狂犬病病毒磷蛋白单抗的制备与应用

摘要

Four monoclonal antibodies against Rabies virus phosphoprotein were produced by immunization with Rabies virus strain CVS in BALB/c mice. After fusion, antibodies in the supernatant of the hybrid cells were respectively detected by ELISA test and western-blot method. Four hybrid cells secreting antibody binding to rabies virus , i.e. 1C9, 4B10, 2G12 and4G5 strains. The 1C9 monoclonal lgG antibody was purified by affinity chromatography method and then labeled with FITC. 501 brain samples collected from ferret badgers, bats, dogs and weasels were detected using the labeled 1C9 antibody and FITC-labeled rabies virus N protein-specific monoclonal antibody (made in our laboratory). The results showed that the labeled 1C9 antibody can detect rabies-related lyssaviruses isolated in China including Rabies virus and Irkut virus, however, the N protein-specific monoclonal antibody only can detect Rabies virus. It suggests that the labeled 1C9 antibody can be used as a rabies diagnostic reagent in China.%以灭活的狂犬病病毒CVS株细胞毒免疫BALB/c小鼠，通过间接ELISA法和Western-blot筛选获得针对磷蛋白的单克隆抗体4株：1C9、4B10、2G12、4G5，其中1C9针对氨基端保守表位。以亲和层析法纯化1C9单抗腹水，异硫氰酸荧光黄标记制备荧光抗体。以1C9磷蛋白荧光抗体与本实验室研制的狂犬病核蛋白免疫荧光抗原检测试剂盒，对本实验室收集的501份疑似狂犬病鼬獾、蝙蝠、犬和黄鼬的脑组织样品进行直接免疫荧光平行检测。结果显示，两种检测手段对基因1型狂犬病毒的检出结果完全一致，而蝙蝠源Irkut病毒仅能以磷蛋白单抗1C9检出。本研究成功获得了与我国现有不同基因型狂犬病毒良好反应的抗狂犬病磷蛋白单抗，并应用于狂犬病的直接免疫荧光检测，为狂犬病诊断提供了敏感性和可靠性良好的诊断试剂。