B3HM细胞免疫小鼠脾细胞scFv噬菌体展示文库的构建及表达

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。

The construction and expression of phage display scFv library from the spleen cells of mice immunized with B3HM cells

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 immunofluorescein. We produce a scFv library containing 5×106 individual clones which showed different patterns after digested with restriction endonuclease BstNI. 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.