The mitochondrial electrochemical gradient is often disturbed during apoptosis and can be detected using cationic dyes such as DePsipher™ (5,5’6,6’- tetrachloro-1,1’,3,3’-tetraethylbenz-imidazolylcarbocyanine iodide) or MitoShift™ (tetramethylrhodamine ethyl ester).
Activation of caspases, or cysteinyl proteases, is a necessary event for execution of the apoptotic response. Some of the caspases are activated early in the apoptotic process and their activation is the first step in a cascade of proteolytic cleavage of key proteins and enzymes, including other caspases and poly (ADP-ribose) polymerase (PARP). Since the substrate specificity of the caspases is high, analysis of substrate cleavage also provides a useful biochemical marker.
The movement of some members of the Bcl-2 family from the cytoplasm to the mitochondria and the subsequent associations that occur between them and other mitochondrial membrane associated proteins are indicated to be crucial steps in apoptosis.
DNA fragmentation occurs as one of the final stages of cell death and has long been considered a hallmark of apoptosis and one of the defining biochemical events of the pathway. For detection of the DNA fragmentation associated with apoptosis by DNA laddering, the DNA is isolated and the cleaved fragments are separated by agarose gel electrophoresis. Our Ethidium Bromide DNA Laddering kit provides the necessary reagents for detection of the DNA ladder.
Utilizing a TUNEL-based assay, a series of kits for the in situ detection of apoptosis with colorimetric and fluorometric options was developed. The TACS® kits are tailored for the detection of DNA fragmentation associated with apoptosis in a variety of cell and tissue types and for analysis by different formats that include microscopy, flow cytometry, and 96 well plates.