title: An optimized method for establishing high purity murine CD8+ T cell cultures.
abstract: 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.
authors.affiliation: T Cell Laboratory, Ludwig Institute for Cancer Research, Austin Health, Melbourne, VIC 3084, Australia.
authors.forename: Damien
authors.initials: D
authors.lastname: Zanker
authors.forename: Kun
authors.initials: K
authors.lastname: Xiao
authors.forename: Sara
authors.initials: S
authors.lastname: Oveissi
authors.forename: Philippe
authors.initials: P
authors.lastname: Guillaume
authors.forename: Immanuel F
authors.initials: IF
authors.lastname: Luescher
authors.forename: Weisan
authors.initials: W
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D007371:Interferon-gamma / Q000276:immunology / Q000378:metabolism; D007376:Interleukin-2 / Q000494:pharmacology; D017493:Leukocyte Common Antigens / Q000276:immunology* / Q000378:metabolism; D018655:Lymphocyte Count; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL
chemical_list: D007376:Interleukin-2; D007371:Interferon-gamma; D017493:Leukocyte Common Antigens