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目的:构建检测抗原特异性细胞毒性T淋巴细胞的单链MHC-肽四聚体。方法:通过在MHC重链基因中引入生物素化位点,并在轻链和重链基因中引入同一限制性酶切位点连接MHC轻链和重链构建可生物素化的单链MHC分子。结果:重组子以包涵体形式表达得到单链MHC分子,重折叠、生物素化、荧光标记后得到活性单链MHC-肽四聚体。结论:单链MHC分子操作简便,提高了四聚体构建效率。
Objective: To construct a single chain MHC-peptide tetramer for detection of antigen-specific cytotoxic T lymphocytes. Methods: Biotinylated single-chain MHC molecules were constructed by introducing a biotinylation site in the MHC heavy chain gene and introducing the same restriction sites in the light and heavy chain genes to link the MHC light and heavy chains . Results: The recombinant protein was expressed in inclusion bodies to obtain single-chain MHC molecules. After refolding, biotinylation and fluorescent labeling, the active single-chain MHC-peptide tetramers were obtained. Conclusion: Single-chain MHC molecules are easy to operate and improve the efficiency of tetramer construction.