人O型外周血淋巴细胞制备人单克隆抗-A细胞株  

Preparation of human monoclonal anti-A cell lines from PBMCs of O type individual

作　　者：郭忠慧[1] 朱自严[1] 赵俸涌[1] 向东[1] GUO Zhonghui;ZHU Ziyan;ZHAO Fengyong;XIANG Dong(Shanghai Blood Center,Shanghai 200051,China)

机构地区：[1]上海市血液中心,上海200051

出　　处：《中国输血杂志》2020年第11期1155-1157,共3页Chinese Journal of Blood Transfusion

基　　金：上海市血液中心科研基金(01J1802-13);上海市卫生健康委员会科研基金面上项目(202040501)。

摘　　要：目的以正常O型人外周血淋巴细胞为实验对象,制备能稳定分泌人单克隆抗-A或抗-B的杂交瘤细胞株,在本实验室建立人血型单克隆抗体制备流程。方法使用淋巴细胞分离液分离外周血单个核细胞(PBMCs),从狨猴B95-8细胞培养上清收获EB病毒(Epstein-Barr virus),用于转化B淋巴细胞,血清学微量板法筛选能够分泌目标抗体的细胞孔。显微镜下挑选分离单个克隆的细胞团,分泌特异性抗体的转化细胞与SHM-D33瘤细胞进行融合,经HOTD选择性培养,血清学筛选,得到分泌特异性抗体的杂交瘤细胞。结果从随机O型个体的外周血淋巴细胞,成功构建持续稳定分泌人源IgM单克隆抗-A的细胞株。结论本研究提供一种可行的技术路线,为从人外周血淋巴细胞制备有诊断和筛选价值的人源血型单克隆抗体细胞株奠定基础。Objective To prepare hybridoma cell lines, which can stably secret human monoclonal anti-A or anti-B antibody from human peripheral blood lymphocytes(PBMCs) of normal O type donors, and standardize the process for preparing human blood group monoclonal antibody in our laboratory. Methods PBMCs were isolated using lymphocyte isolation solution, and Epstein Barr viruses(EBVs) were harvested from the culture supernatant of B95-8 cells for transformation of B lymphocytes. Cell pores that could secrete the target antibody were screened by serological micro plate method. The cell clusters were selected,The transformed lymphoblastoid cell line(LCL) secreting antibodies to A antigens were collected and then hybridized with the myeloma SHM-D33 using electric fusion technique. The hybridoma cells secreting specific antibody were obtained after selective culture by HOTD and serological screening.Results A hybridoma cell line secreting human IgM monoclonal anti-A antibody was successfully constructed from PBMCs of O type individuals. Conclusion The study provides a viable and useful technical route for the preparation of human monoclonal antibody cell lines with diagnostic and screening value from human PBMCs.

关 键 词：红细胞血型 人单克隆抗体 外周血单个核细胞 杂交瘤技术 

分 类 号：R457.11[医药卫生—治疗学]