猪瘟抗体间接ELISA检测方法的建立和优化

Development and optimization of the indirect ELISA method to detect antibody against classical swine fever virus

采用真核系统表达纯化的猪瘟病毒E2蛋白作为包被物,通过对各个反应条件的摸索和优化,建立了检测猪瘟病毒抗体的猪瘟抗体间接ELISA检测方法。研究结果表明,E2蛋白的最佳包被浓度为0.25μg/mL,最佳包被条件为4℃16h;待检血清的最佳稀释倍数为100倍;特异性检测试验证明,该方法与猪常见病原的阳性血清没有交叉反应。通过对大量猪瘟抗体阴、阳性血清的检测,最终确定了本检测方法的判定标准。

Classical swine fever virus（CSFV） E2 protein was expressed in BAC-To-BAC system.After purification,E2 protein was coated to the ELISA plate.Through a series of optimization tests,the indirect ELISA for CSFV antibody was established.The results showed that the optimal concentration of E2 protein for coating was 0.25 μg/mL and coating condition was 4℃ for 16 h.The proper dilution of serum sample was 1∶100.And the specificity tests showed that the purified E2 protein had higher specificity for CSFV antibody in serum.Hundreds of samples were detected by the ELISA and standard of criterion was established by data analysis.