A型塞内卡病毒3AB蛋白原核表达及间接ELISA方法的初步建立OA北大核心

This study was aimed to establish an indirect ELISA method for detecting the antibodies against non-structural protein(NSP)3AB of Seneca virus A(SVA),the recombinant protein of SVA 3AB was expressed by using a prokaryotic expression system.Using the recombinant protein of SVA 3AB as the coating antigen,an indirect ELISA detection method was established.The results showed that the re-combinant protein of SVA 3AB was expressed in a soluble form with a size of 18 kDa and could specifical-ly react with SVA antibodies.The results of established indirect ELISA method showed that:the optimal coating concentration of antigen was 1 µg/mL;the best blocking solution was 3％skimmed milk powder;the optimal dilution ratio of the serum sample was 1:640;the optimal dilution ratio of the enzyme-la-beled antibody was 1∶10 000;and in the darkness development time was 10 minutes.This ELISA method did not cross-react with positive sera of porcine foot-and-mouth disease virus(FMDV),porcine pseudora-bies virus(PRV),porcine reproductive and respiratory syndrome virus(PRRSV),classical swine fever virus(CSFV),and porcine circovirus(PCV);the sensitivity reached 1∶6 400;the intra-batch coefficient of variation was less than 8％,and the inter-batch coefficient of variation was less than 10％.This in-direct ELISA method has strong specificity,high sensitivity,and good repeatability.It can be applied or clinical detection of pig serum samples and lays a foundation for the diagnosis of SVA.