Abstract
The enzyme-linked immunospot (ELISPOT) assay is a highly sensitive tool used to measure the frequency of antigen-specific T-cells in vitro. Among its many applications, this assay is useful for the characterization of cellular immune responses after immunization against measles and other viral pathogens. A description of the measles virus-specific interferon (IFN)-γ ELISPOT assay optimized in our laboratory is described in detail in this chapter. Procedures for the preparation of measles virus, infection of peripheral blood mononuclear cells with measles virus, and the IFN-γ ELISPOT assay are also outlined. These methods can also be broadly adapted to measure activated T-cells to other viral pathogens and/or pathogen-derived peptides. Therefore, the ELISPOT assay can be used for the design, development, and evaluation of new vaccines.
Key Words
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Ovsyannikova I. G., Dhiman N., Jacobson R. M., Vierkant R. A., and Poland G. A. (2003) Frequency of measles virus-specific CD4+ and CD8+ T-cells in subjects seronegative or highly seropositive for measles vaccine. Clin. Diag. Lab. Immunol. 10, 411–416.
Bercovici N., Duffour M-T., Agrawal S., Salcedo M., and Abastado J-P. (2000) New methods for assessing T-cell responses. Clin. Diag. Lab. Immunol. 7, 859–864.
Carter L. L., and Swain S. L. (1997) Single cell analyses of cytokine production. Curr. Opin. Immunol. 9, 177–182.
Nanan R., Rauch A., Kämpgen E., Niewiesk S., and Kreth H. W. (2000) A novel sensitive approach for frequency analysis of measles virus-specific memory T-lymphocytes in healthy adults with a childhood history of natural measles. J. Gen. Virol. 81, 1313–1319.
Carvalho L. H., Hafalla J. C. R., and Zavala F. (2001) ELISPOT assay to measure antigen-specific murine CD8+ T-cell responses. J. Immunol. Methods. 252, 207–218.
Smith J. G., Liu X., Kaufhold R. M., Clair J., and Caulfield M. J. (2001) Development and validation of a gamma interferon ELISPOT assay for quantitation of cellular immune responses to varicella-zoster virus. Clin. Diag. Lab. Immunol. 8, 871–879.
Bailey T, Stark S., Grant A. Hartnett C, Tsang M. and Kalyuzhny A. (2002) A multidonor ELISPOT study of IL-1β, IL-2, IL-4, IL-6, IL-13, IFN-γ, and TNF-α release by cryopreserved human peripheral blood mononuclear cells. J. Immunol. Methods. 270, 171–182.
Kreher C. R., Dittrich M. T., Guerkov R., Boehm B. O., Tary-Lehmann M. (2003) CD4+ and CD8+ cells in cryopreserved human PBMC maintain full functionality in cytokine ELISPOT assays. J. Immunol. Methods. 278, 79–93.
Karber G. (1931) Beitrag zur kollektiven behandlung pharmakologischer reihenversuche. Arch. Exp. Pathol. Pharmakol. 162, 480–483.
Spearman C. (1908) The method of ‘right and wrong cases’ (‘constant stimuli’) without Gauss’s formulae. Br. J. Psychol. 2, 227–242.
Hernandez-Fuentes M. P., Warrens A. N., and Lechler R. I. (2003) Immunologic monitoring. Immunol. Rev. 196, 247–264.
Kalyuzhny A., and Stark S. (2001) A simple method to reduce the background and improve well-to-well reproducibility of staining in ELISPOT assays. J. Immunol. Methods. 257, 93–97.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2005 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Ryan, J.E., Ovsyannikova, I.G., Poland, G.A. (2005). Detection of Measles Virus-Specific Interferon-γ-Secreting T-Cells by ELISPOT. In: Kalyuzhny, A.E. (eds) Handbook of ELISPOT. Methods in Molecular Biology™, vol 302. Humana Press. https://doi.org/10.1385/1-59259-903-6:207
Download citation
DOI: https://doi.org/10.1385/1-59259-903-6:207
Publisher Name: Humana Press
Print ISBN: 978-1-58829-469-2
Online ISBN: 978-1-59259-903-5
eBook Packages: Springer Protocols