用大肠杆菌表达及纯化的戊型肝炎病毒(hepatitis E virus,HEV)P1重组蛋白制备的抗体,建立了检测HEV抗原的双抗体夹心ELISA方法,并对吉林省和辽宁省部分地区猪群HEV的感染进行了病原流行病学调查。方阵滴定法确定鼠源捕获抗体IgG最佳包被量为0.2μg,兔源酶标抗体最佳稀释倍数为1∶500。对大量阴性粪便进行了检测与统计学分析,确定了双抗体夹心ELISA检测HEV的判定标准,即当D490≥0.235判定为阳性。特异性、敏感性和重复性的试验结果表明,本试验建立的方法具有特异、敏感、快速和可重复性好等特点。该方法与RT-PCR方法相比,操作简单、便于基层推广使用。对吉林省、辽宁省不同地区猪群粪便进行检测,发现不同地区不同饲养期的猪群均存在着HEV感染,感染率为9%~33%。本试验建立的双抗体夹心ELISA方法检测HEV抗原,为HEV流行病学调查、临床诊断提供了有效的手段。
Hepatitis E virus(HEV)infection is an emerging zoonotic disease that poses a potential threat to the public health and the pig industry as well.In this study,a sandwich ELISA for detection of HEV antigens was developed using antibodies raised against the recombinant proteins P1 derived from HEV and used to investigate the HEV infections in pig population in Jilin province.The concentrations of coating-antibody and HRP-conjugated anti-HEV P1 were optimized using square matrix titrimetry.The criterion for sandwich ELISA was determined and sample was assessed as positive when its D490≥0.235.The established ELISA had high sensitivity and specificity with a well repeatability and simplicity.Compared with the RT-PCR method,the sandwich ELISA has advantage over RT-PCR with a low cost,easy operation in rural area and elimination of the false negative/positive results.Epidemiological survey indicated that HEV infections in pig herds in Jilin province were ranging from 9%to 33%.The results in this study will provide a sensitive,specific,simple and rapid antigen detection method for diagnosis and epidemiologic survey of HEV infection.
作者
袁悦
李欣
郭昌明
马振乾
胡俊英
王旭
林倩
王新平
YUAN Yue;LI Xin;GUO Chang-ming;MA Zhen-qian;HU Jun-ying;WANG Xu;LIN Qian;WANG Xin-ping(College of Veterinary Medicine,Jilin University,Changchun 130062,China;Qingdao Yibang Biotech Limited Company,Qingdao,Shandong266032,China;Institute for Zoonotic Diseases Research,Jilin University,Changchun 130062,China)
出处
《中国兽医学报》
CAS
CSCD
北大核心
2019年第6期1130-1134,1139,共6页
Chinese Journal of Veterinary Science
基金
国家重点研发计划资助项目(2017YFD0500104
2016YFD0500904)
