sHLA-A*0201-抗原肽四聚体的构建与功能检测  

The preparation and functional test for HLA-A*0201-peptide tetramer

作　　者：翁秀芳[1] 吴雄文[1] 梁智辉[1] 蔡蕾[1] 费世江[1] 陈国安[1] 龚非力[1] 

机构地区：[1]华中科技大学同济医学院免疫学系,武汉430030

出　　处：《中国免疫学杂志》2006年第3期244-247,251,共5页Chinese Journal of Immunology

基　　金：国家自然科学基金(30271201);科技部973计划(2001CB510008)资助

摘　　要：目的:构建有功能的sHLA-A*0201-抗原肽四聚体,建立一套HLAⅠ类分子/抗原肽的四聚体制备技术。方法:利用基因工程及体外折叠技术构建sHLA-A*0201-LMP2A426-434复合物分子,并在BirA酶的作用下生物素化,与荧光标记的亲和素衍生物以分子数4∶1结合而形成HLA-A*0201-LMP2A426-434四聚体。获得的四聚体对长期混合淋巴细胞培养中增殖的抗原特异性CTL进行检测,同时用细胞毒实验验证。结果:荧光标记的亲和素与生物素化的HLA-A*0201-抗原肽复合物分子正确结合,制备的四聚体能够与长期混合淋巴细胞培养出的特异性T细胞结合。结论:从结构与功能上证实成功地获得有功能的HLA-A*0201-抗原肽复合物四聚体。为进一步研究T细胞识别机制与功能奠定了基础,也为过程复杂的HLA-抗原肽四聚体制备提供了可行的免疫学监控方法。Objective：To prepare a sHLA-A2/peptide tetramer and set up the protocols which can be used for preparation of other types of sHLA class Ⅰ molecules with the corresponding peptide. Methods： sHLA-A2/LMP2A426-434 tetramer was prepared by mixing the biotinylated peptide-HLA complex with phycoerythrin（PE） labeled avidin at a molar ratio of 4： 1. We apply our prepared sHLA-A2/LMP2A426-434 tetramer to stain the cells after long term mixed lymphocytes culture. Results：The results indicated sHLA-A2/ LMP2A426-434, tetramer has been correctly constructed in structure and the tetramer was able to discriminate the frequencies of specific CTL induced by HLA-A2/LMP2A426-434. Conclusion：We have successfully constructed the HLA-A＊0201/LMP2A426-434, tetramer and lay the foundation for further study of T cell recognition.

关 键 词：HLA-A＊0201-抗原肽复合物 生物素化 BirA HLA-A＊0201-抗原肽四聚体 抗原特异性CTL 

分 类 号：R392[医药卫生—免疫学]