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中国农学通报

中国农学通报 ›› 2012, Vol. 28 ›› Issue (17): 92-96.doi: 10.11924/j.issn.1000-6850.2011-3654

所属专题： 生物技术

• 畜牧 动物医学 蚕 蜂 • 上一篇    下一篇

新城疫病毒HN蛋白单克隆抗体的制备及其抗原捕捉ELISA方法的建立

田莉莉 李丽   

  • 收稿日期:2011-12-05 修回日期:2012-01-04 出版日期:2012-06-15 发布日期:2012-06-15

Preparation of Monoclonal Antibodies Against HN Protein of Newcastle Disease Virus and Establishment Antigen Capture ELISA

  • Received:2011-12-05 Revised:2012-01-04 Online:2012-06-15 Published:2012-06-15

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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NDV AC-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。

关键词: 经济发展, 经济发展, 土地利用, 保定市

Abstract: 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 κ 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.