Overview

  • Product name

    Caspase-3 Inhibitor Assay Kit
    See all Caspase-3 kits
  • Detection method

    Fluorescent
  • Sample type

    Adherent cells, Suspension cells
  • Assay type

    Quantitative
  • Assay time

    1h 30m
  • Product overview

    Abcam's Caspase-3 Inhibitor Drug Detection Kit provides an effective means for screening caspase inhibitors using fluorometric methods. The assay utilizes synthetic peptide substrate DEVD-AFC (AFC, 7-amino-4-trifluoromethyl coumarin). Active Caspase-3 cleaves the synthetic substrate to release free AFC which can then be quantified by fluorometry. Compounds to be screened can directly be added to the reaction and the level of inhibition of Caspase-3 activity can be determined by comparison of the fluorescence intensity in samples with and without the testing inhibitors.
    Visit our FAQs page for tips and troubleshooting.

  • Notes

    Caspases have been shown to play a crucial role in apoptosis induced by various deleterious and physiologic stimuli. Inhibition of caspases can delay apoptosis, implicating a potential role in drug screening efforts.

  • Platform

    Microplate reader

Properties

  • Storage instructions

    Store at -20°C. Please refer to protocols.
  • Components 100 tests
    2X Reaction Buffer 1 x 10ml
    Active Caspase-3 1 x 100 units
    Caspase Substrate DEVD-AFC 1 x 0.5ml
    Caspase-3 Inhibitor 1 x 10µl
    DTT 1 x 100µl
  • Research areas

  • Relevance

    The 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 (e.g. 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 localization

    Cytoplasmic
  • Alternative names

    • A830040C14Rik
    • Apopain
    • Apopain precursor
    • CASP 3
    • CASP-3
    • CASP3
    • CASP3_HUMAN
    • Casp3a
    • Caspase 3 apoptosis related cysteine protease
    • Caspase 3 p12 subunit
    • Caspase 3 p17 subunit
    • Caspase 3, apoptosis-related cysteine peptidase
    • Caspase 3, apoptosis-related cysteine protease
    • Caspase 3, apoptosis-related cysteine protease a
    • Caspase-3 subunit p12
    • Caspase3
    • CC3
    • CP32B
    • CPP 32
    • CPP-32
    • CPP32
    • CPP32B
    • Cysteine protease CPP32
    • EC 3.4.22.56
    • ICE3
    • LICE
    • mldy
    • OTTHUMP00000165052
    • OTTHUMP00000165053
    • OTTHUMP00000165054
    • PARP cleavage protease
    • Procaspase3
    • Protein Yama
    • SCA 1
    • SCA-1
    • SCA1
    • SREBP cleavage activity 1
    • Yama
    • Yama protein
    see all

Images

  • Titration of the Caspase inhibitor Z-VAD(OMe)-FMK (ab120487) (duplicates; +/- SD).

  • Titration of the Caspase-3 specific inhibitor Q-DEVD-OPh (ab142037) (duplicates; +/- SD).

  • Caspase activity (RFU) in presence of 0µM - 40µM of z-VAD-FMK (generic caspase inhibitor), assessed using DEVD-AFC as caspase-3 substrate and following Caspase-3 Inhibitor Drug Detection Kit (ab102491) protocol. Lanes:

    1.- Background Control: no Caspase-3, no z-VMAD-FMK

    2.- Positive Inhibition Control: no Caspase-3, +z-VAD-FMK

    3.- Positive Control: + Caspase-3, no z-VAD-FMK

    4.- Caspase-3 + 2.5µM z-VAD-FMK

    5.- Caspase-3 + 10µM z-VAD-FMK

    6.- Caspase-3 + 40µM z-VAD-FMK

Protocols

References

This product has been referenced in:

  • Bosc D  et al. A new quinoline-based chemical probe inhibits the autophagy-related cysteine protease ATG4B. Sci Rep 8:11653 (2018). Read more (PubMed: 30076329) »
See 1 Publication for this product

Customer reviews and Q&As

Answer

Thank you for contacting us.

The lab advices as follows:

Onewould have to add DTT to the 2X Reaction buffer before using it for reconstitution of the Active Cappase-3.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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