Rhodamine 110-linked amino acids and peptides as substrates to measure caspase activity upon apoptosis induction in intact cells.
Caspases (cysteine aspartate-specific proteases) are a structurally related group of cysteine proteases that cleave peptide bonds following specific recognition sequences. They play a central role in activating apoptosis of vertebrate cells. To measure apoptosis induced by various stimuli and at an early apoptotic stage, caspases are an ideal target. This is especially the case when apoptotic cells have to be analyzed ex vivo before phagocytes remove them. A new and sensitive caspase assay is based on a substrate that contains only aspartate residues linked to rhodamine 110. With this and similar substrates, we are able to detect intracellular caspase activation by flow cytometry after apoptosis induction in intact hematopoetic cell lines.
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Universitäts-Kinderklinik Ulm, Germany. hubert.hug@medizin.uni-ulm.de
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D000590:Amino Acid Chloromethyl Ketones / Q000378:metabolism; D000596:Amino Acids / Q000378:metabolism*; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017209:Apoptosis / Q000276:immunology; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000378:metabolism; D004789:Enzyme Activation; D005456:Fluorescent Dyes / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D019169:Jurkat Cells / Q000166:cytology* / Q000201:enzymology* / Q000378:metabolism; D010455:Peptides / Q000378:metabolism*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D012235:Rhodamines / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D019311:Staurosporine / Q000494:pharmacology; D013379:Substrate Specificity; D019014:fas Receptor / Q000276:immunology
D000590:Amino Acid Chloromethyl Ketones; D000596:Amino Acids; D000911:Antibodies, Monoclonal; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D005456:Fluorescent Dyes; D010455:Peptides; D012235:Rhodamines; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D019014:fas Receptor; C049025:rhodamine 110; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D019311:Staurosporine