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To Purchase # 5723S

5723S 1 Kit (200 assays) $524.00.0


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PhosphoSitePlus® Resource

  • Additional protein information
  • Analytical tools


Product Includes Quantity (with Count)
Ac-DEVD-AMC 1 x 1 mg
AMC (7-amino-4-methylcoumarin) 1 x 250 µl
PathScan® Sandwich ELISA Lysis Buffer (1X) 7018 1 x 30 ml
Caspase Assay Buffer (2x) 1 x 30 ml
DTT (Dithiothreitol) 7016 1 x 192.8 mg

Product Description

The Caspase-3 Activity Assay Kit is a fluorescent assay that detects the activity of caspase-3 in cell lysates. It contains a fluorogenic substrate (N-Acetyl-Asp-Glu-Val-Asp-7-amino-4-methylcoumarin or Ac-DEVD-AMC) for caspase-3. During the assay, activated caspase-3 cleaves this substrate between DEVD and AMC, generating highly fluorescent AMC that can be detected using a fluorescence reader with excitation at 380 nm and emission between 420 - 460 nm. Cleavage of the substrate only occurs in lysates of apoptotic cells; therefore, the amount of AMC produced is proportional to the number of apoptotic cells in the sample.

Specificity / Sensitivity

Caspase-3 Activity Assay Kit detects fluorescent AMC dye produced from cleavage of Ac-DEVD-AMC by activated caspase-3 in apoptotic cells. This kit is expected to work in most species. Depending on the cell type and the incubation time applied in the assay, 0.5 - 2x105 cells/well (or 100 μg/well of total lysate protein) is sufficient for most experimental setups. For best results, cell number or lysate concentration titrations are recommended (see Figures 1 and 2). Because caspase-7 shares the same susbtrate sequence as caspase-3, this kit also detects caspase-7 activity.

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).

Caspase-7 (CMH-1, Mch3, ICE-LAP3) has been identified as a major contributor to the execution of apoptosis (3-6). Caspase-7, like caspase-3, is an effector caspase that is responsible for cleaving downstream substrates, such as PARP (3,5). During apoptosis, caspase-7 is activated by upstream caspases through proteolytic processsing at Asp23, Asp198, and Asp206, thereby producing the mature subunits (3,5). Similar to caspases-2 and -3, caspase-7 preferentially cleaves substrates following the recognition sequence DEVD (7).

1.  Fernandes-Alnemri, T. et al. (1994) J Biol Chem 269, 30761-4.

2.  Nicholson, D.W. et al. (1995) Nature 376, 37-43.

3.  Cohen, G.M. (1997) Biochem J 326 ( Pt 1), 1-16.

4.  Thornberry, N.A. et al. (1997) J Biol Chem 272, 17907-11.

5.  Fernandes-Alnemri, T. et al. (1995) Cancer Res 55, 6045-52.

6.  Duan, H. et al. (1996) J Biol Chem 271, 1621-5.

7.  Lippke, J.A. et al. (1996) J Biol Chem 271, 1825-8.

Entrez-Gene Id 836
Swiss-Prot Acc. P42574

For Research Use Only. Not For Use In Diagnostic Procedures.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
PathScan is a trademark of Cell Signaling Technology, Inc.

Caspase-3 Activity Assay Kit