牛病毒性腹泻病病毒NS3表位串联蛋白表达及抗体间接ELISA方法的建立

Development of Indirect ELISA of Bovine Viral Diarrhea Virus by Using Tandem Epitopes of NS3

为建立一种有效的牛病毒性腹泻病病毒(Bovine viral diarrhea virus,BVDV)抗体检测方法,作者将BVDVNS3蛋白2个抗原表位的编码核酸序列进行串联,构建重组表达载体pET-30a-EXnN,并在原核表达系统中表达了重组蛋白。选取包含12个表位肽的重组蛋白(r-BVDV-EX6N)作为包被抗原建立NS3蛋白抗体间接ELISA方法。该方法的特异性和稳定性良好,与2种商品化试剂盒的符合率分别为96.05%和78.95%。试验结果表明建立的ELISA方法可用于BVDV NS3蛋白抗体的检测。

The present experiment was performed to establish an indirect ELISA for detection of antibodies against Bovine viral diarrhea virus（BVDV）.Tandem epitopes of NS3 of BVDV was cloned into pET-30a and expressed in E.coli Rosetta.SDS-PAGE analysis revealed a band of protein correspondent with molecular weight of the target protein,and the result of western blot showed that the recombinant protein was recognized specifically by anti-BVDV positive serum.An indirect ELISA（r-BVDV-EX6N-ELISA）was developed using purified protein which contains 12 epitopes as coating antigen to detect BVDV antibodies in cattle.The assay was highly specific and showed no cross-reaction with positive sera of other bovine diseases.Comparison with two commercial kit showed a coincidence rate of 96.05% and 78.95%.The results demonstrated that the indirect ELISA established in this study works well in BVDV detection.