抗人DR5单克隆抗体的制备、鉴定及活性分析

Preparation,characterization,and activity analysis of monoclonal antibodies against human death receptor 5

目的制备抗人DR5单克隆抗体(mAb),鉴定其特性,并进行生物学活性分析。方法以纯化的可溶性DR5(sDR5)免疫Balb/c小鼠,杂交瘤技术制备抗人DR5mAb;运用ELISA、SDS-PAGE电泳方法测定抗DR5mAb与sDR5结合的特性;Ig亚类ELISA试剂盒鉴定抗DR5mAb亚类;间接ELISA法检测腹水mAb效价;流式细胞仪检测肿瘤细胞表面DR5的表达水平;流式细胞仪检测抗DR5单克隆抗体(mAb)诱导肿瘤细胞凋亡的功能。结果获得1株可分泌抗DR5mAb的杂交瘤细胞系R150。SDS-PAGE电泳检测证实,获得的R150可特异性地识别DR5;R150的Ig亚类为IgGI(λ型);腹水效价为1×106;通过流式细胞仪可敏感地检测到肿瘤细胞表面DR5的表达水平及R150诱导肿瘤细胞凋亡情况。结论获得1株可分泌抗DR5mAb的细胞系R150,抗体具有效价高、特异性强等特点并能有效诱导肿瘤细胞凋亡,具有较好的应用价值。

Objective To prepare monoclonal antibodies (mAbs) against DR5,and characterize their biological properties by using ELISA,flow cytometry,and SDS-PAGE. Methods Balb/c mice were immunized with soluble DR5 (sDR5),and then mAbs was prepared by hybridoma technique. Ig subclass and specificity of mAbs were analyzed by ELISA and SDS-PAGE,respectively. The titres of mAbs in ascitic fluid were determined by indirect ELISA. Flow cytometry was utilized to detect the tumor cell apoptosis-inducing activity of the mAbs a...