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目的制备抗流感嗜血杆菌外膜蛋白P6的单克隆抗体(mAb)并进行鉴定。方法以重组P6蛋白腹腔免疫BALB/c小鼠,常规细胞融合技术制备杂交瘤细胞株;间接ELISA筛选阳性细胞克隆并检测染色体数目;测定细胞培养上清的mAb效价及特异性,鉴定mAb的免疫球蛋白类、亚类及抗原结合表位。结果获得2株抗流感嗜血杆菌外膜蛋白P6的杂交瘤细胞株α2G3和γ2C4,染色体众数分别为103与95;间接ELISA结果证明细胞培养上清最高效价分别为1∶256和1∶512,未出现与其他种类细菌的交叉反应;α2G3为IgG2b、γ2C4为IgM,均为κ型;两者可能识别P6蛋白不同表位。结论成功制备了抗流感嗜血杆菌外膜蛋白P6的mAb。
Objective To prepare monoclonal antibodies against Haemophilus influenzae outer membrane protein P6 (mAbs) and identify them. Methods BALB / c mice were immunized intraperitoneally with recombinant P6 protein, and hybridoma cell lines were prepared by conventional cell fusion technique. The positive clones were screened by indirect ELISA and the number of chromosomes was determined. The titer and specificity of mAb in cell culture supernatants were determined. Immunoglobulins, subclasses and antigen binding epitopes. Results Two hybridoma cell lines, α2G3 and γ2C4, which were resistant to Haemophilus influenzae outer membrane protein P6 were obtained. The chromosome numbers were 103 and 95, respectively. The indirect ELISA showed that the highest titer of the cell culture supernatant was 1:256 and 1: 512, no cross-reaction with other species of bacteria; α2G3 is IgG2b, γ2C4 is IgM, are κ; both may identify different epitopes P6 protein. Conclusion The mAb against H. influenzae outer membrane protein P6 was successfully prepared.