Reference - Detail
| RRC ID | 38208 |
|---|---|
| Author | Arakaki A, Ooya K, Akiyama Y, Hosokawa M, Komiyama M, Iizuka A, Yamaguchi K, Matsunaga T. |
| Title | TCR-beta repertoire analysis of antigen-specific single T cells using a high-density microcavity array. |
| Journal | Biotechnol Bioeng |
| 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-beta 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-beta 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. |
| Volume | 106(2) |
| Pages | 311-8 |
| Published | 2010-6-1 |
| DOI | 10.1002/bit.22663 |
| PMID | 20073087 |
| MeSH | Cell Separation / instrumentation* Equipment Design Equipment Failure Analysis Oligonucleotide Array Sequence Analysis / instrumentation* Receptors, Antigen, T-Cell / genetics* Reverse Transcriptase Polymerase Chain Reaction / instrumentation* |
| IF | 4.002 |
| Times Cited | 10 |
| WOS Category | BIOTECHNOLOGY & APPLIED MICROBIOLOGY |
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| Total number of mentions | 0 |
| Resource | |
| Human and Animal Cells | |