Overview

  • Product nameCaspase 3 Assay Kit (Colorimetric)See all Caspase 3 kits ...
  • Tests
    100 x 1 assay
  • Sample type
    Tissue Extracts, Cell Lysate
  • Assay typeEnzyme activity
  • Assay time
    2h 00m
  • Product overview

    Abcam's Caspase 3 Assay Kit (Colorimetric) provides a simple and convenient means for assaying the activity of caspases that recognize the sequence DEVD. The assay is based on spectrophotometric detection of the chromophore p-nitroaniline (p-NA) after cleavage from the labeled substrate DEVD-pNA. The p-NA light emission can be quantified using a spectrophotometer or a microtiter plate reader 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.


    Visit our FAQs page for tips and troubleshooting.

  • Notes

    Activation of ICE-family proteases/caspases initiates apoptosis in mammalian cells.

     

    If you need more DEVD-pNA substrate, it is available for purchase as ab171038.

  • Tested applicationsFunctional Studiesmore details

Properties

  • RelevanceThe caspase family of cysteine proteases play a key role in apoptosis. Caspase 3 (also known as CPP32, YAMA and apopain) is the most extensively studied apoptotic protein among caspase family members. Caspase 3 is synthesized as an inactive pro enzyme that is processed in cells undergoing apoptosis by self proteolysis and/or cleavage by other upstream proteases (e.g. Caspases 8, 9 and 10). The processed form of Caspase 3 consists of large (17kD) and small (12kD) subunits which associate to form an active enzyme. Caspase 3 is cleaved at Asp28 - Ser29 and Asp175 - Ser176. The active Caspase 3 proteolytically cleaves and activates other caspases (e.g. Caspases 6, 7 and 9), as well as relevant targets in the cells (e.g. PARP and DFF). Alternative splicing of this gene results in two transcript variants which encode the same protein. In immunohistochemical studies Caspase 3 expression has been shown to be widespread but not present in all cell types (e.g. commonly reported in epithelial cells of skin, renal proximal tubules and collecting ducts). Differences in the level of Caspase 3 have been reported in cells of short lived nature (eg germinal centre B cells) and those that are long lived (eg mantle zone B cells). Caspase 3 is the predominant caspase involved in the cleavage of amyloid beta 4A precursor protein, which is associated with neuronal death in Alzheimer's disease.
  • Cellular localizationCytoplasmic
  • Alternative names
    • Apopain
    • Apopain precursor
    • CASP 3
    • CASP3
    • Caspase 3 apoptosis related cysteine protease
    • Caspase 3 p12 subunit
    • Caspase 3 p17 subunit
    • Caspase3
    • CPP 32
    • CPP32
    • CPP32B
    • Cysteine protease CPP32
    • Human cysteine protease CPP32 isoform alpha mRNA complete cds
    • ICE3
    • LICE
    • PARP cleavage protease
    • SCA 1
    • SCA1
    • SREBP cleavage activity 1
    • Yama protein
    see all

Applications

Our Abpromise guarantee covers the use of ab39401 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Functional Studies Use at an assay dependent concentration.

Caspase 3 Assay Kit (Colorimetric) images

  • Induction of Caspase-3 Activity by Anti-Fas Antibody in Jurkat–T cells using 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

Protocols

References for Caspase 3 Assay Kit (Colorimetric) (ab39401)

This product has been referenced in:
  • Haghighi F  et al. The cytotoxic activities of 7-isopentenyloxycoumarin on 5637 cells via induction of apoptosis and cell cycle arrest in G2/M stage. Daru 22:3 (2014). Read more (PubMed: 24393601) »
  • Li MM  et al. Genome-wide microRNA expression profiles in hippocampus of rats with chronic temporal lobe epilepsy. Sci Rep 4:4734 (2014). Functional Studies . Read more (PubMed: 24751812) »

See all 9 Publications for this product

Product Wall

It doesn't work for tissue samples

Inconclusive Good 4/5 (Ease of Use)
Abreviews
We tried to quantify Caspase 3 activity in mouse mammary gland during mammary involution after lactation. We tested different protein amounts fron 50 to 200 microgrames (as indicated in protocol booklet), but the product did not work in any case. We wanted to compare Caspase 3 activity in three samples at different timepoints of involution, and we were not able to observe differences (A), which really exist, as we confirmed by western blot of the same samples (B). We checked the kit was not faulty testig in HC11 mammary cell line, inducing apoptosis with cisplatin 15 ug/ml for 24h (C).
The technical service told us it was a problem with our samples and the did not offer a way to solve our problem.
Then, we found a fluorimetric assay ab39383, which is exactly the same but with a fluorometric substrate instead of a colorimetric one, which is more sensitive as abcam scientific support says in the questions and answer section of the product wall in their website. We aquired the fluorometric substrate and repeated the assay with the same samples. This time, the product worked perfectly (D).
So, for mouse mammary gland tissue samples, I would not recommend this product. It seems it has not enough sensitivity. I would try the fluorimetric assay.
Username

Mr. Adrian Blanco

Verified customer

Submitted Jul 17 2014

Freezing and thawing may partially degrade caspase activity but the protocol, in step 1.2., suggests it will not be a problem: "Transfer supernatant (cytosolic extract) to a fresh tube and put on ice for immediate assay or aliquot and store at -80°C fo...

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ab39401 should recognize all mammalian forms of Caspase 3. One kit is $470, which provides enough solution for one 96 well plate.

This kit is good just for cell and tissue lysates but not plasma. If the you are able to collect the platelets from the plasma in a clean manner you will be able to use the kit with the recommended protocol.

Thank you for contacting us.

This kit has not been tested with serum sample yet, so we can provide Abtrial discount if customer is interested in testing this kit.

Please note that this product is "for research use only and are no...

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Thank you for contacting us and your interest in our products.

The Caspase 3 Assay Kit (Colorimetric) (ab39401) can be used with tissue samples and we would suggest following the following protocol:

1. Induce apoptosis in tissue (10-1...

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Thank you for your enquiry.

I am pleased to confirm that this kit (ab39401 Caspase 3 Assay Kit (Colorimetric))is compatible with mouse samples. Here are the references we have from this kit. I hope this will provide some reassurance that thi...

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Thank you for contacting us. I have answered each of your questions below. Please let me know if you have any additional questions about this or any Abcam product.

1. Does this kit recognize rat caspase-3?

This will recognize all ...

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Thank you for contacting Abcam.

The dilution buffer is to dilute the final samples before reading their absorbance, in case of the undiluted readings being above the detection range of the instrument.
Hope this information has been helpful...

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The dilution buffer is used in case you do not have a plate reader an need to read the concentration manually in a cuvette: Step10. "Read samples at 400 or 405 nm in a microtiter plate reader, or
spectrophotometer using a 100 μl micro-qua...

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"