抗血管内皮细胞生长因子受体单抗的制备及鉴定

PREPARATION AND IDENTIFICATION OF MONOCLONAL ANTIBODIES AGAINST HUMAN VASCULAR ENDOTHELIAL CELL GROWTH FACTOR RECEPTOR

目的：制备血管内皮细胞生长因子受体（ｋｉｎａｓｅｉｎｓｅｒｔｄｏｍａｉｎｃｏｎｔａｉｎｉｎｇｒｅｃｅｐｔｏｒ，ＫＤＲ）的单克隆抗体并鉴定其特异性。方法：在大肠杆菌中表达重组ＫＤＲ融合蛋白。融合蛋白经凝胶分离纯化后免疫ＢＡＬＢ／ｃ小鼠，取其脾细胞，用传统杂交瘤技术与小鼠骨髓瘤细胞融合及次黄嘌呤、氨基喋呤与胸苷选择性培养。单克隆杂交瘤上清经ＥＬＩＳＡ双筛和ＳＰ免疫组化筛选并鉴定其亚类。结果：经ＥＬＩＳＡ双筛和组化筛选获得了一株能稳定分泌人ＫＤＲ单克隆抗体的杂交瘤细胞株（命名为ＳＳＷ２），其分泌抗体亚类为ＩｇＧ１，产生的腹水效价可达１×１０－６。结论：ＳＳＷ２同血管内皮细胞尤其是肿瘤组织的血管内皮细胞有较强的阳性反应。

Objective: To prepare monoclonal antibody (McAb) of the vascular endothelial growth factor receptor, kinase insert domain containing receptor (KDR) and identify its specificity. Methods: Fusion protein of KDR was expressed in E. coli. Jm109 and used for immunizing BALB/c mice. The spleen cells from immunized mice were fused with sp2/0 cells and selectively cultured with HAT medium. ELISA and S P immune histochemistry methods were used to select hybridomas. Results: One of hybridomas who secret specific monoclonal antibody against KDR was identified (named SSW2). The immunoglobulin subclass of the McAb was IgG 1, its ascite titer reaching 1×10 -6 . Conclusion: KDR McAb obtained had positive reaction with vascular endothelial cell especially in tumor tissues.