Publication
Title
A real-time fluorometric method for the simultaneous detection of cell death type and rate
Author
Abstract
Several cell death assays have been developed based on a single biochemical parameter such as caspase activation or plasma membrane permeabilization. Our fluorescent apoptosis/necrosis (FANAN) assay directly measures cell death and distinguishes between caspase-dependent apoptosis and caspase-independent necrosis of cells grown in any multiwell plate. Cell death is monitored in standard growth medium as an increase in fluorescence intensity of a cell-impermeable dye (SYTOTOX Green) after plasma membrane disintegration, whereas apoptosis is detected through caspase-mediated release of a fluorophore from its quencher (DEVD-amc). The assay determines the normalized percentage of dead cells and caspase activation per condition as an end-point measurement or in real time (automated). The protocol can be applied to screen drugs, proteins or siRNAs for interference with cell death while simultaneously detecting cell death modality switching between apoptosis and necrosis. Initial preparation may take up to 5 d, but the typical hands-on time is similar to 2 h.
Language
English
Source (journal)
Nature protocols
Publication
2016
ISSN
1754-2189
1750-2799
Volume/pages
11 :8 (2016) , p. 1444-1454
ISI
000379814300010
Pubmed ID
27414760
Full text (Publisher's DOI)
UAntwerpen
Publication type
Subject
External links
Web of Science
Record
Identification
Creation 18.10.2018
Last edited 06.09.2021