Caspase-3 Assay Kit (Colorimetric) (ab39401)
- Datasheet
- References (66)
- Protocols
Overview
-
Product nameCaspase-3 Assay Kit (Colorimetric)
See all Caspase-3 kits -
Detection methodColorimetric
-
Sample typeCell Lysate
-
Assay typeEnzyme activity
-
Assay time2h 00m
-
Product overview
Caspase-3 Assay Kit (Colorimetric) ab39401 provides a simple and convenient means for assaying the activity of caspases that recognize the sequence DEVD.
The Caspase-3 assay protocol is based on the formation of the chromophore p-nitroaniline (p-NA) by cleavage from the labeled substrate DEVD-pNA. The p-NA can be quantified using a spectrophotometer or a microtiter plate reader reading absorbance at 400 or 405 nm.
Comparison of the absorbance of p-NA from an apoptotic sample with an uninduced control allows determination of the fold increase in Caspase-3 activity.
Caspase-3 assay protocol summary:
- add samples to wells
- add reaction buffer and DEVD-p-NA substrate and incubate for 60-120 min at 37ºC
- analyze with microplate reader -
Notes
Due to the nature of the substrate, this assay also detects caspase-7 activity.
Activation of ICE-family proteases/caspases initiates apoptosis in mammalian cells.
Other caspase and apoptosis assays
Review the full set of caspase assays, or the apoptosis assay and apoptosis marker guide.
-
PlatformMicroplate reader
Properties
-
Storage instructionsStore at -20°C. Please refer to protocols.
-
Components 100 tests 2X Reaction Buffer 4 x 2ml Cell Lysis Buffer 1 x 100ml DEVD-pNA 1 x 500µl Dilution Buffer 1 x 100ml DTT 1 x 400µl -
Research areas
-
FunctionInvolved in the activation cascade of caspases responsible for apoptosis execution. At the onset of apoptosis it proteolytically cleaves poly(ADP-ribose) polymerase (PARP) at a '216-Asp-
-Gly-217' bond. Cleaves and activates sterol regulatory element binding proteins (SREBPs) between the basic helix-loop-helix leucine zipper domain and the membrane attachment domain. Cleaves and activates caspase-6, -7 and -9. Involved in the cleavage of huntingtin. -
Tissue specificityHighly expressed in lung, spleen, heart, liver and kidney. Moderate levels in brain and skeletal muscle, and low in testis. Also found in many cell lines, highest expression in cells of the immune system.
-
Sequence similaritiesBelongs to the peptidase C14A family.
-
Post-translational
modificationsCleavage by granzyme B, caspase-6, caspase-8 and caspase-10 generates the two active subunits. Additional processing of the propeptides is likely due to the autocatalytic activity of the activated protease. Active heterodimers between the small subunit of caspase-7 protease and the large subunit of caspase-3 also occur and vice versa.
S-nitrosylated on its catalytic site cysteine in unstimulated human cell lines and denitrosylated upon activation of the Fas apoptotic pathway, associated with an increase in intracellular caspase activity. Fas therefore activates caspase-3 not only by inducing the cleavage of the caspase zymogen to its active subunits, but also by stimulating the denitrosylation of its active site thiol. -
Cellular localizationCytoplasm.
- Information by UniProt
-
Alternative names
- A830040C14Rik
- Apopain
- CASP-3
see all
Associated products
-
Corresponding Antibody
-
Related Products
Images
-
Caspase-3 Assay Kit (Colorimetric) (ab39401)H9c2 cells were either untreated (control) or treated with 2µg/ml of recombinant lipocalin-2 (Lcn2) for 24h. Cell lysates were assayed for caspase-3 activity (n = 3, *, p<0.05).Image obtained from Xu G et al., JBC, 2012 Feb 11;28(7):4808-17
-
Caspase-3 Assay Kit (Colorimetric) (ab39401)Caspase-3 in Jurkat lysates (3.3 x10e6 cells) following 20 hour exposure to 2 uM Camptothecin (ab120115) or 10 ng/mL anti-Fas Ab (MBL).
Protocols
References
This product has been referenced in:
- Ponder KG & Boise LH The prodomain of caspase-3 regulates its own removal and caspase activation. Cell Death Discov 5:56 (2019). Read more (PubMed: 30701088) »
- Li Y et al. HOXB4 knockdown enhances the cytotoxic effect of paclitaxel and cisplatin by downregulating ABC transporters in ovarian cancer cells. Gene 663:9-16 (2018). Read more (PubMed: 29660518) »
