Simultaneous assessment of antigen-stimulated cytokine production and memory subset composition of memory CD8 T cells

J Immunol Methods. 2006 Jun 30;313(1-2):161-8. doi: 10.1016/j.jim.2006.04.005. Epub 2006 May 26.

Abstract

The functional identification of antigen-specific CD8 T cell populations is critical to understanding host responses to infection by intracellular pathogens. Furthermore, assessing the properties of protective memory CD8 T cell populations generated by immunization is necessary in the rational design of vaccines. Recently, a classification scheme was proposed in which memory CD8 T cells were divided into one of two distinct subsets, based on CD62L expression, that have different functional properties and protective capacities. Intracellular cytokine staining functionally identifies antigen-specific CD8 T cell populations after short in vitro stimulation with cognate peptide. This short stimulation, however, results in the cleavage of CD62L from the cell surface of antigen-specific CD8 T cells and precludes distinguishing CD62L(hi)- and CD62L(lo)-expressing memory cell subsets within this population. Here, we describe a method of preserving CD62L expression by the antigen-specific CD8 T cell population during coculture with antigen. This methodology allows for the identification and functional assessment of antigen-specific memory CD8 T cell populations, while simultaneously characterizing the memory subset composition of that population. Using this method, we directly identify differences in IL-2 production capacity by CD62L(hi)- and CD62L(lo)-expressing antigen-specific memory CD8 T cell populations.

MeSH terms

  • Adoptive Transfer
  • Animals
  • Arenaviridae Infections / immunology
  • CD8-Positive T-Lymphocytes / immunology*
  • CD8-Positive T-Lymphocytes / metabolism
  • CD8-Positive T-Lymphocytes / transplantation
  • Cytokines / biosynthesis*
  • Flow Cytometry
  • Hydroxamic Acids / pharmacology
  • Immunologic Memory / immunology*
  • Interferon-gamma / biosynthesis
  • Interleukin-2 / biosynthesis
  • L-Selectin / analysis
  • Listeria monocytogenes / immunology
  • Listeriosis / immunology
  • Lymphocyte Activation / immunology
  • Lymphocytic choriomeningitis virus / immunology
  • Mice
  • Mice, Inbred BALB C
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Oligopeptides / pharmacology
  • Ovalbumin / immunology
  • Receptors, Antigen, T-Cell / genetics
  • Spleen / cytology
  • Spleen / drug effects
  • Spleen / immunology
  • T-Lymphocyte Subsets / drug effects
  • T-Lymphocyte Subsets / immunology*
  • T-Lymphocyte Subsets / metabolism

Substances

  • Cytokines
  • Hydroxamic Acids
  • Interleukin-2
  • Oligopeptides
  • Receptors, Antigen, T-Cell
  • TAPI-2
  • L-Selectin
  • Interferon-gamma
  • Ovalbumin