【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。更多还原

【Abstract】 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/LMP2A_ 426-434 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/LMP2A_ 426-434 tetramer to stain the cells after long term mixed lymphocytes culture.Results:The results indicated sHLA-A2/LMP2A_ 426-434 tetramer has been correctly constructed in structure and the tetramer was able to discriminate the frequencies of specific CTL induced by HLA-A2/LMP2A_ 426-434 .Conclusion:We have successfully constructed the HLA-A*0201/LMP2A_ 426-434 tetramer and lay the foundation for further study of T cell recognition.更多还原