Use of cell-tracking dyes to determine proliferation precursor frequencies of antigen-specific T cells

Methods Mol Biol. 2004;263:109-24. doi: 10.1385/1-59259-773-4:109.

Abstract

The T-cell receptor provides T cells with specificity for antigens of particular molecular structure (the "epitope"); the T-cell pool in an individual responds to the presence of many different antigenic epitopes, but any particular T cell will respond preferentially to one defined epitope. After stimulation of a T cell by the binding of its receptor to its cognate antigen in the context of a major histocompatibility complex (MHC) molecule on an antigen-presenting cell, the T cell will begin to proliferate and synthesize cytokines. Tetramer binding and the enzyme-linked immunospot (ELISPOT) method have been used to determine what proportion of cells in the T-cell pool can bind to a defined antigenic peptide or will secrete cytokines in response to a particular antigenic stimulation. The method described here uses tracking dyes to determine what proportion of T cells will proliferate in response to stimulation. As a flow cytometric "single-cell" method, it can be combined with tetramer and cytokine staining to determine the precursor frequencies of cells in the T-cell pool able to bind tetramer, to synthesize cytokines, and to proliferate in response to antigen.

MeSH terms

  • Animals
  • Antigen-Presenting Cells / metabolism
  • Antigens / chemistry
  • CD8 Antigens / chemistry
  • Cell Division
  • Coloring Agents / pharmacology*
  • Cytokines / metabolism
  • Enzyme-Linked Immunosorbent Assay
  • Epitopes / chemistry
  • Flow Cytometry
  • Fluoresceins / chemistry
  • Fluorescent Dyes / pharmacology
  • Humans
  • Interferon-gamma / metabolism
  • Models, Statistical
  • Peptides / chemistry
  • Phenotype
  • Receptors, Antigen, T-Cell / chemistry
  • Succinimides / chemistry
  • T-Lymphocytes / chemistry*

Substances

  • 5-(6)-carboxyfluorescein diacetate succinimidyl ester
  • Antigens
  • CD8 Antigens
  • Coloring Agents
  • Cytokines
  • Epitopes
  • Fluoresceins
  • Fluorescent Dyes
  • Peptides
  • Receptors, Antigen, T-Cell
  • Succinimides
  • Interferon-gamma