Overview

  • Product name

    Cleaved Caspase-3 Staining Kit (FITC)
    See all Caspase-3 kits
  • Sample type

    Adherent cells, Suspension cells
  • Assay type

    Enzyme activity
  • Assay time

    2h 00m
  • Product overview

    Cleaved Caspase-3 Staining Kit (FITC) ab65613 provides a convenient means for sensitive detection of activated Caspase-3 in living cells.


    The cleaved caspase 3 assay utilizes the Caspase-3 inhibitor, DEVD-FMK, conjugated to FITC (FITC-DEVD-FMK) as a marker. FITC-DEVD-FMK is cell permeable, non-toxic, and irreversibly binds to activated Caspase-3 in apoptotic cells. The FITC label allows detection of activated / cleaved caspase-3 in apoptotic cells directly by fluorescence microscopy, flow cytometry, or fluorescence plate reader.

  • Notes

    Previously called Caspase-3 (active) FITC Staining Kit

    Other caspase and apoptosis assays

    Review the full set of caspase assays, or the apoptosis assay and apoptosis marker guide.

  • Platform

    Microplate reader, Fluor. microscope, Flow cyt.

Properties

  • Storage instructions

    Store at -20°C. Please refer to protocols.
  • Components 100 tests
    FITC-DEVD-FMK 1 x 100µl
    Wash Buffer 2 x 100ml
    Z-VAD-FMK 1 x 10µl
  • 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

  • Caspase-3 activation was measured by flow cytometry, using ab65613, in cells that were left alone or pretreated with QVD.oph and then treated with I-CRP (1.25 U/ml) or a drug that induces DNA strand breaks (100 µM) for 24 h, data was then analyzed and graphed.

  • Caspase-3 in Jurkat cells following 24 hour exposure to 2 uM Camptothecin (CPT; ab120115) with or without 20 nM caspase inhibitor Z-VAD(OMe)-FMK (zVAD; ab120487). Background signal subtracted, duplicates; +/- SD.

Protocols

References

This product has been referenced in:

  • Martínez-Torres AC  et al. IMMUNEPOTENT CRP induces cell cycle arrest and caspase-independent regulated cell death in HeLa cells through reactive oxygen species production. BMC Cancer 18:13 (2018). Flow Cyt ; Human . Read more (PubMed: 29298674) »
  • Våtsveen TK  et al. Artesunate shows potent anti-tumor activity in B-cell lymphoma. J Hematol Oncol 11:23 (2018). Read more (PubMed: 29458389) »
See all 12 Publications for this product

Customer reviews and Q&As

Abreviews
Attempted to use this stain for an in-tact invertebrate organ - specifically, the light organ of the Hawaiian Bobtail squid, Euprymna scolopes. The stain was easy to use, and worked beautifully on some surface-level tissues of the animal. E.g, apoptotic cells with caspase 3 in the tentacle/arms were clearly visible. However, the light organ - which is in the squid body cavity and overlaid on one side with the gills and the other with the funnel - was not compatible with this stain, hence the 2-star review. The following adaptations of the prescribed protocol were attempted:

1) Whole animals incubated with the dye for 30/60 minutes at 37, washed with the wash buffer.
2) Ventrally-dissected animals (light organ exposed) incubated with the dye for 30 and 60 minutes at 37, then washed with wash buffer.
3) Completely removed light organs incubated with the dye for 30 and 60 minutes at 37, then washed with wash buffer.
4) All of the above repeated, but permeabilized by 1 and 2% Triton X 100 for 30 minutes before staining.

Apoptosis in the light organ tissues was always verified before caspase staining by staining with acridine orange. For some reason, however, the FITC-conjugated inhibitor peptide was never observed to penetrate into the light organ. In the picture attached, the arms of the light organ extending outwards showed signs of apoptotic cell death when examined by arcridine orange (image not shown). However, nothing was visible with the caspase dye (image shown).

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Verified customer

Submitted Aug 08 2018

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