抗人大肠癌HCAb嵌合抗体在CHO细胞中的高效表达

目的 在CHO细胞中高效表达抗人大肠癌HCAb人-鼠嵌合抗体.方法 从分泌鼠源单抗CAb的杂交瘤细胞中克隆、扩增可变区基因,测序鉴定后连入真核表达载体,转染二氢叶酸还原酶缺陷型中国仓鼠卵巢（dihydrofolate reductase-deficient Chinese hamster ovary,CHO-dhfr-）细胞进行表达.经G418及氨甲喋呤（MTX）梯度加压筛选进行嵌合抗体的高效表达,并用夹心ELISA方法测定嵌合抗体表达产量.结果 采用RT-pCR、夹心ELISA等实验证实了所表达的抗人大肠癌HCAb嵌合抗体的人源性.培养上清中的抗体产量可达20mg/L.结论 成功的实现了抗人大肠癌HCAb人-鼠嵌合抗体在CHO细胞中高效表达,为其进一步在大肠癌临床诊治中的应用奠定了基础.

High-level expression of human-mouse chimeric antibody HCAb against human colorectal cancer in CHO cells

[Objective] To high-level express human-mouse chimeric antibody against human colorectal cancer. [Methods] Human-mouse chimeric antibody HCAb was expressed by cloning the light and heavy chain variable region genes from the hybridoma which secretes murine antibody. Then we constructed the light and heavy chain expression vectors and transfected CHO-dhfr-cells. The transfected cells were screened with G418 and subjected to gene amplification in medium containing gradually increasing methotrexate (MTX). The expressed chimeric antibody was quantitated by ELISA. [Results] The chimeric antibody was expressed in CHO cells successfully. The results of RT-PCR and ELISA proved that the expression product was humanized. The antibody production achieved 20 mg/L. [Conclusion] Our work lays a good foundation for its further application into diagnosis reagents and immunotherapeutic tracers against human colorectal cancer.