猪脑心肌炎病毒重组抗原间接ELISA诊断方法的建立与应用

以猪脑心肌炎病毒VP1重组蛋白为抗原,建立了检测猪脑心肌炎病毒(EMCV)血清抗体的间接ELISA诊断方法。经优化后抗原最适包被浓度为2μg/mL,血清最适稀释度为1∶50,其作用时间为90 min,酶标抗体最适稀释度为1∶20 000,最适作用时间为30 min。判定标准为OD450≥0.427为阳性,OD450≤0.35为阴性,介于二者之间为可疑。该重组蛋白与PRRSV、猪瘟、PCV2、FMDV抗体反应呈阴性,证明具有良好的特异性。应用该方法检测了来自我国不同地区的26家猪场的156份临床血清,结果证明我国部分规模化猪场已有猪脑心肌炎疾病存在。

Development of an indirect ELISA with recombinant antigen for detecting antibody against Encephalomyocarditis virus

An indirect ELISA for detecting Encephalomyocarditis virus (EMCV) antibodies was developed with the recombinant VP1 protein used as antigen.The optimized concentration of coated antigen was 2μg/mL.The serum samples were diluted in 1∶50 and incubated for 90 minutes.The dilution of conjugate was 1∶20000,and the reaction time was 30 minutes.The standard of validation was that the serum samples with OD450≥0.427 were positive and the samples with OD450≤0.35 were negative.The recombinant antigen had no cross reaction with other 4 antibodies to porcine diseases.This indirect ELISA was used to detect EMCV antibody in 156 serum samples from26 pig farms.The results showed that EMCV existed in some scaled pig farms of China.