Combined megaplex TCR isolation and SMART-based real-time quantitation methods for quantitating antigen-specific T cell clones in mycobacterial infection.

ABSTRACT: Despite recent advances in measuring cellular immune responses, the quantitation of antigen-specific T cell clones in infections or diseases remains challenging. Here, we employed combined megaplex TCR isolation and SMART-based real-time quantitation methods to quantitate numerous antigen-specific T cell clones using limited amounts of specimens. The megaplex TCR isolation covered the repertoire comprised of recombinants from 24 Vbeta families and 13 Jbeta segments, and allowed us to isolate TCR VDJ clonotypic sequences from one or many PPD-specific IFNgamma-producing T cells that were purified by flow cytometry sorting. The SMART amplification technique was then validated for its capacity to proportionally enrich cellular TCR mRNA/cDNA for real-time quantitation of large numbers of T cell

SUBMITTER: Du G 

PROVIDER: S-EPMC2884368 | biostudies-literature | 2006 Jan

REPOSITORIES: biostudies-literature