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Author:
Date:
2016-06-20
[Abstract] Cytotoxic CD8+ T cells are responsible for the lysis of cells expressing peptides associated with MHC class I molecules and derived from infection with a pathogen or from mutated antigens. In order to quantify in vivo this antigen-specific CD8+ T cell killing activity, we use the in vivo killing assay (IVK). Here we describe the protocol for the lysis of cells expressing a CD8+ T cell epitope of the OVA protein (SIINFEKL). Mice are previously immunized with the OVA protein and 7 days after immunization, they receive a mix of target cells, prepared from naive C57BL/6 spleen cells pulsed with the SIINFEKL peptide and labeled with high level of CFSE and of non-pulsed control cells labeled with low level of CFSE. One day later, the spleen ...
[摘要] 细胞毒性CD8 + T细胞负责裂解表达与MHC I类分子相关并且源自病原体感染或突变抗原的肽的细胞。为了在体内定量该抗原特异性CD8 + T细胞杀伤活性，我们使用体内杀伤试验（IVK）。在这里，我们描述了用于裂解表达OVA蛋白（SIINFEKL）的CD8 + T细胞表位的细胞的方案。小鼠先前用OVA蛋白免疫，并且在免疫后7天，它们接受从用SIINFEKL肽脉冲并用高水平的CFSE标记的天然C57BL/6脾细胞制备的靶细胞和标记的非脉冲对照细胞的混合物具有低水平的CFSE。一天后，分离受体小鼠的脾细胞并通过FACS分析以测量CFSE高度细胞和CFSE低度细胞的量。通过在免疫的和未免疫的小鼠中CFSE高与低之间的差计算裂解的百分比。
测量抗原特异性CD8 + T细胞在体内裂解其抗原的能力对评价诱导针对病原体或肿瘤抗原的适应性细胞毒性应答是非常重要的。
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