CST's PathScan^® Cleaved Caspase-3 (Asp175) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of cleaved caspase-3 protein. A Cleaved Caspase-3 (Asp175) Rabbit mAb has been coated onto the microwells. After incubation with cell lysate, the cleaved caspase-3 protein is captured by the coated antibody. Following extensive washing, a Biotinylated Caspase-3 Rabbit Antibody is added to detect the captured cleaved caspase-3 protein. HRP-linked streptavidin is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of cleaved caspase-3 protein.

*Antibodies in this kit are custom formulations specific to kit.

CST's PathScan^® Cleaved Caspase-3 (Asp175) Sandwich ELISA Kit detects endogenous levels of cleaved caspase-3 protein in human cell lines. Using this ELISA kit #7190, a significant induction of cleaved caspase-3 at aspartic acid 175 can be seen in staurosporine-treated HeLa cells, while the level of total caspase-3, analyzed by western blot using Caspase-3 Antibody #9662, remains unchanged (Figure 1). Jurkat cells treated with etoposide show similar results (data not shown). This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

Caspase-3 (CPP-32, Apopain, Yama, SCA-1) is a critical executioner of apoptosis, as it is either partially or totally responsible for the proteolytic cleavage of many key proteins, such as the nuclear enzyme poly (ADP-ribose) polymerase (PARP) (1). Activation of caspase-3 requires proteolytic processing of its inactive zymogen into activated p17 and p12 fragments. Cleavage of caspase-3 requires the aspartic acid residue at the P1 position (2).