TCR-β repertoire analysis of antigen-specific single T cells using a high-density microcavity array

Atsushi Arakaki, Kaori Ooya, Yasuto Akiyama, Masahito Hosokawa, Masaru Komiyama, Akira Iizuka, Ken Yamaguchi, Tadashi Matsunaga

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

The antigen specificity of cytotoxic T cells, provided by T-cell receptors (TCRs), plays a central role in human autoimmune diseases, infection, and cancer. As the TCR repertoire is unique in individual cytotoxic T cells, a strategy to analyze its gene rearrangement at the single-cell level is required. In this study, we applied a high-density microcavity array enabling target cell screening of several thousands of single cells for identification of functional TCR-β gene repertoires specific to melanoma (gp100) and cytomegalovirus (CMV) antigens. T cells expressing TCRs with the ability to recognize fluorescent-labeled antigen peptide tetramers were isolated by using a micromanipulator under microscopy. Regularly arranged cells on the microcavity array eased detection and isolation of target single cells from a polyclonal T-cell population. The isolated single cells were then directly utilized for RT-PCR. By sequencing the amplified PCR products, antigen-specific TCR-β repertoires for gp100 and human cytomegalovirus antigens were successfully identified at the single-cell level. This simple, accurate, and cost-effective technique for single-cell analysis has further potential as a valuable and widely applicable tool for studies on gene screening and expression analyses of various kinds of cells.

Original languageEnglish
Pages (from-to)311-318
Number of pages8
JournalBiotechnology and Bioengineering
Volume106
Issue number2
DOIs
Publication statusPublished - 2010 Jun 1
Externally publishedYes

Keywords

  • Gene expression analysis
  • Microarray analytical devices
  • Single-cell
  • T-cell antigen receptor specificity
  • T-cell receptor beta genes

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

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  • Cite this

    Arakaki, A., Ooya, K., Akiyama, Y., Hosokawa, M., Komiyama, M., Iizuka, A., Yamaguchi, K., & Matsunaga, T. (2010). TCR-β repertoire analysis of antigen-specific single T cells using a high-density microcavity array. Biotechnology and Bioengineering, 106(2), 311-318. https://doi.org/10.1002/bit.22663