兔支气管败血波氏杆菌单克隆抗体的制备及鉴定

目的建立能稳定分泌抗兔支气管败血波氏杆菌（Bb）的单克隆抗体杂交瘤细胞株,为今后进一步建立该菌的免疫检测技术奠定基础。方法以Bb分离株BLJ05的灭活菌液为免疫原,腹腔免疫BALB/c小鼠,采用常规杂交瘤技术制备Bb单克隆抗体（McAb）,用间接ELISA、Western-blot等方法对McAb特性进行鉴定。结果获得两株能稳定分泌抗Bb单克隆抗体的杂交瘤细胞株,分别命名为A7D5和D6B2,其小鼠腹水抗体效价分别为1∶409600和1∶102400;且不与兔大肠杆菌、多杀性巴氏杆菌、产气荚膜梭菌等兔的常见病原菌反应,特异性强。两株单抗亲和力实验表明A7D5亲和力略高于D6B2。ELISA相加试验表明它们针对相同的抗原表位。结论成功建立了两株能稳定分泌抗兔支气管败血波氏杆菌单克隆抗体的杂交瘤细胞株,效价高、特异性强,为今后建立该菌的免疫检测技术建立奠定了基础。

Preparation and Identification of Monoclonal Antibodies against Bordetella bronchiseptica from Rabbits

Objective To prepare the special monoclonal antibodies against Bordetella bronchiseptica from rabbits, and to establish a good immune diagnose methods for further research. Method BALB/c mice were immunized by using the immunogen of Bb BLJ05 which was inactivated by methanal. The monoclonal antibodies were prepared by using the traditional hybridoma technique, and the biology characteristic of each monoclonal antibody was identified by indirect enzyme linked immunosorbont assay （ELISA）, western-blot analysis and so on. Result Two strains of hybridoma which named A7 D5 and D6 ]32 were obtained. The ascites titers of Bb specific ELISA antibodies obtained by mice were 1：409600 and 1 ： 102400. The specificity identification revealed that the monoclonal antibodies did not react with E. coli, P. psicicida and Cl. Perfringens. Moreover, the afllnitv of A7 D5 was tighter than that of D6 B2, and the results of epitope analysis with addition ELISA revealed that the two MeAbs could recognize the same epitope. Conclusion The McAbs with high ELISA titer and good specificity were successfuUy prepared.