Millipore Sigma Vibrant Logo
Attention: We have moved. Merck Millipore products are no longer available for purchase on MerckMillipore.com.Learn More

APT129 Caspase 8 Colorimetric Activity Assay Kit 25, IETD

Caspase-8 Colorimetric Activity Assay Kits provide a simple & convenient means for assaying the activity of caspases that recognize the sequence IETD.

MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.
Key Applications: Activity Assay
View Products on Sigmaaldrich.com
APT129
25 assays  
Purchase on Sigma-Aldrich

Overview

Replacement Information

Key Spec Table

Species ReactivityKey ApplicationsDetection Methods
MaACTChromogenic
Description
Catalogue NumberAPT129
Brand Family Chemicon®
Trade Name
  • Chemicon
DescriptionCaspase 8 Colorimetric Activity Assay Kit 25, IETD
OverviewActivation of ICE-family proteases/caspases initiates apoptosis in mammalian cells. Chemicon's Caspase-8 Colorimetric Activity Assay Kits provide a simple and convenient means for assaying the activity of caspases that recognize the sequence IETD. The assay is based on spectophotometric detection of the chromophore p-nitroaniline (pNA) after cleavage from the labeled substrate IETD-pNA. The free pNA can be quantified using a spectrophotometer or a microtiter plate reader at 405 nm. Comparison of the absorbance of pNA from an apoptotic sample with an uninduced control allows determination of the fold increase in caspase-8 activity.
Alternate Names
  • FLICE
Materials Required but Not Delivered· Microcentrifuge and 1.5 mL Microcentrifuge tubes

· 37°C Waterbath or Incubator

· Spectrophotometer or Microplate Reader

· 96 well Microtiter Plate
References
Product Information
Components
  • 5X Cell Lysis Buffer (Part No. 90065): 5 mL
  • 5X Assay Buffer (Part No. 90066): 10 mL
  • Caspase-8 Substrate (Ac-IETD-pNA) (Part No. 90081): 250 µL of 2.5 mg/mL solution
  • Caspase-8 Inhibitor (Ac-IETD-CHO) (Part No. 90086): 50 µL of 100 mM (0.05 mg/mL) in DMSO
  • pNA Standard (Part No. 90085): 250 µL of 10 mM in DMSO
Detection methodChromogenic
Quality LevelMQ100
Applications
ApplicationCaspase-8 Colorimetric Activity Assay Kits provide a simple & convenient means for assaying the activity of caspases that recognize the sequence IETD.
Key Applications
  • Activity Assay
Biological Information
Species Reactivity
  • Mammals
Entrez Gene Number
Entrez Gene SummaryThis gene encodes a member of the cysteine-aspartic acid protease (caspase) family. Sequential activation of caspases plays a central role in the execution-phase of cell apoptosis. Caspases exist as inactive proenzymes composed of a prodomain, a large protease subunit, and a small protease subunit. Activation of caspases requires proteolytic processing at conserved internal aspartic residues to generate a heterodimeric enzyme consisting of the large and small subunits. This protein is involved in the programmed cell death induced by Fas and various apoptotic stimuli. The N-terminal FADD-like death effector domain of this protein suggests that it may interact with Fas-interacting protein FADD. This protein was detected in the insoluble fraction of the affected brain region from Huntington disease patients but not in those from normal controls, which implicated the role in neurodegenerative diseases. Many alternatively spliced transcript variants encoding different isoforms have been described, although not all variants have had their full-length sequences determined.
Gene Symbol
  • CASP8
  • MGC78473
  • CASP-8
  • MACH
  • procaspase-8
  • MCH5
  • ALPS2B
  • FLICE
  • CAP4
  • EC 3.4.22.61 [Contains: Caspase-8 subunit p18
  • Caspase-8 subunit p10].
UniProt Number
UniProt SummaryFUNCTION: SwissProt: Q14790 # Most 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. Isoforms 5, 6, 7 and 8 lack the catalytic site and may interfere with the pro-apoptotic activity of the complex.
SIZE: 479 amino acids; 55391 Da
SUBUNIT: Heterotetramer that consists of two anti-parallel arranged heterodimers, each one formed by a 18 kDa (p18) and a 10 kDa (p10) subunit. Interacts with FADD, CFLAR and PEA15. Isoform 9 interacts at the endoplasmic reticulum with a complex containing BCAP31, BAP29, BCL2 and/or BCL2L1.
SUBCELLULAR LOCATION: Cytoplasm.
TISSUE SPECIFICITY: Isoforms 1, 5 and 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.DOMAIN:SwissProt: Q14790 Isoform 9 contains a N-terminal extension that is required for interaction with the BCAP31 complex.
PTM: 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.
DISEASE: SwissProt: Q14790 # Defects 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.
SIMILARITY: Belongs to the peptidase C14 family. & Contains 2 DED (death effector) domains.
Physicochemical Information
Dimensions
Materials Information
Toxicological Information
Safety Information according to GHS
Safety Information
Product Usage Statements
Usage Statement
  • Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Storage and Shipping Information
Storage ConditionsStore kit materials at -20°C up to their expiration date.

Special Precautions:

· After thawing reagents, use immediately or aliquot and freeze at -20oC for longer storage. Avoid repeated freeze/thaw cycles.

· The Caspase-8 substrate and pNA standard are especially light sensitive. Maintain these reagents in amber or covered containers.
Packaging Information
Material Size25 assays
Transport Information
Supplemental Information
Specifications
Global Trade Item Number
Catalogue Number GTIN
APT129 04053252740350