Product Pathways - PathScan ELISA
PathScan® Cleaved Caspase-3 (Asp175) Sandwich ELISA Kit #7190
When ordering five or more kits, please contact us for processing time and pricing at firstname.lastname@example.org.
|Kit Includes||Volume||Solution Color|
|Cleaved-Caspase 3 (Asp175) Rabbit mAb coated Microwells||96 tests|
|Biotinylated Caspase-3 Rabbit Detection Antibody||11 ml||Green|
|HRP-linked Streptavidin||11 ml||Red|
|TMB Substrate #7004||11 ml||Colorless|
|STOP Solution #7002||11 ml||Colorless|
|Sealing Tape||2 sheets|
|ELISA Wash Buffer (20X)||25 ml||Colorless|
|ELISA Sample Diluent||25 ml||Blue|
|Cell Lysis Buffer (10X) #9803||15 ml||Yellowish|
Note: 12 8-well modules –Each module is designed to break apart for 8 tests.
Note: Kit should be stored at 4°C with the exception of Cell Lysis Buffer, which is stored at –20°C (packaged separately).
Species enclosed in parentheses are predicted to react based on 100% sequence homology.
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 lysates, 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 kit are custom formulations specific to kit.
Specificity / Sensitivity
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.
ELISA - Western correlation
Figure 1: Treatment of HeLa cells with staurosporine stimulates cleavage of caspase-3 protein, detected by PathScan® Cleaved Caspase-3 (Asp175) Sandwich ELISA Kit #7190 (top figure), but does not affect the level of total caspase-3 protein detected by western blot (bottom figure), using Cleaved Caspase-3 (Asp175) Antibody #9661 (upper panel) or Caspase-3 Antibody #9662 (lower panel).
Caspase-3 (CPP-32, Apoptain, 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).
- Fernandes-Alnemri, T. et al. (1994) J. Biol. Chem. 269, 30761-30764.
- Nicholson, D. W. et al. (1995) Nature 376, 37-43.
- Steiner, P. et al. (2007) Clin Cancer Res 13, 1540-51.
- Levy, D.S. et al. (2009) Blood 113, 1723-9. Applications: ELISA
Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!
- 9661 Cleaved Caspase-3 (Asp175) Antibody
- 9664 Cleaved Caspase-3 (Asp175) (5A1E) Rabbit mAb
- 9662 Caspase-3 Antibody
- 9953 Staurosporine
- 9803 Cell Lysis Buffer (10X)
- 7004 TMB Substrate
- 7002 STOP Solution
For Research Use Only. Not For Use In Diagnostic Procedures.