基于猪A型塞内卡病毒VP2蛋白的间接ELISA抗体检测方法的建立与应用OA

In order to establish a detection method that can be used to diagnose SVA infection in clinical practice,this study expressed VP2 protein by prokaryotic expression and established an indirect ELISA method.The results showed that pET-28a-VP2 protein was expressed in the form of inclusion body,and could react specifically with SVA antibody;and the detection method was established with VP2 protein as the coating primitive.The optimal antigen coating amount was 500 ng/well.2.5%BSA was used as blocking solution for 24h at 4℃.The optimal serum dilution ratio was 1∶100.The optimal enzyme-labeled secondary antibody dilution ratio was 1∶40 000.The optimal color develop-ment time was 15 min.The method did not cross-react with the positive serum of common porcine pathogens such as foot-and-mouth disease virus,porcine reproductive and respiratory syndrome virus,pseudorabies virus,porcine circovirus,porcine rotavirus,and swine fever virus,with a sensitivity of 1∶800 and a coefficient of variation within and between batches of less than 10%.The established method was used to test serum samples from 13 provinces in 2024,among which the positive rate of pig herd antibody was 23%,and the positive rate of pig farm antibody was 65%.SVA infection was seasonal,showing intensification in winter and spring,and subsidence in summer and autumn.