Abstract
A major objective of immune analysis in the setting of cancer, cancer vaccination, and therapy is to accurately characterize and isolate functional T-cells elicited by a tumor. Secretion of cytokines is an important function of activated effector and memory T-cells. The cytokine secretion assay (CSA) directly assesses T-cells secreting cytokines after a short restimulation in vitro. Cells can be stained for surface markers enabling characterization of subsets. Viable cytokine-secreting T-cells can be isolated for expansion and further functional testing or adoptive transfer. Since the method is not restricted to any antigen or major histocompatibility complex (MHC) haplotype, it enables us to detect CD8+as well as CD4+T-cells reacting against a wide vary of tumor-associated antigens, ranging from particular known tumor antigens to whole tumor cells and crude tumor lysates. Thus, the method provides a valuable tool to analyze and isolate functional tumor-responsive T-cells according to one of their effector functions.
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References
Goedegebuure, P. S. and Eberlein, T. J. (1995) The role of CD4+tumor-infiltrating lymphocytes in human solid tumors. Immunol. Res. 14, 119–131.
Hung, K., Hayashi, R., Lafond-Walker, A., Lowenstein, C., Pardoll, D., and Levitsky, H. (1998) The central role of CD4+T cells in the antitumor immune response. J. Exp. Med. 188, 2357–2368.
Marzo, A. L., Kinnear, B. F., Lake, R. A., et al. (2000) Tumor-specific CD4+T cells have a major "e;post-licensing"e; role in CTL mediated anti-tumor immunity. J. Immunol. 165, 6047–6055.
Surman, D. R., Dudley, M. E., Overwijk, W. W., and Restifo, N. P. (2000) Cutting edge: CD4+T cell control of CD8+T cell reactivity to a model tumor antigen. J. Immunol. 164(bd2), 562–565.
Ossendorp, F., Mengede, E., Camps, M., Filius, R., and Melief, C. J. (1998) Specific T helper cell requirement for optimal induction of cytotoxic T lymphocytes against major histocompatibility complex class II negative tumors. J. Exp. Med. 187(bd5), 693–702.
Nishimura, T., Nakui, M., Sato, M., et al. (2000) The critical role of Th1-dominant immunity in tumor immunology. Cancer Chemother. Pharmacol. 46(bdSuppl), S52–61.
Manz, R., Assenmacher, M., Pflüger, E., Miltenyi, S., and Radbruch, A. (1995) Analysis and sorting of live cells according to secreted molecules relocated to a cell-surface affinity matrix. Proc. Natl. Acad. Sci. USA 92, 1921–1925.
Brosterhus, H., Brings, S., Leyendeckers, H., et al. (1999) Enrichment and detection of live antigen-specific CD4+and CD8+T cells based on cytokine secretion. Eur. J. Immunol. 29, 4053–4059.
Oelke, M., Moehrle, U., Chen, J. L., et al. (2000) Generation and purification of CD8+su Melan-A-specific cytotoxic T lymphocytes for adoptive transfer in tumor immunotherapy. Clin. Cancer Res. 6, 1997–2005.
Oelke, M., Kurokawa, T., Hentrich, I., et al. (2000) Functional characterization of CD8+ antigen-specific cytotoxic T lymphocytes after enrichment based on cytokine secretion: comparison with the MHC-tetramer technology. Scand. J. Immunol. 52, 544–549.
Pittet, M. J., Zippelius, A., Speiser, D. E., et al. (2001) Ex vivo IFN-γ secretion by circulating CD8 T lymphocytes: implications of a novel approach for T cell monitoring in in-fectious malignant diseases. J. Immunol. 166, 7634–7640.
Becker, C., Pohla, H., Frankenberger, F., et al. (2001) Adoptive tumor therapy with T lymphocytes enriched through an IFN-γ capture assay. Nat. Med. 7(bd10), 1159–1162.
Ge, Q., Palliser, D., Eisen, H. N., and Chen, J. (2002) Homeostatic T cell proliferation in a T cell-dendritic cell coculture system. Proc. Natl. Acad. Sci. USA 99, 2983–2988.
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Becker, C. (2005). Isolation and Expansion of Tumor-Specific CD4+T-Cells by Means of Cytokine Secretion. In: Ludewig, B., Hoffmann, M.W. (eds) Adoptive Immunotherapy: Methods and Protocols. Methods in Molecular Medicine™, vol 109. Humana Press. https://doi.org/10.1385/1-59259-862-5:257
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DOI: https://doi.org/10.1385/1-59259-862-5:257
Publisher Name: Humana Press
Print ISBN: 978-1-58829-406-7
Online ISBN: 978-1-59259-862-5
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