猪传染性胃肠炎病毒重组N蛋白抗原间接ELISA抗体检测方法的建立

Development of an indirect ELISA for detection antibodies to porcine transmissible gastroenteritis virus using recombinant nucleoprotein antigen

本研究以纯化的原核表达猪传染性胃肠炎病毒N蛋白为诊断抗原,建立了猪传染性胃肠炎病毒抗体检测的间接ELISA诊断方法,将其命名为rnTGE-ELISA。该抗原不与其他常见10种猪病的阳性血清发生交叉反应,批内和批间重复性试验的变异系数均小于15%;对仔猪免疫后不同时间的血清检测结果表明rnTGE-ELISA与纯化病毒ELISA符合率达95.0%;rnTGE-ELISA相对于VN试验的敏感性为96.3%、特异性为92.2%;现地试验中,rnTGE-ELISA与SvanovaTGEV/PRCVantibodydiagnosisKit的符合率达87.0%,通过中和试验复核结果表明,rnTGE-ELISA的假阳性低于SvanovaTGEV/PRCVantibodydiagnosisKit。本试验建立的rnTGE-ELISA诊断方法具有良好的敏感性和特异性,为免疫猪群抗体监测和TGE流行病学调查提供了一种快速、简便的血清学诊断方法。

Using the recombinant and purified N protein as antigen expressed in Escherichia coli BL21, an indirect ELISA was successfully developed to detect antibody to transmissible gastroenteritis virus （TGEV）, the indirect ELISA was named rnTGE-ELISA.The recombinant N protein antigen showed no cross-reaction with the positive sera of other ten kinds of swine diseases, coefficient of variability percent（C. V %） of intro-batch duplicativity test and inter-batch duplicativity test was less 15 %; Result of immune piglets were tested with rnTGE-ELISA and purified virus ELISA showed that 95.0 % concordance was obtained; Sensitivity and specificity of rnTGE-ELISA relative to VN was respectively 96.3 % and 92.2 %; in field test rnTGE-ELISA compared with Svanova TGEV/PRCV antibody diagnosis Kit, 87.0 % concordance was obtained. Therefore,this rnTGE-ELISA based on recombinant N protein antigen has good sensitivity and specificity, can afforded a simple and rapid means for assessment of vaccination in the field and investigation of TGE epidemiology.