An optimized method for establishing high purity murine CD8+ T cell cultures

J Immunol Methods. 2013 Jan 31;387(1-2):173-80. doi: 10.1016/j.jim.2012.10.012. Epub 2012 Oct 22.


Establishing CD8(+) T cell cultures has been empirical and the published methods have been largely individual laboratory based. In this study, we optimized culturing conditions and show that IL-2 concentration is the most critical factor for the success of establishing CD8(+) T cell cultures. High IL-2 concentration encouraged T cells to non-specifically proliferate, express a B cell marker, B220, and undergo apoptosis. These cells also lose typical irregular T cell morphology and are incapable of sustaining long-term cultures. Using tetramer and intracellular cytokine assessments, we further demonstrated that many antigen-specific T cells have been rendered nonfunctional when expanded under high IL-2 concentration. When IL-2 is used in the correct range, B220-mediated cell depletion greatly enhanced the success rate of such T cell cultures.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Apoptosis / drug effects
  • Apoptosis / immunology*
  • CD8-Positive T-Lymphocytes / drug effects
  • CD8-Positive T-Lymphocytes / immunology*
  • CD8-Positive T-Lymphocytes / metabolism
  • Cell Culture Techniques / methods*
  • Cell Proliferation / drug effects
  • Cells, Cultured
  • Dose-Response Relationship, Drug
  • Interferon-gamma / immunology
  • Interferon-gamma / metabolism
  • Interleukin-2 / pharmacology
  • Leukocyte Common Antigens / immunology*
  • Leukocyte Common Antigens / metabolism
  • Lymphocyte Count
  • Mice
  • Mice, Inbred BALB C
  • Mice, Inbred C57BL


  • Interleukin-2
  • Interferon-gamma
  • Leukocyte Common Antigens