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目的制备整合素阻断剂AP25的单克隆抗体,鉴定其效价和特异性。方法从抗原免疫BALB/c小鼠分离免疫脾细胞,通过杂交瘤技术将免疫脾细胞与小鼠骨髓瘤细胞SP2/0融合,筛选杂交瘤细胞株并进行扩大培养。采用小鼠体内诱生法制备腹水,进行纯化后得到抗AP25单克隆抗体,用ELISA和Western blot鉴定其效价和特异性。结果 ELISA检测抗AP25单克隆抗体效价可达1∶2×106,Western blot鉴定抗体特异性良好。结论获得了能够高表达抗AP25单克隆抗体的杂交瘤细胞株;腹水制备抗体,纯化后得到效价高、特异性良好的AP25单克隆抗体。
Objective To prepare a monoclonal antibody against integrin blocker AP25 and identify its potency and specificity. Methods Immunized splenocytes were isolated from BALB / c mice immunized with antigen. The immunized spleen cells were fused with mouse myeloma cells SP2 / 0 by hybridoma technique. The hybridoma cell lines were screened and expanded. The ascites was prepared by in vivo inoculation in mice, and the anti-AP25 monoclonal antibody was obtained after purification. The titer and specificity of the anti-AP25 monoclonal antibody were determined by ELISA and Western blot. Results The titer of McAb against AP25 was up to 1: 2 × 106 by ELISA, and the antibody specificity was good by Western blot. Conclusion The hybridoma cell line which can express anti-AP25 monoclonal antibody was obtained. The ascites was used to prepare the antibody. The AP25 monoclonal antibody with high titer and good specificity was obtained after purification.