Caspase activity can be detected within whole living cells using the Magic Red substrate-based MR-Caspase assay kit. Designated as the MR-Caspase product line, this user-friendly kit enables researchers to quickly visualize intracellular caspase activity within their particular experimental cell line.

The Magic Red detection kit utilizes the fluorophore cresyl violet. As the 4 amino acid sequence, aspartylglutamylalanylaspartic acid (DEVD) is the optimal sequence for caspase 3 as well as 7, it was coupled to cresyl violet to create the caspase 3/7 substrate, MR-(DEVD)2 (synthesized by Enzyme Systems Products). When bi-substituted via amide linkage to two target caspase sequence groups {(DEVD)2}, cresyl violet does not fluoresce. Following enzymatic hydrolysis at one or both of the aspartic acid amide linkage sites, the mono and non-substituted cresyl violet fluorophores fluoresce red when excited at 550-590 nm.

Detection of intercellular caspases 3 and 7 activities in Staurosporine-induced Jurkat cells using MR (DEVD)2 . Intracellular structures were detected on a Nikon Eclipse E 800 photomicroscope using a 510-560 nm excitation filter and a 570-620 nm emission/barrier filter set at approximately 700X magnification. Photo A shows the corresponding DIC image of the cells.

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References:

• Boonacker, E. and C. J. F. Van Noorden. 2001. Enzyme cytochemical techniques for metabolic mapping in living cells, with special reference to proteolysis. J. Histochem. Cytochem. 49:1473-1486.
• Van Noorden, C. J. F., E. Boonacker, E. R. Bissel, A. J. Meijer, J. Marle and R. E. Smith. 1997. Ala-Pro-cresyl violet, a synthetic fluorogenic substrate for the analysis of kinetic parameters of dipeptidyl peptidase IV (CD26) in individual living rat hepatocytes. Anal. Biochem. 252:71-77.