Raji 细胞系特异性Fab噬菌体抗体库构建及筛选.docVIP

Raji 细胞系特异性Fab噬菌体抗体库构建及筛选 　 ; 作者：谢小芳, 申咏梅, 杨晓春, 董宁征, 白霞, 石怡珍 毕业论文 【关键词】; B细胞淋巴瘤;,Fab;,噬菌体抗体库, 毕业论文 　　Construction and screening of the specific Fab phage antibody library against Raji cell strain 毕业论文 　　[Abstract]; AIM: To construct and screen the specific Fab phage antibody library against human Raji cell strain in Blymphoma. METHODS: BALB/c mice were immunized with Raji cells, and the antibody light chain к genes and heavy chain genes Fd from the spleen cells were amplified by RTPCR. After restrictive digestion with Sac I/Xba I and Xho I/Spe I, the light chain кgenes and heavy chain genes Fd were inserted into the phagemid vector pComb3HSS successively and then electroporated into E.coli XL1Blue. The specific Fab phage antibody library against Raji cell strain in human Blymphoma was constructed by infection of helper phage VCSM13. The specific antibodies against Raji cells were obtained after selected with Raji cells. The binding activity with antigens was identified by ELISA and the positive clones were sequenced. RESULTS: The Fab phage antibody library with 2.18×107 volume was constructed and eight positive clones which specifically recognized Raji cell strain were isolated. Sequence analysis of the two positive clones showed that the variable heavy domains (VH) and variable light domains (VL) were highly homologous with the registered murine Ig heavy chain V region sequences and kappa light chain sequences, respectively. CONCLUSION: Fab phage antibody library was successfully constructed and specific antibodies against membrane antigens in Raji cells were obtained, which will provide an experimental foundation for the further investigation of Blymphoma immunotherapy. 毕业论文 　　[Keywords]Blymphoma; Fab; phage antibody library 毕业论文 　　[摘 要]; 目的: 构建针对B细胞淋巴瘤Raji细胞系噬菌体抗体库并从中筛选出特异性的抗体。方法: 以Raji细胞免疫BALB/c小鼠, RTPCR法从脾淋巴细胞中扩增抗体轻链к和重链Fd基因。经Sac I/Xba I和Xho I/Spe I双酶切, 依次克隆入噬菌体载体pComb3HSS中, 并电转化大肠杆菌XL1Blue, 以辅助噬菌体VCSM13进行超感染, 构建B细胞淋巴瘤Raji细胞系特异性Fab噬菌体抗体库。以Raji细胞为