抗CD_3基因工程嵌合抗体片段Fab′的构建、表达和活性测定

Construction and Expression of Anti-CD_3 Chimeric Antibody Fab′ Fragment in Ecoli

PCR扩增抗CD3单抗轻链可变区 (VL)和重链可变 (VH)片段基因 ,将其重组到Fab′表达载体中 ,构建成抗CD3嵌合抗体Fab′表达载体 ,转化大肠杆菌 16C9进行可溶性表达。产物经蛋白G亲和层析柱纯化。免疫荧光竞争结合实验和3H掺入实验证实能与小鼠抗CD3IgGHIT3a竞争性结合表达CD3的T淋巴细胞 。

WT5BZ]The gene of variable region of heavy chain(V Ⅱ)and light chain (V L) were amplified from the expression vector of anti CD 3 single chain antibody (Scfv) into the vector sspB11 and sspB12,respectively.The Fab′ fragment expression vector was constructed by cloning the gene of VL+CL and VH+CH 1 into expression vector PAYZ19.The Fab′ fragment was expressed in Ecoli 16C9 solublly and purified by protein G affinity chromatography.The chimeric Fab′ was found to be equipotent to the murine parent Ab in binding CD 3 expressing cells and stimulating T cell prolifcration.