论文部分内容阅读
目的构建并表达β2微球蛋白-绿色荧光蛋白融合蛋白,并应用该蛋白建立一种全新的MHC-Ⅰ肽亲和力实验方法。方法构建β2M-ZsGreen-6×His标签的融合蛋白真核表达质粒,转染293FT细胞表达,并以镍亲和层析法纯化,将该融合蛋白与表位肽共同与T2细胞反应,通过流式细胞术检测T2细胞标记绿色荧光的情况鉴定表位。结果构建了β2微球蛋白-绿色荧光蛋白真核表达质粒并表达、纯化得到融合蛋白,该蛋白可应用于亲和力实验,进行表位鉴定。结论β2微球蛋白-绿色荧光融合蛋白应用于亲和力实验鉴定CTL表位方法可行,步骤简便,适用于大通量表位鉴定。
Objective To construct and express β2-microglobulin-green fluorescent protein fusion protein and to establish a novel MHC-Ⅰ peptide affinity test using this protein. Methods The eukaryotic expression plasmid of β2M-ZsGreen-6 × His tag fusion protein was constructed and transfected into 293FT cells. The recombinant protein was purified by nickel affinity chromatography. The fusion protein and epitope peptide were reacted with T2 cells. Cytometric detection of T2 cells labeled with green fluorescence identifies epitopes. Results The eukaryotic expression plasmid of β2-microglobulin-GFP was constructed and expressed. The fusion protein was purified and confirmed by affinity test. Conclusion The β2-microglobulin-green fluorescent fusion protein can be used in affinity experiments to identify CTL epitopes. The method is simple and convenient and is suitable for the identification of large-flux epitopes.