猪传染性胃肠炎病毒N蛋白原核表达和间接ELISA检测方法的建立

试验构建了pET28a-TGEV-N重组表达质粒，完成TGEV N蛋白的表达和纯化，通过SDS-PAGE和Western blotting分析，表明表达产物N蛋白可被猪TGEV阳性血清识别。用TGEV N蛋白作为诊断抗原，建立了检测TGEV血清抗体的间接ELISA检测方法，该方法批内、批间重复试验变异系数均小于5%；检测血清的特异性为100%，假阳性率为0，假阴性率为5%，与7种常见猪病血清无交叉反应。针对临床血清，用该方法检测结果与中和试验结果相比，符合率为100%。该方法能够快速、有效地检出TGEV抗体，重复性和特异性好，为标准化诊断试剂盒的开发奠定了基础。

Prokaryotic Expression and Development of an Indirect ELISA Detection Method of Porcine Transmissible Gastroenteritis Virus N Protein

In the study,the recombinant expression plasmid pET28a-TGEV-N was constructed,and expression and purification of the TGEV N protein was completed.Through SDS-PAGE and Western blotting analysis,the N protein could be identified by transmissible gastroenteritis virus (TGEV) positive serum.Using the TGEV N protein as diagnosis antigen,we established an indirect ELISA method for detecting TGEV serum antibodies.The coefficients of variation of intro-batch and inter-batch duplicability test were less than 5%.The specificity was 100%,and the rate of false positive and the false negative rate were 0 and 5%,respectively.No cross reactions with seven porcine diseases positive serum were detected.Using this method,the clinical serum samples were detected.The results showed 100% coincidence rate compared with neutralization test.The method could detect TGEV antibody quickly and effectively,and had good repeatability and specificity,which laid the foundation for the development of standardized diagnostic kits.