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抗肝癌单链抗体的活性检测
引用本文:孙志伟,刘彦仿,袁清安,俞炜源,黄翠芬.抗肝癌单链抗体的活性检测[J].生物技术通讯,1999,10(4):270-273.
作者姓名:孙志伟  刘彦仿  袁清安  俞炜源  黄翠芬
作者单位:1. 军事医学科学院生物工程研究所,北京,100071
2. 第四军医大学病理学教研室,西安,710032
摘    要:将抗肝癌单克隆抗体 HAb25的轻、重链可变区基因通过Linker连接起来,构建成抗肝癌单链抗体scFv25,然后亚克隆入含 E-tag检测标签的 pET15b表达载体进行融合表达。表达产物(scFv25-E-tag)采用 SABC法对肝癌细胞涂片进行染色,利用鼠抗E-tag抗体间接检测初步纯化的scFv25的活性,同时采用竞争结合实验检测scFv25的亲和活性。scFv25能够与靶细胞特异性结合,阳性部位主要定位于细胞膜上;scFv25可封闭53%的亲本抗体的亲和活性。scFv25较好地保持了亲本抗体的特异性和亲和活性。

关 键 词:肝细胞肝癌  单克隆抗体  单链抗体  E-tag

Activity assay of the single chain Fv of anti-hepatocellular carcinoma
Sun Zhiwei,LiuYanfang,Yuan Qing''an,Huang Cuifen.Activity assay of the single chain Fv of anti-hepatocellular carcinoma[J].Letters in Biotechnology,1999,10(4):270-273.
Authors:Sun Zhiwei  LiuYanfang  Yuan Qing'an  Huang Cuifen
Abstract:To construct and express mouse anti-HCC single chain Fv(scFv25) and evaluate the ac- tivity of scFv25. The HAb25 VL and VH gene were linked by a (Gly4Ser)3 polypeptide linker to form scFv25 gene, then it was subcloned into prokaryotic expression vector pET15b which con- tains E-tag to perform fusion expression. Using expression product scFv25-E-tag and anti-mouse E-tag antibody to stain HCC cell smearing slides by immunochemical methods SABC to evaluate the activity of partially purified scFv25 indirectly. Competitive inhibition trial was performed to evaluate the affinity of the scFv25 Competitive inhibition trial confirmed that the scFv25 can in- hibit 53% affinity of the antibody HAb25. scFv25 can combine with targeted cell specifically, and positive reaction position was on the membrane. scFv25 was same its native antibody in specificity and affinity.
Keywords:hepatocellular carcinoma  monoclonal antibody  single chain Fv  E-tag
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