Sensolyte homogeneous rh110 caspase 3 7 assay kit

Caspase activity was determined fluorometrically by using cell type specific assay kits. To evaluate the activity of caspases 3/7, 8, 9, and 12 in the HL-60 cells, we used a SensoLyte® Homogeneous Rh110 Caspase-3/7 Assay Kit (AnaSPEC, San Jose, CA, USA); an APOPCYTO™ Caspase-8 Fluorometric Assay Kit (MBL, Nagoya, Japan); an APOPCYTO™ Caspase-9 Fluorometric Assay Kit; and a Caspase 12 Assay Kit (Fluorometric; ab65664, Abcam Japan, Tokyo, Japan). Alternatively, to evaluate caspase 3 activity in the T98G, MDA-MB-231 and KATO III cells an APOPCYTO™ Caspase-3 Colorimetric Assay Kit was used. Cellular fluorescence was measured using a fluorescence microplate reader (Varioskan Flash, Thermo Fisher Scientific).

Executioner caspase (DEVDase) activity was measured using a SensoLyte Homogeneous Rh110 Caspase‐3/7 Assay Kit (Anaspec, Inc., 71114) according to the manufacturer's instructions. Briefly, 1.0 × 10⁴ cells were plated into each well of a 96‐well plate for overnight incubation and subjected to the experimental treatments indicated. At the designated time‐points, 33 μL of freshly prepared fluorogenic caspase substrate (DEVD‐Rho110) was added to wells containing 100 μL cell culture media and incubated at room temperature for 18 hours. Following incubation, fluorescence was determined using a microplate fluorometer (ex. 496 nm, em. 520 nm; Molecular Devices, SpectraMax M2).