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Human/Mouse Caspase-8 Antibody

Catalog #: AF705
9 Citations Datasheet
Catalog # Availability Size / Price Qty
AF705
AF705-SP
Detection of Human and Mouse Caspase‑8 by Western Blot.
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Product Details
Citations (9)
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Summary Data Examples Preparation and Storage Reconstitution Calculator Background Product Datasheets Related Research Areas

Human/Mouse Caspase-8 Antibody Summary

Species Reactivity
Human, Mouse
Specificity
Detects human and mouse Caspase-8 in Western blots.
Source
Polyclonal Goat IgG
Purification
Antigen Affinity-purified
Immunogen
E. coli-derived recombinant human Caspase-8
Ser234-Asp496
Accession # AAC50645
Formulation
Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
Label
Unconjugated

Applications

Recommended Concentration
Sample
Western Blot
0.5 µg/mL
See below
Simple Western
5 µg/mL
See below
Knockout Validated
Caspase‑8 is specifically detected in HeLa human cervical epithelial carcinoma parental cell line but is not detectable in Caspase‑8 knockout HeLa cell line.
 

Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.

Scientific Data

Western Blot Detection of Human and Mouse Caspase-8 antibody by Western Blot. View Larger

Detection of Human and Mouse Caspase‑8 by Western Blot. Western blot shows lysates of Jurkat human leukemic T cell line and DA3 mouse myeloma cell line treated with 1 µM staurosporine for the indicated time. PVDF membrane was probed with 0.5 µg/mL Goat Anti-Human/Mouse Caspase-8 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF705) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). Specific bands for Caspase-8 precursor were detected at approximately 60 kDa (as indicated in upper panal) and specific bands for cleaved Caspase-8 were detected at approximately 14-18 kDa (as indicated in lower panal). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 4.

Knockout Validated Western Blot Shows Human Caspase-8 Antibody Specificity by Using Knockout Cell Line. View Larger

Western Blot Shows Human Caspase‑8 Specificity by Using Knockout Cell Line. Western blot shows lysates of HeLa human cervical epithelial carcinoma parental cell line and Caspase-8 knockout HeLa cell line (KO). PVDF membrane was probed with 0.5 µg/mL of Goat Anti-Human/Mouse Caspase-8 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF705) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). A specific band was detected for Caspase-8 at approximately 58 kDa (as indicated) in the parental HeLa cell line, but is not detectable in knockout HeLa cell line. GAPDH (Catalog # AF5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Simple Western Detection of Human Caspase-8 antibody by Simple Western<sup>TM</sup>. View Larger

Detection of Human Caspase‑8 by Simple WesternTM. Simple Western lane view shows lysates of Jurkat human acute T cell leukemia cell line untreated or treated with Staurosporine (STS) for 2 hours, loaded at 0.2 mg/mL. Specific bands were detected for Caspase-8 at approximately 59 kDa (as indicated) using 5 µg/mL of Goat Anti-Human/Mouse Caspase-8 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF705) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Preparation and Storage

Reconstitution
Reconstitute at 0.2 mg/mL in sterile PBS.
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Shipping
Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.

Background: Caspase-8

Caspase-8 (Cysteine-aspartic acid protease 8/Casp8a; also named MCH5, FLICA and MACH alpha 1) is a 28 kDa member of the peptidase C14A family of enzymes (1, 2, 3). It is widely expressed and is considered an initiating caspase for the apoptotic cascade (4). Caspase-8 acts on a wide variety of substrates, including procaspases-3, 4, 6, 7, 9 and 10, c-FLIPL and procaspase‑8 itself (1, 5, 6). Human procaspase-8a is a 54‑56 kDa, 479 amino acid (aa) protein (4, 7, 8, 9). It contains two N-terminal death domains (aa 1‑177), followed by a catalytic site that utilizes His317Gly318 plus Cys360. Normally, it is an inactive, cytosolic monomer (1, 10, 11). But following death-domain (DD) containing receptor oligomerization, Caspase-8 is recruited to the death-inducing signaling complex (DISC) that forms around the death domains of the oligomerized receptor (12). FADD/CAP-1 is recruited first, followed by procaspase-8/CAP-4 and, possibly, c-FLIPL and procaspase‑10 (12). The recruitment, or concentration, of procaspase-8 induces homodimerization. This act alone is sufficient for activation. However, the activity level is modest at best, and appears to be directed towards either itself, or c-FLIPL, which is known to form a functional heterodimer with procaspase-8 (5, 11). When directed towards itself, autocleavage occurs first between Asp374Ser375, generating a 43 kDa (p43) N-terminal (aa 1‑374) and an 11 kDa C‑terminal (aa 375‑479) fragment. The C‑terminus is further cleaved between Asp384Leu385 to generate a mature p10 subunit (aa 385‑479). The p43 subunit is next cleaved twice, once between Asp216Ser217, and again between Asp210Ser211 to generate a 26 kDa DD-containing prodomain (aa 1‑210) with an additional 18 kDa mature p18 subunit (aa 217‑374) (12). p18 and p10 noncovalently associate to form a 28 kDa heterodimer, which subsequently associates with another p18:p10 heterodimer to form an active, mature Caspase-8 molecule. This leaves the DISC to act on downstream apoptotic procaspases. In the event procaspase-8 comes to the DISC complexed with c‑FLIPL, c‑FLIPL will be cleaved by procaspase-8, generating a p43 fragment that is analogous to the Caspase-8 p43 subunit. This fragment, however, appears not to be an intermediate in a proteolytic cascade. Rather, it serves as a functional subunit, interacting with TRAF2 and activating NF kappa B. This may account for many of the nonapoptotic activities associated with Caspase-8 (5, 6, 13). Mature human and mouse Caspase-8a heterodimers are 73% aa identical (14).

