【摘要】 以抗癌胚抗原 （Carcinoembryonicantigen ,CEA）单链抗体与假单胞菌外毒素 （PseudomonasexotoxinA ,PEA）的截短和修饰形式PE38 KDEL构建重组免疫毒素CEA PE38 KDEL ,并在大肠杆菌菌株BL2 1（DE3）_star中表达。采用镍离子螯合层析法纯化变性的包涵体样品 ,并用连续梯度透析的方法对纯化后的包涵体进行复性。采用流式细胞术鉴定复性产物与靶细胞的结合活性 ,结果表明免疫毒素CEA PE38 KDEL具有与靶细胞特异性结合的活性。以MTT法检测免疫毒素对肿瘤细胞的体外杀伤活性 ,结果表明该免疫毒素对SW1116和CNE₂细胞具有特异性杀伤活性。证明了经包涵体复性的抗CEA免疫毒素CEA PE38 KDEL对表达CEA抗原的肿瘤细胞具有良好的结合和杀伤活性。更多还原

【Abstract】 A recombinant immunotoxin named CEA/PE38/KDEL was constructed, which was composed of anti-CEA single-chain Fv and the truncated and modified form of Pseudomonas exotoxin (PE38/KDEL). The CEA/PE38/KDEL immunotoxin was expressed in the E.coli strain BL21(DE3)-star as inclusion bodies. The denatured inclusion bodies were purified with Ni-NTA chelate agarose, then the constant gradient dialysis was used to perform the refolding of the CEA/PE38/KDEL immunotoxin. Results of FACS and MTT assay indicate that the refolded immunotoxins keep potent and specific cytotoxicity to tumor cells bearing CEA antigens.更多还原