hFcεRIα/RBL-2H3细胞株的构建与评价  

Construction and identification of recombinant RBL-2H3 cells transfected with hFcεRIα

作　　者：王南南[1] 刘中成[1] 张艳芬[2] 刘玉欣[1] 崔哲[1] 陈瑶[1] 

机构地区：[1]河北大学药学院,河北保定071002 [2]河北大学科学技术处,河北保定071002

出　　处：《中国药理学通报》2015年第8期1174-1179,共6页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 81202338);河北省自然科学基金资助项目(No H2013201128);中国博士后科学基金资助项目(No 2013M530885);河北省教育厅项目(No Z2015013)

摘　　要：目的构建稳定表达人FcεRIα(h FcεRIα)亚基的RBL-2H3细胞株。方法利用RT-PCR技术克隆h FcεRIα基因,构建真核表达载体p CI-neo-h FcεRIα。脂质体介导法转染RBL-2H3细胞,G418筛选获得稳定转染细胞株。利用RT-PCR、Western blot及免疫荧光法鉴定转染结果。结果通过对脂质体介导转染体系进行优化,转染效率可高达75.38%。经Western blot、免疫荧光及RT-PCR鉴定,h FcεRIα基因在RBL-2H3细胞内成功表达。结论成功构建h FcεRIα/RBL-2H3细胞模型,为深入研究Ig E与FcεRI作用机制及相关药物开发提供了实验基础。Aim To construct the stable hFcεRIα/RBL-2H3 cell line expressing human FcεRIα（hFcaRIct）. Methods The human FcεRIα gene was obtained by RT-PCR and cloned into the eukaryotic expression vector pCI-neo. Then, the pCI-neo- hFcεRIα vector was transfected into RBL-2H3 cells by lipo- somes, and the transfected cells were screened through G418 fil- tration subsequently. Finally, RT-PCR, Western blot and immu- nofiuoreseence assay were used to determine the result of trans- fection. Results According to the optimized transfection param-eters, the transfection efficiency reached 75.38%. The results of Western blot, immunofluoreseence and RT-PCR showed that hFcεRIα could be expressed in RBL-2H3 cells successfully. Conclusion HFeεRIα/RBL-2H3 cells were successfully con- structed, which will be the experimental basis for further study on the mechanism of IgE/FcεRI and drugs for allergy diseases.

关 键 词：hFcεRIα IGE RBL-2H3细胞 脂质体转染 G418筛选 稳定细胞株 

分 类 号：R329.24[医药卫生—人体解剖和组织胚胎学] R392.11[医药卫生—基础医学]