人脑心肌炎病毒免疫球蛋白M捕获酶联免疫吸附试验的建立及初步应用

Establishment and preliminary application of EMCV IgM capture ELISA method

目的建立脑心肌炎病毒(EMCV)免疫球蛋白M(IgM)捕获酶联免疫吸附试验(ELISA)检测方法,用于感染EMCV血清学早期诊断。方法用抗人IgM(μ链)单抗进行包被,辣根过氧化物酶(HRP)标记的EMCV为酶标抗原,初步建立EMCV IgM捕获ELISA;对建立的方法进行条件优化及特异性、精密性、稳定性等验证。结果建立的EMCV IgM捕获ELISA检测方法抗体包被浓度为1μg/mL,EMCV-HRP工作浓度和反应时间分别为1∶400和45min,封闭液为10%马血清时该法检测效果可达最佳,3批试剂批内及批间变异系数均小于5%,且特异性、精密性及稳定性较好。结论建立的EMCV IgM捕获ELISA法特异、精密且稳定,可用于人感染EMCV的早期检查,为EMCV的诊断奠定基础。

Objective To establish the encephalomyocarditis virus（EMCV） immunoglobulin M （IgM ） captured enzyme‐linked immunosorbent assay （ELISA ） detection method for the application in early serological diagnosis of EMCV infection .Methods Anti‐human（μ chain） monoclonal antibody was used to conduct the coating and the horse radish peroxidase（HRP） labelled EMCV served as the enzyme labeled antigen ,the EMCV IgM capture ELISA meth‐od was preliminarily established ;The established method was performed the conditional optimization and verification of the specificity ,precision and stability .Results The antibody coating concentration in the established EMCV IgM captured ELISA method was 1 μg/mL ,the EMCV‐HRP work concentration and reaction times were 1 ∶ 400 and 45 min respectively .When the blocking solution was 10% horse serum ,this method achieved the best effect .The intra‐assay and inter‐assay coefficients of variation in 3 batches of reagents were less than 5% ,and the specificity ,precision and stability were better .Conclusion The established EMCV IgM capture ELISA method is specific ,precise and sta‐ble ,can be used for the detection of early human EMCV infection and lays the foundation for the EMCV diagnosis .