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A DELFIA time-resolved fluorescence cell-mediated cytotoxicity assay performed on the PHERAstar FS

NM Patterson, T. Cunningham, X. Jiang, DJ Shapiro University of Illinois 06/2009

Immunosurveillance in tumors is brought about by natural killer cells and cytotoxic T lymphocytes that release perforin and granzyme B to induce apoptosis in tumor cells. However, cancer cells are able to escape this mechanism. In breast cancer cells the hormone estrogen is suspected to protect against granzyme B-induced cell death.

 

A cytotoxicity assay based on time-resolved fluorescence measures cell death in one cell type despite of being co-cultured. To this end, cells of interest are loaded with a fluorescence-enhancing ligand which is trapped in living cells. Only upon cell lysis, an event during cell death, the molecule is released into the medium and forms a Europium-chelate. The chelate is a long-lived fluorophore and its emission signal relates to cell death of the initially labeled cells.

 

The DELFIA® cytotoxicity assay measured on the PHERAstar® FS enabled to investigate NK cell mediated cytolysis. Increasing estrogen was clearly related to reduced cell lysis.

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