猪流行性腹泻病毒血清中特异性IgG抗体间接ELISA检测方法的建立  

Development of Indirect ELISA Assay for Detection of Porcine Epidemic Diarrhea Virus Antibodies in Sera of Naturally Infected Pigs

作　　者：谷长维 谷长乐 胡博[1,2] GU Changwei;GU Changle;HU Bo(Institute of Special Wild Economic Animal and Plant Science,CAAS,Changchun 130112,China;Key Laboratory of Special Animal Epidemic Disease,Ministry of Agriculture and Rural Affairs,Changchun 130112,China)

机构地区：[1]中国农业科学院特产研究所,长春130112 [2]农业农村部经济动物疫病重点实验室,长春130112

出　　处：《中国动物传染病学报》2022年第3期106-112,共7页Chinese Journal of Animal Infectious Diseases

基　　金：国家重点研发计划项目(2017YFD0501600)。

摘　　要：本研究旨在开发和优化新型间接ELISA,使其成为一种简单、快速、灵敏、特异的检测抗PEDV抗体的方法,并评价其作为诊断PED方法的有效性。以Vero细胞培养全病毒抗原(LNCT2株)间接ELISA检测方法为基础,以中和试验(NT)为参照,通过受试者工作特征曲线(ROC)分析,确定截断值为0.320,敏感性为92.6,特异性为90.1%。曲线下面积(AUC)为0.949,说明间接ELISA检测具有较好的准确性。间接ELISA与中和试验呈正相关(R=0.815,P<0.05)。此外,kappa静校正结果也显示出良好的一致性。抗原包被保存板干燥后不同时间(1 d、2 W、1、2、3、4、5、6个月)的敏感性和特异性分别为100%和80%~100%。本研究开发的间接ELISA检测可作为PEDV感染的血清学调查和疾病控制的可靠检测手段,且我们的抗原包被ELISA板可在4℃保存至少6个月。The objective of this study was to develop and optimize a novel indirect ELISA as a simple,rapid,sensitive and specific method for the detection of anti-PEDV antibodies and evaluate the efficiency of the assay as a diagnostic method for PED.The indirect ELISA test was developed based on whole virus antigen prepared from Vero cell cultures infected with the NK94P6 strain and compared with neutralization test(NT).Subsequently,this method was evaluated by receiver operating characteristic(ROC)analysis and the cut-offvalue was determined as 0.320 with sensitivity and specificity of 92.6%and 90.1%,respectively.The area under curve(AUC)was 0.949,indicating excellent accuracy of this test.There was significant positive correlation between indirect ELISA and neutralization test(R=0.815,P<0.05).Furthermore,the kappa statics showed the excellent agreement between these two tests(kappa value=0.815).In addition,the coated plates were air-dried and stored for different periods(1 day,2 weeks,1,2,3,4,5 and 6 months)and the sensitivity and specificity were 100%and 80%-100%,respectively.In summary,the indirect ELISA test developed in our study would be useful as a reliable method for serological survey and control of PEDV infection and the pre-coated ELISA plates could be stored at 4°C for at least 6 months.

关 键 词：猪流行性腹泻病毒 ELISA 中和试验 预包被板 长期保存 

分 类 号：S852.651[农业科学—基础兽医学]