Caspase3/7ActPlateReAssay, Grn, 200tests

The kit from STEMCELL Technologies is designed for the detection of caspase-3/7 activity in cell lysates prepared from mammalian cells undergoing apoptosis. Caspases are a family of proteases that play a central role in cellular apoptosis through the cleavage of select proteins, resulting in the disassembly of the cell. This process is initiated by pro-apoptotic signals from death receptors or stimuli such as cytotoxic reagents, which activate the initiator caspases and lead to activation of the effector caspases—caspase-3 and caspase-7. Caspase-3 and caspase-7 have a proteolytic substrate selectivity for the peptide sequence Asp-Glu-Val-Asp (DEVD).

The kit includes a fluorogenic caspase-3/7 substrate (Z-DEVD-Rh 110-DVED-Z) and an assay buffer. When added to apoptotic cells, the substrate is cleaved by activated caspase-3/7, generating a green fluorescent signal that can be detected by standard plate readers. The assay is useful for quantifying caspase-3/7 activity in apoptotic cells and for screening caspase-3/7 inhibitors.

Specifications include an excitation wavelength of 500 nm and an emission wavelength of 522 nm with a cutoff of 515 nm, an extinction coefficient of 80,000 cm⁻¹M⁻¹, and a liquid format. Stained cells are observed using a fluorescence microplate reader at Ex/Em = 490/525 nm (cutoff = 515 nm). Camptothecin, a topoisomerase I inhibitor, can be used to induce apoptosis, with Z-VAD-FMK serving as an optional pan-caspase inhibitor for confirming inhibition of caspase-3/7-like activities.