Caspase 8 Assay Kit (Colorimetric) (ab39700)

Overview

  • Product nameCaspase 8 Assay Kit (Colorimetric)
    See all Caspase 8 kits
  • Detection methodColorimetric
  • Tests
    100 x 1 assay
  • Sample type
    Cell Lysate
  • Assay typeEnzyme activity
  • Assay time
    2h 00m
  • Product overview

    Caspase 8 Assay Kit (colorimetric) (ab39700) is a simple and convenient assay to quantify the activity of caspase 8 in cell lysates, based on the recognition of the sequence Ile-Glu-Thr-Asp (IETD). The assay is based on spectrophotometric detection of the chromophore p- nitroanilide (pNA) after it is cleaved from the labeled substrate IETD-pNA. The pNA light emission can be quantified using a spectrophotometer or a microtiter plate reader at OD 400 – 405 nm. Comparison of the absorbance of pNA from an apoptotic sample with an un-induced control allows determination of the fold increase in Caspase 8 activity.



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  • Notes

    The caspase family of highly conserved cysteine proteases play an essential role in programmed cell death (including apoptosis, pyropoptosis and necroptosis).

    Mammalian caspases can be subdivided into three functional groups: initiator caspases (Caspase 2, 8, 9 and 10), executioner caspases (Caspase 3, 6 and 7), and inflammatory caspases (Caspase 1, 4, 5, 11 and 12). Initially synthesized as inactive pro-caspases, caspases become rapidly cleaved and activated in response to granzyme B, death receptors and apoptosome stimuli. Caspases will then cleave a range of substrates, including downstream caspases, nuclear proteins, plasma membrane proteins and mitochondrial proteins, ultimately leading to cell death.

    Caspase 8 (CASP8/FLICE, EC:3.4.22.61) is the most upstream caspase involved in the activation of apoptosis through the extrinsic pathway, mediated by CD95 (Fas) receptor and TNFR. Caspase 8 is recruited to the receptors through the adapter molecule FADD, resulting in the formation of the aggregate called death-inducing signaling complex (DISC) and proteolytic activation of caspase 8. The active dimeric enzyme is then liberated from the DISC and free to activate downstream apoptotic proteases. Inhibition or inactivation of caspase 8 is required for induction of necroptosis.

  • Tested applicationsSuitable for: Functional Studiesmore details

Properties

  • RelevanceCaspases are cysteine proteases, expressed as inactive precursors, that mediate apoptosis by proteolysis of specific substrates. Caspases have the ability to cleave after aspartic acid residues. There are two classes of caspases involved in apoptosis; initiators (activation by receptor cluster) and effectors (activation by mitochondrial permeability transition). Proapoptotic signals autocatalytically activate initiator caspases, such as Caspase 8 and Caspase 9. Activated initiator caspases then process effector caspases, such as Caspase 3 and Caspase 7, which in turn cause cell collapse.
  • Cellular localizationCytoplasmic
  • Alternative names
    • Apoptotic cysteine protease
    • CAP4
    • CASP8
    • Caspase 8
    • CED 3
    • FLICE
    • MACH
    • MACH5
    see all

Applications

Our Abpromise guarantee covers the use of ab39700 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 8 Assay Kit (Colorimetric) images

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

  • Active caspase 8 in control (CTRL) Jurkat cells (10e6/mL) or in cells after four hours exposure to 50 ng/mL anti-Fas Ab (αFas) (MBL), or pretreated one hour with 50 μM Z-VAD(OMe)-FMK (ab120487) followed by four hours with αFas. Background signal subtracted, duplicates; +/- SD.

Protocols

References for Caspase 8 Assay Kit (Colorimetric) (ab39700)

ab39700 has not yet been referenced specifically in any publications.

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