特异性O1群稻叶型霍乱弧菌单克隆抗体的制备、筛选及识别抗原表位的分析

目的制备并筛选出高特异性高活性的抗01群稻叶型霍乱弧菌（Vibrio cholerae O1 Serotype Inaba）单克隆抗体（monoclonal antibody，McAb），为霍乱的预防诊断提供有力的抗体工具。方法应用杂交瘤技术，通过灭活的O1群稻叶型霍乱弧菌免疫的BALB／c小鼠的脾细胞与SP2／O细胞融合制备特异性McAb，以间接ELISA法对所需的杂交瘤细胞株进行了特异性及排除性筛选，对筛选出的特异性McAb进行了包括亚型鉴定，效价、亲和常数的测定以及特异性的鉴定与分析，并通过单克隆抗体的位点相加实验、与O1群稻叶型霍乱弧菌脂多糖（LPS）的间接ELISA和Western blot实验对McAb识别的抗原表位进行了初步分析。结果融合了386株能分泌抗01群稻叶型霍乱弧菌McAb的杂交瘤细胞株，通过用O1群小川型、0139霍乱弧菌及9种非霍乱弧菌的细菌进行的筛选，最后得到4株能稳定分泌特异性的针对该McAb的细胞株，其分泌的抗体亚类分别为2株IgG1，1株IgG2，1株IgG2b；腹水效价均达10^-6；亲和常数达10^8以上。间接ELISA法及Western blot证实所获的McAb可与O1群稻叶型霍乱弧菌发生特异性反应。ELISA相加实验结果显示4株McAb识别不同的抗原表位，与LPS的反应表明其中2株针对O1群稻叶型霍乱弧菌脂多糖，2株针对非脂多糖抗原位点。结论获得霍乱弧菌O1群稻叶型特异性McAb，初步定位单克隆抗体识别表位所在区域。

Preparation and characterization of monoclonal antibodies against Vibrio cholerae O1 Serotype Inaba and analysis on targeting antigenic epitopes of the monoclone antibodies

Objective To prepare and select monoclonal antibodies (McAbs) with high specificity against Vibrio cholera O1 Serotype Inaba and to provide a potent medical tool for the prevention and diagnosis of cholera. Methods BALB/c mice were immunized with Vibrio cholerae O1 Serotype Inaba. Hybridoma cell lines secreting monoclonal antibodies against Vibrio cholerae O1 Serotype Inaba were established after cell fusion of mouse splenic cells and SP2/0. Both of these McAbs were characterized for their antibody titre in...