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目的 :探讨补体系统在T细胞活化中的作用。方法 :构建补体组分C3活化片段C3b的受体结合片段C3bAN42 与OVA2 57-2 64CTL表位融合基因表达质粒 ,并对融合蛋白的表达以及生物学活性进行检测。结果 :以重组质粒转染COS 7细胞后 ,用FACS检测融合蛋白在COS 7细胞中得到表达。将转染细胞的培养上清与巨噬细胞系Ana 1共孵育 ,通过激光共聚焦显微镜扫描观察 ,发现融合蛋白可与Ana 1发生特异性结合 ,并且重组的融合蛋白被Ana 1细胞内吞后 ,可释放出表位肽OVA2 57-2 64。后者与MHC Ⅰ类分子结合后 ,以OVA2 57-2 64 MHC Ⅰ类分子复合物的形式被交叉呈递于Ana 1细胞的表面。结论 :C3bAN42 与OVA2 57-2 64CTL表位融合基因在真核细胞中能正确表达 ,且重组的融合蛋白具有其天然的生物学活性
Objective: To investigate the role of complement system in T cell activation. Methods: The fusion fragment of C3bAN42 and OVA2 57-2 64CTL fusion protein was constructed and the expression and biological activity of the fusion protein was detected. Results: After transfection of COS 7 cells with recombinant plasmids, the fusion proteins were expressed in COS 7 cells by FACS. The cultured supernatants of the transfected cells were co-incubated with the macrophage cell line Ana 1. Scanning confocal laser scanning microscopy showed that the fusion protein could specifically bind to Ana 1, and the recombinant fusion protein was endocytosed by Ana 1 cells , Releases epitope peptide OVA2 57-264. The latter is bound to MHC class I molecules and cross-presented on the surface of Ana 1 cells in the form of a complex of OVA2 57-264 MHC class I molecules. Conclusion: The fusion gene of C3bAN42 and OVA2 57-2 64CTL can be correctly expressed in eukaryotic cells, and the recombinant fusion protein has its natural biological activity