摘要
目的 :构建并在真核细胞中表达抗人重组碱性成纤维生长因子 (rh bFGF)人 鼠嵌合抗体。方法 :从分泌抗rh bFGF鼠单抗(mAb) 1F11的杂交瘤细胞中扩增、克隆VL、VH 基因 ,从质粒pMDHC和pMDLC中扩增、克隆CL、CH 基因 ,测序鉴定后插入真核表达载体 ,并转化二氢叶酸还原酶缺陷型中国仓鼠卵巢(CHO dhfr- )细胞进行表达。采用间接ELISA检测表达产物的人源性和其抗原结合活性。结果 :序列测定表明所克隆的抗体基因是功能性抗体的V区基因 ,在转化细胞的培养上清中可检测到抗rh bFGF嵌合抗体的表达。ELISA试验证实 ,该嵌合抗体具有人源C区 ,并具有鼠mAb 1F11的抗原结合活性。结论 :在真核细胞中成功地表达抗rh bFGF的人 鼠嵌合抗体 。
AIM: To express a human-mouse chimeric antibody against rh-bFGF antige n in eukaryotic cells. METHODS: The V L and V H genes were amp lified and cloned from the hybridoma 1F11 secreting anti-rh-bFGF mouse monoclo nal antibody (mAb) and the C L and C H genes were amplified and cloned from th e plasmid pMDHC and pMDLC. They were inserted into eukaryotic expression vectors after sequencing. The recombinant plasmids were then transformed into CHO-dhfr - cells for expression. The humanization and antigen specificity of expressed products were identified by indirect ELISA. The relative molecular mass (M r) of expressed products was determined by SDS-PAGE. RESULTS: The cloned antibody genes were identified to be functional by sequencing. The chimeric antibody could be detected in the culture supernatant of transformed CH O cells.The humanization and specificity of the chimeric antibody to rh-bFGF we re confirmed by ELISA. CONCLUSION: The mouse-human chimeric ant ibody against rh-bFGF has been expressed successfully in eukaryotic cells, whic h lays the foundation for its further clinical research.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2004年第2期163-167,共5页
Chinese Journal of Cellular and Molecular Immunology
基金
广州市科技局重点课题资助 (No .2 0 0 3J1 C0 1 71 2 )
国家计委部门重点项目资助 (No .1 997 2 0 71 )
