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Updated: Aug 11, 2026
Cell-based Flow Cytometry Assay to Measure Cytotoxic Activity
Published on: December 17, 2013
Sensitive and viable identification of antigen-specific CD8+ T cells by a flow cytometric assay for degranulation
Michael R Betts1, Jason M Brenchley, David A Price
1Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 40 Convent Drive, Bethesda, MD 20892, USA. mbetts@mail.nih.gov
Insights
A new method detects antigen-specific CD8+ T cells by measuring cell surface CD107a/b exposure after activation-induced degranulation, reflecting cytolytic potential. This assay offers a more complete assessment of T cell functionality.
Area of Science:
- Immunology
- Cellular Biology
Background:
- Assessing antigen-specific CD8+ T cell function traditionally relies on MHC-class I tetramer staining or intracellular cytokine production.
- These methods do not directly measure the cytolytic potential of CD8+ T cells.
Purpose of the Study:
- To introduce a novel flow cytometry assay for enumerating antigen-specific CD8+ T cells based on activation-induced degranulation.
- To validate this assay as a measure of cytolytic potential.
Main Methods:
- Utilized flow cytometry to detect cell surface expression of CD107a and CD107b following T cell activation.
- Investigated the relationship between CD107a/b surface exposure, intracellular perforin levels, and inhibition by colchicine.
- Correlated CD107a/b expression with IFN-gamma production and antigen-specific cytolytic activity.
Main Results:
- Cell surface CD107a/b exposure, a marker of degranulation, was successfully measured on activated CD8+ T cells.
- CD107a/b surface expression correlated with loss of intracellular perforin and was dependent on degranulation.
- CD107-expressing CD8+ T cells demonstrated antigen-specific cytolytic activity.
Conclusions:
- The novel assay accurately enumerates antigen-specific CD8+ T cells by measuring CD107a/b surface expression, indicating cytolytic potential.
- This method provides a more comprehensive evaluation of CD8+ T cell functionality when combined with other markers like MHC-class I tetramers and intracellular cytokines.
Abstract:
Flow cytometric detection of antigen-specific CD8+ T cells has previously been limited to MHC-class I tetramer staining or intracellular cytokine production, neither of which measure the cytolytic potential of these cells. Here we present a novel technique to enumerate antigen-specific CD8+ T cells using a marker expressed on the cell surface following activation induced degranulation, a necessary precursor of cytolysis. This assay measures the exposure of CD107a and b, present in the membrane of cytotoxic granules, onto the cell surface as a result of degranulation. Acquisition of cell surface CD107a and b is associated with loss of intracellular perforin and is inhibited by colchicine, indicating that exposure of CD107a and b to the cell surface is dependent on degranulation. CD107a and b are expressed on the cell surface of CD8+ T cells following activation with cognate peptide, concordant with production of intracellular IFNgamma. Finally, CD107-expressing CD8+ T cells are shown to mediate cytolytic activity in an antigen-specific manner. Measurement of CD107a and b expression can also be combined with MHC-class I tetramer labeling and intracellular cytokine staining to provide a more complete assessment of the functionality of CD8+T cells expressing cognate T cell receptors (TCR).
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