抗肝癌hdsFv融合RC-RNase重组免疫毒素的表达、纯化及活性测定  

Expression, purification and functional detection of anti-HCC hdsFv fused with RC-RNase

作　　者：付勇[1] 苏雪梅[2] 刘彦仿[3] 杨守京[3] 赵君[3] 邹赛英[1] 

机构地区：[1]兰州军区乌鲁木齐总医院病理科,830000 [2]兰州军区乌鲁木齐总医院妇产科,830000 [3]第四军医大学基础部病理学教研室

出　　处：《肿瘤防治研究》2006年第9期626-628,700,共4页Cancer Research on Prevention and Treatment

基　　金：国家自然科学基金资助项目(30370810)

摘　　要：目的制备有临床应用前景的特异性高、稳定性强的新型抗肝癌重组免疫毒素。方法利用IPTG诱导抗肝癌hdsFv-RC-RNase重组免疫毒素在大肠杆菌中表达。表达产物经Ni-NTA亲合层析法纯化并复性,应用免疫细胞化学和MTT的方法分别检测其对人肝癌细胞的特异性结合和杀伤活性。结果重组免疫毒素以可溶性形式在大肠杆菌中获得表达。免疫细胞化学和细胞毒实验表明其能够特异性地结合并杀伤肝癌细胞,并且能够保持较强的稳定性。结论我们抗肝癌hdsFv-RC-RNase重组免疫毒素的成功制备,为其进一步的临床应用研究奠定了一定的实验基础。Objective To obtain a new kind of anti-HCC recombinant immunotoxin, which has potentialities for clinical application, high specificity and increased stability. Methods The prokaryotic expression vector pTIH-hdsFv RC-RNase was induced to express in E. coli by IPTG. The expressed product was purified by Ni-NTA agarose affinity chromatography under native conditions and mildly refolded. The immunocytochemical staining and MTT colorimetry were used respectively to detect the specifically binding activity and killing effect of the recombinant immunotoxin. Results The fusion gene hdsFv-RC RNase was induced to express effectively in soluble form in E. coli. The purity of the expressed product was near to homogeneity. The recombinant immunotoxin has the activity to bind specifically and kill obviously the HCC cells. The recombinant irnmunotoxin could maintain high stability after stored at 4℃ for 3 months. Conclusion The recombinant immutoxin of anti-HCC hdsFv-RC-RNase was prepared successfully, which lay the foundation for the further research on its clinical application.

关 键 词：肝细胞癌 免疫毒素 二硫键稳定单链抗体 牛蛙核糖核酸酶 

分 类 号：R735.7[医药卫生—肿瘤]