Sensitive and viable identification of antigen-specific CD8+T cells by a flow cytometric assay for degranulation
Sensitive and viable identification of antigen-specific CD8+T cells by a flow cytometric assay for degranulation
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DOI：
10.1016/s0022-1759(03)00265-5
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发表时间：
2003-10-01
影响因子：
2.2
通讯作者：
Koup, RA
中科院分区：
文献类型：
--
作者：
Betts, MR;Brenchley, JM;Koup, RA
抗原特异性CD8+ T细胞的流式细胞术检测以前仅限于mhc - I类四聚体染色或细胞内细胞因子产生，这两种方法都不能测量这些细胞的细胞溶解潜能。在这里，我们提出了一种新的技术来枚举抗原特异性CD8+ T细胞，使用在细胞表面表达的标记，在活化诱导的脱颗粒后，细胞溶解的必要前体。本试验测量CD107a和b的暴露，存在于细胞毒性颗粒的膜，到细胞表面的脱粒结果。细胞表面CD107a和In的获取与细胞内功能的丧失有关，并被秋水仙碱抑制，这表明CD107a和b暴露于细胞表面依赖于脱颗粒。CD107a和b被同源肽激活后在CD8+ T细胞表面表达，与细胞内IFNgamma的产生一致。最后，表达cd107的CD8+ T细胞以抗原特异性的方式介导细胞溶解活性。CD107a和b表达的测量也可以与mhc - I类四聚体标记和细胞内细胞因子染色相结合，以更完整地评估表达同源T细胞受体（TCR）的CD8+ T细胞的功能。(C) 2003 Elsevier B.V.版权所有
Flow cytometric detection of antigen-specific CD8+ T cells has previously been limited to MHC-class I tetramer staining or intracellular cytokine production, neither of which measure the cytolytic potential of these cells. Here we present a novel technique to enumerate antigen-specific CD8+ T cells using a marker expressed on the cell surface following activation induced degranulation, a necessary precursor of cytolysis. This assay measures the exposure of CD107a and b, present in the membrane of cytotoxic granules, onto the cell surface as a result of degranulation. Acquisition of cell surface CD107a and In is associated with loss of intracellular perform and is inhibited by colchicine, indicating that exposure of CD107a and b to the cell surface is dependent on degranulation. CD107a and b are expressed on the cell surface of CD8+ T cells following activation with cognate peptide, concordant with production of intracellular IFNgamma. Finally, CD107-expressing CD8+ T cells are shown to mediate cytolytic activity in an antigen-specific manner. Measurement of CD107a and b expression can also be combined with MHC-class I tetramer labeling and intracellular cytokine staining to provide a more complete assessment of the functionality of CD8+ T cells expressing cognate T cell receptors (TCR). (C) 2003 Elsevier B.V. All rights reserved.