人源噬菌体抗体库的构建及抗D-二聚体抗体的获得  

Construction of Hominine Antibody Library and Screening of the Hominine Anti-D-dimer Antibody

作　　者：陈颖[1] 田朝伟[2] 杨晓仪[1] 贾宗剑[1] 许健[1] 黄少华[2] 陈敏生[2] 吴文言[1] 

机构地区：[1]中山大学生命科学学院分子生物学实验室,广东广州510275 [2]广州医学院第二附属医院,广东广州510260

出　　处：《中山大学研究生学刊（自然科学与医学版）》2004年第2期107-114,共8页Journal of the Graduates Sun YAT-SEN University(Natural Sciences.Medicine)

基　　金：广州市重点科技攻关项目(1999-×-006-01)

摘　　要：目前国际和国内筛选蛋白所用的技术主要是噬菌体展示技术和酵母双杂交技术。噬菌体展示技术在抗体的筛选方面已显示出良好的应用前景。本实验应用噬菌体展示技术展示出血栓病人的抗体,形成对血栓病人的抗体库。并从中淘选出能与血栓降解产物之一的 D-二聚体有特异结合作用的抗体 Fabp13。抗 D-二聚体的 Fab 在 XL1-Blue 中得到可溶表达。再利用抗 D-二聚体抗体进行竞争 ELISA 检验,证实 p13具有抗 D-二聚体的活性。从而我们筛选得到了抗 D-二聚体抗体 Fab p13。Human phage-displayed Fab library was constructed by using phase surface display technology.We selected an Fab-phage specifically interacting with D-dimer from this library.Immunnglobulin heavy and light chains genes from human lymphocytes were amplified respectively by RT-PCR using family specific primers and signal-peptide sequences of immunoglobulin as semi-nested PCR primers.The PCR resulting gene fragments were cloned into phagemid vector pComb3H and the recombinant library of human immunoglobulin is generated with helper phage VCSM13.The human anti-D-dimer Fab-phage p13 is isolated via micropanning and then successfully expressed in E. coil XL1-Blue.The success of isolating human anti-D-dimer Fab proves the practicability of phage display system in human McAb preparation.

关 键 词：D-二聚体 FAB 噬菌体展示 导向溶栓 抗体库 血栓类疾病 导向治疗 溶栓剂 

分 类 号：R543[医药卫生—心血管疾病] R96[医药卫生—内科学]