【摘要】 目的:构建能同时特异性靶向人骨肉瘤细胞表面肿瘤相关抗原TP-3及人T细胞表面TCR/CD3复合体CD3ε-链的双特异性单链抗体anti-CD3×anti-TP-3 diabody,对其生物学特性进行研究。方法:通过抗体可变区基因构建表达质粒pKIDCD3×TP-3,继而转染大肠杆菌K12系XL1-Blue株,表达活性抗体diabody,通过固定化金属离子亲和层析法纯化,流式细胞术、蛋白电泳进行特异性及亲和力的测定,并将其与人T细胞、骨肉瘤细胞共同培养,观察其诱导T细胞杀伤肿瘤细胞的活性。结果:diabody纯度高,在人血清中较为稳定,活性强,能够同时特异性地结合人骨肉瘤细胞及人T细胞,且将二者的距离拉近,特异性地溶解靶细胞。结论:双特异性单链抗体diabody能充分诱导人细胞毒T细胞活性,使骨肉瘤细胞溶解。更多还原

【Abstract】 Objective:To construct a bispecific single chain antibody fragment anti-CD3×anti-TP-3 diabody with dual specificity for both the human osteosarcoma cell antigen TP-3 and CD3ε-chain of CD3/TCR.Methods: The expression plasmid pKIDCD3 × TP-3 was constructed with gene of antibody variable fragment,and then it was transinfected into the E.coli K12 strain XL1-Blue to obtain diabody. The diabody was purified by IMAC and its affinity was determined by flow cytometry and PAGE,at last,it was incubated with human T cell and osteosarcoma cell.Results: The diabody was absolute and stable in human plasma with high specificity for human T cells and osteosarcoma cells,and it also makes T cells approach target cells and lyse them.Conclusion: The diabody not only stimulates and retargets CTLs to osteosarcoma cells but also lyses them.更多还原