鸡传染性支气管炎病毒M蛋白单克隆抗体的制备及鉴定  

Preparation and identification of monoclonal antibodies against M protein of avian infectious bronchitis virus

作　　者：王富妍[1] 任雅 邹年莉[1] 郭明萍[1] 吴瑞婷[1] 付润饶[1] 文心田[1] 曹三杰[1] 黄勇[1] 

机构地区：[1]四川农业大学动物医学院动物预防医学生物工程实验室,四川雅安625014 [2]四川省雅安市天全县农业局,四川雅安625500

出　　处：《中国生物制品学杂志》2013年第10期1463-1466,共4页Chinese Journal of Biologicals

摘　　要：目的制备鸡传染性支气管炎病毒(infectious bronchitis virus,IBV)M蛋白单克隆抗体,并进行鉴定。方法利用IBV Sczy3株免疫BALB/c小鼠,取其脾细胞与SP2/0骨髓瘤细胞融合,间接ELISA法筛选杂交瘤细胞,强阳性孔经有限稀释法亚克隆,制备抗IBV M蛋白单克隆抗体。对制备的单抗进行单抗亚型、腹水ELISA效价、特异性、交叉反应性鉴定及Western blot分析。结果获得2株能稳定分泌抗IBV M蛋白单克隆抗体的杂交瘤细胞株,命名为2C10和7H6,单抗亚型分别为IgM和IgG1;腹水ELISA效价分别为1∶25 600和1∶12 800;2株单抗均只与IBV发生特异性反应,而与新城疫病毒(Newcastle disease virus,NDV)、禽流感病毒(avian influenza virus,AIV)H9亚型、AIV H5抗原和鸡传染性法氏囊病毒(infectious bursal disease virus,IBDV)均不反应;2株单抗与5种基因型的其他10株IBV均有良好的反应性。结论制备出2株针对IBV M蛋白的单抗,为今后更深入地研究该蛋白的分子生物学功能、抗原表位的鉴定及抗原捕获ELISA方法的建立等提供了有力的工具。Objective To prepare and identify the monoclonal antibodies （McAbs） against M protein of avian infectious bronchitis virus （IBV）. Methods BALB/c mice were immunized with IBV Sczy3 strain, of which the spleen cells were fused with myeloma SP 2 / 0 cells. Hybridoma cells were screened by indirect ELISA, of which the strong positive ones were further subcloned by limiting dilution method. The prepared McAbs were identified for subtype, ELISA titer in ascites, specificity and cross reactivity, and analyzed by Western blot. Results Two hybridoma cell strains stably secreting McAbs against IBV M protein were obtained and named as 2C10 and 7H6, of which the subtypes were IgM and IgG1, and ELISA titers in ascites were 1：25 600 and 1：12 800, respectively. Either of the McAbs showed specific reaction only with IBV, which showed no reactions with Newcastle disease virus （NDV）, avian influenza virus （AIV） subtype H9, AIV H5 or avian infectious bursal disease virus （IBDV）. The two MeAbs showed good reactivity with other ten IBV strains of five genotypes. Conclusion Two McAbs against IBV M protein were prepared, which provided a powerful tool for further study on the molecular biological function and identification of antigenic epitope of the protein and development of antigen-capture ELISA for IBV.

关 键 词：鸡传染性支气管炎病毒 M蛋白 单克隆抗体 酶联免疫吸附测定 

分 类 号：S858.31[农业科学—临床兽医学] R392-33[农业科学—兽医学]