Single-Cell Analysis of T-Cell Receptor αβ Repertoire
Single-Cell Analysis of T-Cell Receptor αβ Repertoire
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DOI：
10.1007/978-1-4939-2963-4_15
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发表时间：
2015-01-01
期刊：
影响因子：
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通讯作者：
Thomas, Paul G.
中科院分区：
文献类型：
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作者：
Dash, Pradyot;Wang, George C.;Thomas, Paul G.
在单细胞水平上对T细胞受体(TCR)α链和β链使用的无偏见的配对分析为了解TCR谱系和免疫反应的性质提供了一个有价值的窗口。早期的TCR谱系分析技术往往局限于检测TCR互补决定区3(CDR3)β的表达，或者需要体外克隆程序，这可能会人为地使TCR谱系从体内状态偏离。在这里，我们描述了一种直接的体外、基于单细胞的策略，用于TCRαβ谱系的克隆型分析，该策略利用嵌套聚合酶链式反应中TCRα和TCRβ特异性引物的多路面板来扩增单个表位特异性T细胞的表达转录。这一策略产生了感兴趣的任何给定群体的α-βT细胞的配对TCRα-β序列。
The unbiased, paired analysis of T-cell receptor (TCR) alpha- and beta-chain usage at the single-cell level provides a valuable window of understanding into the TCR repertoire and the nature of the immune response. Earlier technologies for TCR repertoire analysis were often limited to examining TCR complementarity-determining region 3 (CDR3) beta expression or required in vitro cloning procedures that can artificially skew the TCR repertoire from its in vivo state. We describe here a direct ex vivo, single-cell-based strategy for the clonotypic analysis of TCR alpha beta repertoires that utilizes multiplexed panels of TCR alpha and TCR beta-specific primers in a nested PCR to amplify expressed transcripts from individual, epitope-specific T cells. This strategy yields the paired TCR alpha beta sequences of any given population of alpha beta T cells of interest.