Caspase-3/7 Assay: A Luminescence-Based Assay to Quantify Apoptosis by Measuring Caspase-3/7 Activities in Frog Skin Explants

To extract proteins from a frozen frog toe sample, place the sample in a 1.5-milliliter screw cap microcentrifuge tube, containing 100 microliters of sample buffer and two stainless steel beads.

Use a bead beater at maximum speed to lyse the cells. After this, place the tube on ice for 3 minutes, then, centrifuge the tube to collect the supernatant.

To perform the caspase 3/7 assay in a 384-well plate, add 10 microliters of sample buffer as blank, and 10 microliters of protein extract in triplicate. Then, pipette 10 microliters of commercially available caspase 3/7 reagent to the protein extract and blank wells. Then, shake the plate gently for 15 seconds to mix the two reagents.

Incubate at room temperature and dark for 30 minutes. Finally, use a luminescent plate reader to measure the luminescence of the sample.