title: Detection of programmed cell death in cells exposed to genotoxic agents using a caspase activation assay.
abstract: Many toxins that individuals are exposed to cause DNA damage. Cells that have sustained DNA damage may attempt to repair the damage prior to replication. However, if a cell has sustained serious damage it cannot repair, it will commit suicide through a genetically regulated programmed cell death (PCD) pathway. Crucial to the ultimate execution of PCD is a family of cysteine proteases called caspases. Activation of these enzymes occurs late enough in the PCD pathway that a cell can no longer avoid cell death, but still earlier than PCD-associated morphological changes or DNA fragmentation. This protocol details a method for using fluorochrome-conjugated caspase inhibitors for the detection of activated caspases in intact cells. The analysis and documentation is performed using fluorescence microscopy.
authors.affiliation: Department of Pediatrics, Wake Forest University School of Medicine, Winston-Salem, NC, USA.
authors.forename: Madhu
authors.initials: M
authors.lastname: Gupta
authors.forename: Madhumita
authors.initials: M
authors.lastname: Santra
authors.forename: Patrick P
authors.initials: PP
authors.lastname: Koty
mesh_terms: D000818:Animals; D017209:Apoptosis; D061945:Caspase Inhibitors / Q000737:chemistry; D020169:Caspases / Q000378:metabolism*; D002478:Cells, Cultured; D004249:DNA Damage; D004789:Enzyme Activation; D057075:Enzyme Assays; D005452:Fluoresceins / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D008856:Microscopy, Fluorescence; D009153:Mutagens / Q000633:toxicity*; D013194:Staining and Labeling; D016707:Tissue Fixation
chemical_list: D061945:Caspase Inhibitors; D005452:Fluoresceins; D005456:Fluorescent Dyes; D009153:Mutagens; C024098:6-carboxyfluorescein; D020169:Caspases