摘要
为建立一种快速的新城疫病毒病原检测方法。本研究以原核表达的重组HN蛋白免疫7周龄BALB/c雌鼠,取其脾细胞与骨髓瘤细胞SP2/0进行融合,经间接ELISA方法筛选,成功获得了1株能稳定分泌抗新城疫病毒HN蛋白的McAb杂交瘤细胞,命名为4E8,4E8亚类鉴定为重链属于IgG2a,轻链属于κ链。以多克隆抗体作为包被抗体、单克隆抗体4E8作为检测抗体,通过双抗夹心ELISA各个反应条件的优化,建立检测新城疫病毒抗原捕捉ELISA方法。该方法对EDSV、ITLV、IBV、IBDV不发生交叉反应,其敏感性比HA试验要高4倍以上,与RT-PCR相比较,符合率、敏感性和特异性分别为95.6%、93.3%、96.1%。本研究建立的NDVAC-ELISA有良好的重复性、敏感性和特异性,可应用于新城疫病毒感染的早期诊断。
To produce a rapid Newcastle disease virus （NDV） pathogen detection method. BALB/c mice were immunized with HN recombination protein and the mouse splenic cells were fused with SP2/0 cells, hybridoma cell stably secreting anti-HA McAb was screened by ELISA designated 4F8, the immunoglobulin type of McAb identification shown was IgG2a type with K chain. An antigen capture ELISA （AC-ELISA） was developed for detection of NDV using polyclonal antibody as capture antibody and specific monoclonal antibody 4F8 as detecting antibody. The AC-ELISA showed no cross-reaction with other four avian viruses （EDSV, ITLV, IBV, IBDV）, The sensitivity four times higher than HA test, Comparing with RT-PCR, the concordance, sensitivity, specificity was 95.6%, 93.3%, 96.1%, respectively. The NDV AC-ELISA had good specificity, sensitivity and repeatability, could be used for diagnosing NDV infections.
出处
《中国农学通报》
CSCD
2012年第17期92-96,共5页
Chinese Agricultural Science Bulletin
