基于重组E^(rns)蛋白的猪瘟E^(rns)-ELISA的建立和优化

Development and Optimization of an E^(rns)-ELISA Based on Recombinant E^(rns) Protein for Detection of Antibodies against Classical Swine Fever

将大肠杆菌表达的猪瘟病毒重组Erns蛋白经纯化后作为包被抗原,建立了检测猪瘟抗体的Erns-ELISA方法。经优化试验,确定了最适抗原包被量为0.15μg;血清最适稀释度为1∶100。经阻断试验、交叉反应试验和重复性试验证明,建立的Erns-ELISA简便、特异、稳定,与基于重组E2蛋白的E2-ELISA的符合率高达96.7%,与基于全病毒抗原的Dot-ELISA符合率为77.5%。可以用于猪瘟抗体的血清学监测以及用作基于E2蛋白的标记疫苗配套的鉴别诊断。

This study aims to develop a differential assay for popularizing the use of E2-based marker vaccines against classical swine fever （CSF）. Based on the recombinant E^rns produced by E.coli, an indirect ELISA （E^rns-ELISA） was established and optimized using the E^rns fusion protein as coating antigens. The optimal amount of coating antigen was optimized to be 0.15 μg, and the serum dilution was optimized to be 1：100. The established E^rns-ELISA was shown to be sensitive, specific and reproducible, and has a good agreement with E2-ELISA based on the recombinant fusion E2 protein （96.7%） and Dot-ELISA based on whole viral antigens （77.5%）. The E^rns-ELISA combined with E2-ELISA can be used as differential diagnostic assays accompanying marker or DIVA vaccines against CSF to differentiate animals infected with wild-type viruses from those vaccinated with E2-based marker vaccines.