FL-CTL assay: Fluorolysometric determination of cell-mediated cytotoxicity using green fluorescent protein and red fluorescent protein expressing target cells

Kong Chen, Liansheng Chen, Peilin Zhao, Luis Marrero, Elizabeth Keoshkerian, Alistair Ramsay, Yan Cui

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

Cytotoxic T lymphocytes (CTLs) are crucial effectors against intracellular pathogens and cancer. Accurate and efficient assessment of CTL activity is important for basic and clinical studies. Widely used CTL assays, including the chromium release, JAM test and ELISPOT, involve either radioisotopes or lengthy procedures. Here, we developed a new fluorolysometric CTL assay based on cell-mediated cytolysis of fluorescent protein (GFP or DsRed) expressing cells quantified by one of the fluoro-based methods: flow cytometry, fluorescence microplate reader, or fluorescence microscopy. With flexible detection methods and lentiviral vector transduced stable lines of either GFP+ or DsRed+ cells as targets for antigen presentation and equal number of the other as internal reference for consistency and accuracy, this assay is easy to perform and to scale-up for simultaneous multi-sample analyses. Using two different antigen systems, we demonstrated that this assay is very sensitive to determine primary CTL activity of both in vitro and in vivo primed antigen-specific T cells. Thus, this FL-CTL assay is highly sensitive, reliable, reproducible, economical, convenience and supports broad applications compared to conventional CTL assays.

Original languageEnglish (US)
Pages (from-to)100-114
Number of pages15
JournalJournal of Immunological Methods
Volume300
Issue number1-2
DOIs
StatePublished - May 1 2005
Externally publishedYes

Keywords

  • Antigen-specific CTL function
  • CTL assay
  • Cytotoxic T lymphocytes
  • Fluorescent proteins
  • Fluorometric-based CTL
  • GFP

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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