title: Optimization and validation of a robust human T-cell culture method for monitoring phenotypic and polyfunctional antigen-specific CD4 and CD8 T-cell responses.
abstract: OBJECTIVE
  Monitoring cellular immune responses is one prerequisite for the rational development of cancer vaccines.
  
  
  METHODS
  We describe an extensive effort to optimize and validate quantitatively an in vitro T-cell culture method by determining the phenotype and function of both CD4(+) and CD8(+) T cells, including measurement of the phenotype markers CCR7, CD45RA, CD28 and CD27 and the functional markers interferon (IFN)-gamma, interleukin (IL)-2, macrophage inflammatory protein (MIP)-1beta, tumor necrosis factor (TNF)-alpha and CD107a.
  
  
  RESULTS
  Autologous peripheral blood mononuclear cells (PBMC) were potent stimulators that expanded antigen (Ag)-specific CD8(+) T cells during short-term culture with the addition of IL-2 and IL-15 cytokines. Polyfunctional Ag-specific CD4(+) and CD8(+) T cells were detectable using this method.
  
  
  CONCLUSIONS
  Our culture system represents a robust human T-cell culture protocol that permits phenotypic, quantitative and qualitative evaluation of vaccine-induced CD4(+) and CD8(+) T-cell responses.
authors.affiliation: Ludwig Center for Cancer Immunotherapy, Immunology Program, Sloan-Kettering Institute, Newy York, New York 10021, USA.
authors.forename: Yun
authors.initials: Y
authors.lastname: Lin
authors.forename: Humilidad F
authors.initials: HF
authors.lastname: Gallardo
authors.forename: Geoffrey Y
authors.initials: GY
authors.lastname: Ku
authors.forename: Hao
authors.initials: H
authors.lastname: Li
authors.forename: Gregor
authors.initials: G
authors.lastname: Manukian
authors.forename: Teresa S
authors.initials: TS
authors.lastname: Rasalan
authors.forename: Yinyan
authors.initials: Y
authors.lastname: Xu
authors.forename: Stephanie L
authors.initials: SL
authors.lastname: Terzulli
authors.forename: Lloyd J
authors.initials: LJ
authors.lastname: Old
authors.forename: James P
authors.initials: JP
authors.lastname: Allison
authors.forename: Alan N
authors.initials: AN
authors.lastname: Houghton
authors.forename: Jedd D
authors.initials: JD
authors.lastname: Wolchok
authors.forename: Jianda
authors.initials: J
authors.lastname: Yuan
mesh_terms: D000941:Antigens / Q000276:immunology; D015703:Antigens, CD / Q000378:metabolism; D000943:Antigens, Differentiation / Q000378:metabolism; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D019496:Cancer Vaccines; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D005240:Feasibility Studies; D006801:Humans; D009369:Neoplasms / Q000276:immunology* / Q000628:therapy
chemical_list: D000941:Antigens; D015703:Antigens, CD; D000943:Antigens, Differentiation; D019496:Cancer Vaccines; D016207:Cytokines