小鼠HSP27/HSPB1单克隆抗体的制备和鉴定  

Preparation and Identification of Mouse Monoclonal Antibody against HSP27/HSPB1

作　　者：田方艳 曹辉 张培湘 黄欢 尹秀峰 王杨 周辉[2] 

机构地区：[1]湖南远泰生物技术有限公司,湖南长沙410219 [2]中南大学湘雅医学院附属肿瘤医院,湖南长沙410013

出　　处：《肿瘤药学》2013年第1期26-29,43,共5页Anti-Tumor Pharmacy

基　　金：长沙市科技局科技计划重点项目资助(编号:K1112022);湖南省医药卫生科研计划项目资助(B2012-098);湖南省科技厅科技计划项目资助(2012SK3249)

摘　　要：目的制备针对热休克蛋白27/热休克蛋白B1(HSP27/HSPB1)的特异性单克隆抗体并对其进行鉴定。方法用HSP27/HSPB1作为免疫抗原免疫BalB/c小鼠,通过常规杂交瘤技术将免疫后的小鼠脾细胞与SP2/0细胞经PEG融合,筛选阳性杂交瘤细胞株,进行克隆化培养后接种于小鼠腹腔制备HSP27/HSPB1单克隆抗体,并利用间接酶联免疫吸附试验、免疫印迹、免疫组织化学、免疫荧光技术对所制备的抗体进行鉴定。结果获得了一株可稳定分泌HSP27/HSPB1蛋白单克隆抗体的杂交瘤细胞株,命名为4C9C7,其亚类是IgG1,分泌的单克隆抗体能特异性结合组织和细胞中的HSP27/HSPB1蛋白。结论本实验成功制备了可稳定分泌HSP27/HSPB1单克隆抗体的杂交瘤细胞及HSP27/HSPB1特异性的单克隆抗体,可应用于各种相关的科学研究。Objective To prepare and indentify specific monoclonal antibody （McAb） against heat shock protein 27/heat shock protein B1 （HSP27/HSPB1）. Methods BalB/c mice were immunized with purified HSP27/HSPB1 protein expressed by prokaryotic expression system. Then the spleen cells from immunized mice were fused with SP2/0 myeloma cells by conventional hybridoma technique. The positive hybridoma cells were chosen to clone and culture, and then were injected into the abdominal cavities of BalB/c mice. The prepared antibody was identified by indirect enzyme linked immunosorbent assay （ELISA）, western blotting, immunohistochemistry and immunofluorescence technique. Results A hybridoma cell line that was able to stably secrete HSP27/HSPB1 monoclonal antibody was obtained and named 4C9C7. Its subtype was IgG1. Its secreted antibodies could spe-cifically bind to HSP27/HSPB1 protein from different types of tissues and cells. Conclusion A hybridoma cell line that was capa-ble to stably secret HSP27/HSPB1 monoclonal antibody was successfully prepared in our study and it may be applied to various related scientific researches.

关 键 词：热休克蛋白27 热休克蛋白B1 单克隆抗体 制备 鉴定 

分 类 号：Q511[生物学—生物化学]