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Optimization and validation of a robust human T-cell culture method for monitoring phenotypic and polyfunctional antigen-specific CD4 and CD8 T-cell responses.

BACKGROUND AIMS
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.