Visualization and quantification of T cell–mediated cytotoxicity using cell-permeable fluorogenic caspase substrates (original) (raw)

Nature Medicine volume 8, pages 185–189 (2002)Cite this article

Abstract

We have developed a non-radioactive flow-cytometry assay to monitor and quantify the target-cell killing activities mediated by cytotoxic T lymphocytes (CTLs). This flow-cytometry CTL (FCC) assay is predicated on measurement of CTL-induced caspase activation in target cells through detection of the specific cleavage of fluorogenic caspase substrates. Here we show that this assay reliably detects antigen-specific CTL killing of target cells, and demonstrate that it provides a more sensitive, more informative and safer alternative to the standard 51Cr-release assay most often used to quantify CTL responses. The FCC assay can be used to study CTL-mediated killing of primary host target cells of different cell lineages, and enables the study of antigen-specific cellular immune responses in real time at the single-cell level. As such, the FCC assay can provide a valuable tool for studies of infectious disease pathogenesis and development of new vaccines and immunotherapies.

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Acknowledgements

We thank R. Ahmed for provision of reagents and helpful advice; P. Henkart, R. Mittler and S. Staprans for critical review of the manuscript; and T. Cinotte for technical assistance. This study was supported by NIH grant P01AI46007 and R2AI49089. M.B.F. is an Elizabeth Glaser Scientist of the Pediatric AIDS Foundation.

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Authors and Affiliations

  1. Emory Vaccine Center and Emory University School of Medicine, Atlanta, Georgia, USA
    Luzheng Liu, Ann Chahroudi, Guido Silvestri, Mary E. Wernett, William J. Kaiser, Jeffrey T. Safrit, John D. Altman & Mark B. Feinberg
  2. OncoImmunin, Gaithersburg, Maryland, USA
    Akira Komoriya & Beverly Z. Packard

Authors

  1. Luzheng Liu
  2. Ann Chahroudi
  3. Guido Silvestri
  4. Mary E. Wernett
  5. William J. Kaiser
  6. Jeffrey T. Safrit
  7. Akira Komoriya
  8. John D. Altman
  9. Beverly Z. Packard
  10. Mark B. Feinberg

Corresponding author

Correspondence toMark B. Feinberg.

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Liu, L., Chahroudi, A., Silvestri, G. et al. Visualization and quantification of T cell–mediated cytotoxicity using cell-permeable fluorogenic caspase substrates.Nat Med 8, 185–189 (2002). https://doi.org/10.1038/nm0202-185

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