IFN-γ Secretion Assay – Detection Kits, human

IFN-γ Secretion Assay – Detection Kits, human

The IFN-γ Secretion Assay was developed for the sensitive detection of human IFN-γ-secreting cells.

Background information

IFN-γ (interferon-gamma) is predominantly secreted by activated CD8
and CD4
memory and effector T cells and by NK cells. It is mainly involved in the regulation of inflammatory immune responses. These T
1 types of immune mechanisms are effective against intracellular pathogens and tumors. IFN-γ-secreting T cells can also be involved in immunological disorders, such as autoimmune reactions.

Downstream applications

Virus-specific T cells were investigated after stimulation with peptides or proteins derived from influenza virus
, and ADV
Virus-specific T cells were expanded
in vitro1,2,3,5,6,9
showing highly specific and very efficient killing of target cells and have been analyzed for TCR clonotypes
The IFN-γ Secretion Assay was used for the isolation and analysis of antigen-specific T cells from PBMCs after stimulation with Tetanus Toxoid
, minor histocompatibility antigens, and tumor antigens
. The assay was also used to purify and analyze tumor-specific T cells from T cell lines
,for the isolation of functional antigen-specific, IFN-γ-secreting T cells reacting to other tumor antigens, e.g. SSX
, or HER2
from PBMCs or TILs (tumor infiltrating lymphocytes)
The IFN-γ Secretion Assay was used to counterstain peptide-MHC-tetramer-labeled Melan A-specific CD8
T cells to analyze functionality of the tetramer-positive cells.
The IFN-γ Secretion Assay was also used for isolation and functional characterization of allergen-specific T cells.
Furthermore, the IFN-γ Secretion Assay was used for epitopemapping of MHC class II peptides.
IFN-γ-secreting human NK cells were isolated using the IFN-γ Secretion Assay.
IFN-γ Secretion Assay reagents were reported to cross-react with chimpanzee cells
but not with rhesus macaque cells.
The IFN-γ Secretion Assay can also be used for two-color cytokine analysis
and allows counterstaining of peptide- MHC-tetramer-labeled cells
. It can also be combined with flow cytometric proliferation assays
  • Selected references

    1. Moody, D. B. et al. (2017) CD1b-mycolic acid tetramers demonstrate T-cell fine specificity for mycobacterial lipid tails. Eur. J. Immunol. 47(9): 1525-1534
    2. Lehrnbecher, T. et al. (2017) Impact of human mesenchymal stromal cells on antifungal host response against Aspergillus fumigatus. Oncotarget 8(56): 95495-95503
    3. Brosterhus, 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
    4. Bissinger, A. L. et al. (2002) Isolation and expansion of human cytomegalovirus- specific cytotoxic T lymphocytes using interferon-gamma secretion assay. Exp. Hematol. 30: 1178-1184
    5. Bitmansour, A. D. et al. (2002)
      ex vivo
      analysis of human CD4(+) memory T cell activation requirements at the single clonotype level.
      J. Immunol. 169: 1207-1218
    6. Bickham, K. et al. (2001)
      EBNA1-specific CD4
      T cells in healthy carriers of Epstein-Barr virus are primarily Tʜ1 in function.
      J. Clin. Invest. 107: 121-130
    7. Cohen, G. B. et al. (2002) Clonotype tracking of TCR repertoires during chronic virus infections. Virology 304: 474-484
    8. Koehne, G. et al. (2002) Quantitation, selection, and functional characterization of Epstein-Barr virus-specific and alloreactive T cells detected by intracellular interferon-gamma production and growth of cytotoxic precursors. Blood 99: 1730-1740
    9. Altfeld, M. et al. (2001) Vpr is preferentially targeted by CTL during HIV-1 infection. J. Immunol. 167: 2743-2752
    10. Douek, D. C. et al. (2002)
      HIV preferentially infects HIV-specific CD4
      T cells.
      Nature 417: 95-98
    11. Lichterfeld, M. et al. (2004)
      HIV-1-specific cytotoxicity is preferentially mediated by a subset of CD8
      T cells producing both interferon-gamma and tumor necrosis factor-alpha.
      Blood 104: 487-494
    12. Lee, S. K. et al. (2002) The functional CD8 T cell response to HIV becomes type-specific in progressive disease. J. Clin. Invest. 110: 1339-1347
    13. Desombere, I. et al. (2003) The interferon gamma secretion assay: a reliable tool to study interferon gamma production at the single cell level. J. Immunol. Methods 286: 167-185
    14. Meidenbauer, N. et al. (2003) Survival and tumor localization of adoptively transferred Melan-A-specific T cells in melanoma patients. J. Immunol. 170: 2161-2169
    15. Ayyoub, M. et al. (2004)
      An immunodominant SSX-2-derived epitope recognized by CD4
      T cells in association with HLA-DR.
      J. Clin. Invest. 113: 1225-1233
    16. Pittet, M. J. et al. (2001)
      Ex vivo
      IFN-gamma secretion by circulating CD8 T lymphocytes: implications of a novel approach for T cell monitoring in infectious and malignant diseases.
      J. Immunol. 166: 7634-7640
    17. Märten, A. et al. (2002) Generation of activated and antigen-specific T cells with cytotoxic activity after co-culture with dendritic cells. Cancer Immunol. Immunother. 51: 25-32
    18. Meyer zu Büschenfelde, C. M. et al. (2001) The generation of both T killer and Tʜ cell clones specific for the tumor-associated antigen HER2 using retrovirally transduced dendritic cells. J. Immunol. 167: 1712-1719
    19. Becker, C. et al. (2001) Adoptive tumor therapy with T lymphocytes enriched through an IFN-gamma capture assay. Nat. Med. 7: 1159-1162
    20. Novak, E. J. et al. (2001)
      Tetramer-guided epitope mapping: rapid identification and characterization of immunodominant CD4
      T cell epitopes from complex antigens.
      J. Immunol. 166: 6665-6670
    21. Deniz, G. et al. (2002) Human NK1 and NK2 subsets determined by purification of IFN-gamma-secreting and IFN-gamma-nonsecreting NK cells. Eur. J. Immunol. 32: 879-884
    22. Akdis, M. et al. (2004) Immune responses in healthy and allergic individuals are characterized by a fine balance between allergen-specific T regulatory 1 and T helper 2 cells. J. Exp. Med. 199: 1567-1575
    23. Meyer-Olson, D. et al. (2003) Analysis of the TCR beta variable gene repertoire in chimpanzees: identification of functional homologs to human pseudogenes. J. Immunol. 170: 4161-4169
    24. Gutzmer, R. et al. (2003) Human dendritic cells express the IL-18R and are chemoattracted to IL-18. J. Immunol. 171: 6363-6371
    25. Feuchtinger T. et al. (2004)
      Isolation and expansion of human adenovirus-specific CD4
      and CD8
      T cells according to IFN-gamma secretion for adjuvant immunotherapy.
      Exp. Hematol. 32: 282-289
  • Special protocols

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