B3HM细胞免疫小鼠脾细胞scFv噬菌体展示文库的构建及表达  

The Construction and Expression of Phage Display scFv Library from the Spleen Cells of Mice Immunized With B3HM Cells

作　　者：许静[1] 张磊[1] 刁世勇[1] 刘斌[1] 孟磊[1] 姜学英[1] 韩忠朝[1] 

机构地区：[1]中国医学科学院中国协和医科大学血液学研究所实验血液学国家重点实验室,天津300020

出　　处：《中国生物工程杂志》2007年第7期12-16,共5页China Biotechnology

基　　金：国家自然科学基金资助项目(39870291)

摘　　要：目的:利用噬菌体展示技术构建B3HM细胞免疫小鼠的脾细胞表达scFv文库。方法:用人骨髓细胞系B3HM细胞免疫小鼠,取其脾细胞采用RT-PCR方法扩增VH和Vk基因并克隆入噬菌体展示表达载体,构建scFv文库,测定文库的库容量,BstNI酶切单克隆分析文库的多样性,对文库进行富集检测,鉴定单克隆噬菌体与B3HM细胞结合反应。结果:文库的库容为5×106cfu,单克隆的BstNI酶切图谱显示多样性,单克隆噬菌体抗体与B3HM细胞呈阳性反应。结论:噬菌体展示文库的成功构建为寻找新的致白血病相关基因,阐明白血病发病机理奠定了基础。To construct a scFv library by phage display technique from the spleen cells of mice immunized with B3HM cells. Three mice were immunized with B3HM cells, and their spleen cells were harvested. The genes of VH and Vk were amplified by RT-PCR from the cDNA of the immunized spleen cells and a scFv-phage display antibody library was constructed. The capacity of library was measured, and the variety of the library was analyzed by digesting with restriction endonuclease BstNI. ScFv phage clones were randomly picked and identified phage-scFv clone by binding B3HM cells using clones which showed different patterns after immunofluorescein. A scFv library containing 5 × 10^6 individual digested with restriction endonuclease BstNI was produced. Individnal phage-scFv clone showed B3HM cells positive using immunofluorescein. A scFv library of anti-B3HM cell surface molecules has been constructed. It will be useful for finding out some novel genes of causing leukemia, and establishs the infarctate foundation of clarifying the pathogenesis of leukemiagenesis.

关 键 词：B3HM细胞 噬菌体展示 抗体库 单链抗体 

分 类 号：R392.11[医药卫生—免疫学]