Overview

  • Product nameCaspase-8 (active) FITC Staining Kit
    See all Caspase-8 kits
  • Sample type
    Adherent cells, Suspension cells
  • Assay typeEnzyme activity
  • Assay time
    2h 00m
  • Product overview

    Caspase 8 (active) FITC Staining Kit (ab65614) provides a convenient means for sensitive detection of activated caspase 8 in living cells. The assay utilizes the caspase 8 inhibitor, IETD-FMK, conjugated to FITC (FITC-IETD-FMK) as a marker. FITC-IETD-FMK is cell permeable, non-toxic, and irreversibly binds to activated caspase 8 in apoptotic cells. The FITC label allows detection of activated caspase-8 in apoptotic cells directly by fluorescence microscopy, flow cytometry, or fluorescence plate reader.
    Visit our FAQs page for tips and troubleshooting.

  • Notes

    Activation of caspases plays a central role in apoptosis.

  • Tested applicationsSuitable for: FM, Functional Studies, Flow Cytmore details

Properties

  • FunctionMost upstream protease of the activation cascade of caspases responsible for the TNFRSF6/FAS mediated and TNFRSF1A induced cell death. Binding to the adapter molecule FADD recruits it to either receptor. The resulting aggregate called death-inducing signaling complex (DISC) performs CASP8 proteolytic activation. The active dimeric enzyme is then liberated from the DISC and free to activate downstream apoptotic proteases. Proteolytic fragments of the N-terminal propeptide (termed CAP3, CAP5 and CAP6) are likely retained in the DISC. Cleaves and activates CASP3, CASP4, CASP6, CASP7, CASP9 and CASP10. May participate in the GZMB apoptotic pathways. Cleaves ADPRT. Hydrolyzes the small-molecule substrate, Ac-Asp-Glu-Val-Asp-
    -AMC. Likely target for the cowpox virus CRMA death inhibitory protein. Isoform 5, isoform 6, isoform 7 and isoform 8 lack the catalytic site and may interfere with the pro-apoptotic activity of the complex.
  • Tissue specificityIsoform 1, isoform 5 and isoform 7 are expressed in a wide variety of tissues. Highest expression in peripheral blood leukocytes, spleen, thymus and liver. Barely detectable in brain, testis and skeletal muscle.
  • Involvement in diseaseDefects in CASP8 are the cause of caspase-8 deficiency (CASP8D) [MIM:607271]. CASP8D is a disorder resembling autoimmune lymphoproliferative syndrome (ALPS). It is characterized by lymphadenopathy, splenomegaly, and defective CD95-induced apoptosis of peripheral blood lymphocytes (PBLs). It leads to defects in activation of T-lymphocytes, B-lymphocytes, and natural killer cells leading to immunodeficiency characterized by recurrent sinopulmonary and herpes simplex virus infections and poor responses to immunization.
  • Sequence similaritiesBelongs to the peptidase C14A family.
    Contains 2 DED (death effector) domains.
  • DomainIsoform 9 contains a N-terminal extension that is required for interaction with the BCAP31 complex.
  • Post-translational
    modifications
    Generation of the subunits requires association with the death-inducing signaling complex (DISC), whereas additional processing is likely due to the autocatalytic activity of the activated protease. GZMB and CASP10 can be involved in these processing events.
    Phosphorylated upon DNA damage, probably by ATM or ATR.
  • Cellular localizationCytoplasm.
  • Information by UniProt
  • Alternative names
    • ALPS2B
    • Amyotrophic lateral sclerosis 2 chromosomal region candidate gene 12 protein
    • Apoptotic cysteine protease
    • Apoptotic protease Mch-5
    • Apoptotic protease Mch5
    • CAP4
    • CASP-8
    • CASP8
    • CASP8_HUMAN
    • Caspase 8
    • Caspase 8 apoptosis related cysteine peptidase
    • Caspase-8 subunit p10
    • CED 3
    • FADD Like ICE
    • FADD-homologous ICE/CED-3-like protease
    • FADD-like ICE
    • FLICE
    • FLJ17672
    • ICE-like apoptotic protease 5
    • MACH
    • MACH alpha 1/2/3 protein
    • MACH beta 1/2/3/4 protein
    • MCH5
    • MGC78473
    • MORT1 associated ced 3 homolog
    • MORT1-associated CED-3 homolog
    • OTTHUMP00000163717
    • OTTHUMP00000163720
    • OTTHUMP00000163724
    • OTTHUMP00000163725
    • OTTHUMP00000165062
    • OTTHUMP00000165063
    • OTTHUMP00000165064
    • OTTHUMP00000206552
    • OTTHUMP00000206582
    see all

Applications

Our Abpromise guarantee covers the use of ab65614 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
FM Use at an assay dependent concentration.
Functional Studies Use at an assay dependent dilution.
Flow Cyt Use at an assay dependent dilution.

Caspase-8 (active) FITC Staining Kit images

  • Active caspase 8 in control Jurkat cells (10e6/mL) or cells treated for five hours with 10 ug/mL Cyclohexamide (CHX) (ab120093) or four hours with 25 ug/mL Mitomycin C (MitoC) (ab120797). Background signal subtracted, duplicates; +/- SD.

  • Active caspase 8 in Jurkat cells following four hours exposure to 50 ng/mL anti-Fas Ab (αFas) (MBL), five hours with 10 μg/mL cyclohexamide (CHX) (ab120093), or one hour pretreatment with CHX followed by four hours with αFas. Background signal subtracted, duplicates; +/- SD.

  • Caspase 8 activation of Hela (left) and SiHa (right) cells after in vitro treatment with pentoxylline (PTX) or cisplatin (CIS) either alone or in combination. Results represent the mean ± SD of three independent experiments carried out in triplicate. (*) p<0.001 vs CTL.

    Image obtained from Hernandez-Flores G et al; BMC Cancer, 2011 Nov 11; 11:483

Protocols

References for Caspase-8 (active) FITC Staining Kit (ab65614)

This product has been referenced in:
  • Hernandez-Flores G  et al. Pentoxifylline sensitizes human cervical tumor cells to cisplatin-induced apoptosis by suppressing NF-kappa B and decreased cell senescence. BMC Cancer 11:483 (2011). Read more (PubMed: 22074157) »

See 1 Publication for this product

Product Wall

Thank you for contacting us. 1.) You may use Caspase-8 negative cells (cells that were not stimulated to induce Caspase-8) for your negative control in Flow Cytometry. 2.) For a viability dye, you may try using PI but this will need to be optimi...

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