一种改进的分泌抗半抗原抗体杂交瘤细胞株筛选方法  

An Improved Method for Screening Hybridoma Cells Secreting AntiHapten Antibodies

作　　者：王云龙 段江波[2] 李恒思 王霈[4] 毛烈[2] 张怡青[5] 王国强 王继创 

机构地区：[1]郑州市职业技术学院,河南郑州450046 [2]郑州大学生物工程系,河南郑州450001 [3]河南省生物工程技术研究中心,河南郑州450001 [4]中国药科大学,江苏南京211198 [5]河南师范大学,河南新乡453002

出　　处：《生物技术通讯》2014年第1期94-96,共3页Letters in Biotechnology

基　　金：河南省科技攻关项目(122102310633)

摘　　要：目的:研究解决半抗原分子单克隆抗体制备技术路径中遇到的在阳性杂交瘤细胞株筛选时无法排除载体蛋白间交叉反应影响的问题,以半抗原去甲肾上腺素(norepinephrine,NE)为例。方法:在NE完全抗原免疫小鼠实施细胞融合后,分别包被牛血清白蛋白(BSA)、卵清白蛋白(OVA)、BSA-NE、OVA-NE等4种不同抗原的酶标板平行检测细胞培养上清液;挑选BSA、OVA检测结果为阴性,BSA-NE、OVA-NE检测结果为阳性的孔内细胞进行克隆化筛选单克隆细胞。结果:本筛选方法可一次性从8板96孔板中筛选到13个符合要求的阳性孔,经3次克隆化后获得6株特异性强的杂交瘤细胞株。结论:本方法避免了载体蛋白间交叉反应对筛选的影响,改进了传统的单一指标筛选方法,筛选效率更高。Objective： Take norepinephrine（NE） as an example to establish an improved hybridoma screen mode which increasing specificity and efficiency of screening procedure when preparing for anti-hapten antibodies. Meth ods： After cell fusion steps, four kinds of antigens, bovine serum alhumin（BSA）, ovalbumin（OVA）, BSA-NE, OVA-NE, were coated separately on 96 wells ELISA plate and simultaneously test the supernatant of hybridoma culture. Those hyhridomas wells with results of BSA-NE and OVA-NE positive reaction, but BSA and OVA nega tive reaction were selected for a further cloning procedure to get monoclonal cell lines. Results： 13 satisfied mono clonal hybridoma cells were obtained from 8 ELISA plates. After the cloning procedure 6 high specific monoclonal antibody secreting hybridomas cell lines acquired. Conclusion： This improved ELISA screening mode directly en- hanced the specificity towards hapten structure compared with currently commonly used method. This mode can suf ficiently avoid interaction between protein carriers.

关 键 词：半抗原 杂交瘤细胞筛选 去甲肾上腺素 半抗原载体蛋白 

分 类 号：R331[医药卫生—人体生理学]