抗人死亡受体5单链抗体的构建表达纯化及活性分析  

The preparation and tumor cell apoptosis-inducing activity assay of anti-human DR5 single-chain antibody

作　　者：张佳锴[1,2] 程小峰[1] 庄国洪[1] 

机构地区：[1]厦门大学医学院抗癌研究中心,福建厦门361005 [2]厦门大学生命科学学院,福建厦门361005

出　　处：《中国生化药物杂志》2012年第2期97-100,105,共5页Chinese Journal of Biochemical Pharmaceutics

基　　金：福建省自然科学基金资助项目(C0710046)

摘　　要：目的构建、表达、纯化抗人死亡受体5(DR5)单链抗体,并检测其诱导肿瘤细胞凋亡的活性。方法采用RT-PCR获取鼠源性抗人DR5单克隆抗体重链和轻链可变区基因序列,以一柔性连接肽连接二者,转入表达载体,以大肠杆菌表达融合蛋白,亲和色谱纯化后用MTT实验和凋亡检测试剂盒检测其凋亡诱导活性。结果获得的序列经比对为抗体重链和轻链可变区基因,表达纯化后的重组蛋白具有接近完整抗体的肿瘤细胞凋亡诱导活性。结论抗人DR5 scFv可作为诱导肿瘤细胞凋亡的候选药物,为肿瘤免疫学研究提供材料。Purpose To construct,express,purify anti-human DR5 scFv,and to assay its activity of apoptosis induction for tumor cell lines.MethodsVariable region sequences of heavy chain and light chain to murine anti-human DR5 monoclonal antibody were acquired by RT-PCR,then they were linked through a flexible linker peptide and was cloned into the expression vector.After being expressed by E.coli and purified using affinity chromatography,the recombinant protein was employed to MTT assay as well as apoptosis assay kit in order to detect its apoptosis-inducing activity.ResultsThe sequences we′ve got were established as the variable region genes of antibody′s heavy and light chain,while the recombinant proteins expressed and purified possessing the apoptosis-inducing activity come near to complete antibody.ConclusionAnti-human DR5 scFv can supply material for tumor immunology research as drug candidate inducing tumor cell apoptosis.

关 键 词：死亡受体5 单克隆抗体 SCFV 细胞凋亡 

分 类 号：Q78[生物学—分子生物学]