Abstract
An assay based on two-color flow cytometry has been developed to measure CTL and NK cell-mediated cytotoxicity. After effector/target cells are incubated together, CTL or NK populations are stained with an effector cell specific PE-conjugated mAb. Subsequently, annexin V-FITC binds to cells expressing phosphatidylserine (an early marker of apoptosis) on the cell surface. Target cells are gated upon as PE-negative and quantified with respect to their annexin V positivity. The shift from annexin V(neg) to annexin V(hi) is a discrete event such that all target cells fall within discernible populations with respect to annexin V. There is a strong correlation between cytotoxicity measured with our assay and a standard 51Cr release assay (r2 = 0.989). The PE/annexin V assay shows increased sensitivity at early timepoints after target/effector cell mixing. In addition, this method allows for analysis of target cells at the single cell level. Therefore, we have described a promising new technique to measure in vitro cell-mediated cytotoxicity. It avoids the potential difficulties of working with radioactive isotopes, and offers increased sensitivity and versatility.
Original language | English (US) |
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Pages (from-to) | 1-9 |
Number of pages | 9 |
Journal | Journal of Immunological Methods |
Volume | 224 |
Issue number | 1-2 |
DOIs | |
State | Published - Apr 22 1999 |
Keywords
- Annexin V
- Cr release
- Flow cytometry
- Human cytotoxic T cells
ASJC Scopus subject areas
- Immunology and Allergy
- Immunology