摘要
目的进行ＴＮＦ－α单抗Ｆａｂ段基因的克隆并在大肠杆菌中表达。方法用逆转录－聚合酶链反应技术（ＲＴ－ＰＣＲ）从分泌抗人ＴＮＦ－α的鼠单抗杂交瘤细胞系中克隆重链Ｆｄ段和κ链基因；并用ＥＬＩＳＡ和免疫印迹分析证明表达的Ｆａｂ段特异结合ＴＮＦ－α。结果从分泌抗人ＴＮＦ－α的鼠单抗杂交瘤细胞系中克隆出了重链Ｆｄ段和κ基因。经ＤＮＡ序列测定表明，ＶＨ、Ｄ、ＪＨ分别属于ＶＨ３Ｄ、ＤＳＰ２和ＪＨ４；Ｖκ和Ｊκ分别属于Ｖκ１和Ｊκ１。将该Ｆｄ和κ链ｃＤＮＡ克隆到表达载体ｐＣｏｍｂ３Ｈ中，在大肠杆菌中获得了表达。结论ＥＬＩＳＡ和免疫印迹分析表明，表达的Ｆａｂ段可特异地和ＴＮＦ－α结合。
Fd and κ gene from a murine hybridoma secreting antiTNFα monoclonal antibody was cloned by RTPCR. Sequencing analysis showed that the VH,D and JH were derived from VH3D,DSP2 and JH4,respectively, and the Vκ and Jκ from Vκ 1 and Jκ1.Fd and κ gene were cloned into the expressing vector pComb3H and expressed in E.coli.ELISA and Western blotting analysis demonstrated that bacterial expressed Fab specifically bound to recombinant human TNFα.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
1998年第2期149-151,共3页
Chinese Journal of Microbiology and Immunology
基金
国家科委863计划
