小鼠抗人CD52功能性抗体的制备与鉴定  

Preparation and identification of mouse anti-human CD52 functional antibody

作　　者：刘友[1,2] 屈浩 石琳 黄丽华 张宇光[1,3] 朱卫彬 

机构地区：[1]北京协和医学院中国医学科学院血液学研究所血液病医院,天津300020 [2]内蒙古科技大学包头医学院,内蒙古包头014040 [3]协和干细胞基因工程有限公司,天津300384

出　　处：《细胞与分子免疫学杂志》2013年第1期56-59,64,共5页Chinese Journal of Cellular and Molecular Immunology

基　　金：天津市科技创新专项资金项目(08FDZDSH03000)

摘　　要：目的鼠抗人CD52抗血清的制备与功能鉴定。方法由人Hut-78细胞提取细胞总RNA,运用RT-PCR方法扩增CD52基因,定向克隆入真核表达载体pcDNA3.1(+),用脂质体转染法转染CHO细胞,建立稳定转染的CHO细胞系,用RT-PCR、FACS、免疫组织化学技术检测目的蛋白的表达。用CD52合成肽免疫小鼠,通过间接ELISA和流式细胞术分析抗血清,用MTS实验鉴定抗体对白血病LCL细胞系的增殖抑制作用。结果建立了稳定转染的CHO细胞系。鉴定出有高效价抗体存在。MTS实验检测免疫血清有抑制LCL细胞生长作用。结论功能性抗体制备成功并建立了稳定转染CD52的CHO细胞系。Objective To prepare and identify mouse anti-human CD52 antibody. Methods The RNA was extracted from Hut-78 cells. CD52 gene was amplified by RT-PCR. With the double-enzyme digestion, CD52 gene was cloned into pcDNA3. 1 （＋） eukaryotic expression vector, named as pcDNA3. I （＋）/CD52. We transfected the recombinant into CHO cell by LipofectamineTM 2000. Stable transfected CHO cell line was established, and the CD52 expression in the transfected cells was detected by RT-PCR, FACS and immunocytochemical staining. BALB/c mice were immunized with synthetic peptide CD52. The titer of anti-CD52 serum was detected with indirect ELISA. Inhibition of leukemia LCL cell proliferation by immu- nized mouse serum was identified with MTS assay. Results The eukaryotic expression vector pcDNA3.1 （＋）/CD52 was constructed, stable transfected CHO cell line was established. High titer anti-CD52 antibody was identified. The immunized mouse serum inhibits proliferation of LCL cells by MTS assay. Conclusion Preparation of functional antibody against CD52 and stable transfected CHO cell line have established successfully.

关 键 词：CD52抗体 真核表达载体 稳定转染细胞 抗体制备 

分 类 号：R392.11[医药卫生—免疫学]