References
  1. Chowdhury, I. et al. (2008) Comp. Biochem. Physiol. B 151:10.
  2. Boatright, K.M. & G.S. Salvesen (2003) Curr. Opin. Cell Biol. 15:725.
  3. Launay, S. et al. (2005) Oncogene 24:5137.
  4. Srinivasula, S.M. et al. (1996) Proc. Natl. Acad. Sci. USA 93:14486.
  5. Hughes, M.A. et al. (2009) Mol. Cell 35:265.
  6. Lamkanfi, M. et al. (2007) Cell Death Differ. 14:44.
  7. Fernandes-Alnemri, T. et al. (1996) Proc. Natl. Acad. Sci. USA 93:7464.
  8. Boldin, M.P. et al. (1996) Cell 85:803.
  9. Muzio, M. et al. (1996) Cell 85:817.
  10. Donepudi, M. et al. (2003) Mol. Cell 11:543.
  11. Boatright, K.M. et al. (2003) Mol. Cell 11:529.
  12. Golks, A. et al. (2006) Cell Death Differ. 13:489.
  13. Scaffidi, C. et al. (1997) J. Biol. Chem. 272:26953.
  14. Sakamaki, K. et al. (1998) Eur. J. Biochem. 253:399.
Entrez Gene IDs
841 (Human); 12370 (Mouse)
Alternate Names
AIS; androgen receptor; CASP8; Caspase8; Caspase-8; DHTRTFM; Dihydrotestosterone receptorHYSP1; HUMARA; Mch5; NR3C4KD; Nuclear receptor subfamily 3 group C member 4; SMAX1SBMA; spinal and bulbar muscular atrophy

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Citations for Human/Mouse Caspase-8 Antibody

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

9 Citations: Showing 1 - 9
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  1. NF-kB functions as a molecular link between tumor cells and Th1/Tc1 T cells in the tumor microenvironment to exert radiation-mediated tumor suppression.
    Authors: Simon PS, Bardhan K, Chen MR et al.
    Oncotarget.
  2. Significance of calreticulin as a prognostic factor in endometrial cancer
    Authors: Q Xu, C Chen, G Chen, W Chen, D Zhou, Y Xie
    Oncol Lett, 2018-04-13;15(6):8999-9008.
  3. Contrasting roles of H3K4me3 and H3K9me3 in regulation of apoptosis and gemcitabine resistance in human pancreatic cancer cells
    Authors: C Lu, D Yang, ME Sabbatini, AH Colby, MW Grinstaff, NH Oberlies, C Pearce, K Liu
    BMC Cancer, 2018-02-06;18(1):149.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  4. Saturated fatty acids activate caspase-4/5 in human monocytes, triggering IL-1 beta and IL-18 release
    Authors: Nicolas J. Pillon, Kenny L. Chan, Shitian Zhang, Marios Mejdani, Maya R. Jacobson, Alexandre Ducos et al.
    American Journal of Physiology-Endocrinology and Metabolism
  5. Multiplexed, targeted profiling of single-cell proteomes and transcriptomes in a single reaction
    Genome Biol, 2016-09-19;17(1):188.
    Species: Human
    Sample Types: Whole Cells
    Applications: ICC
  6. Ectodomain shedding of EGFR ligands and TNFR1 dictates hepatocyte apoptosis during fulminant hepatitis in mice.
    Authors: Murthy A, Defamie V, Smookler DS
    J. Clin. Invest., 2010-07-12;120(8):2731-44.
    Species: Mouse
    Sample Types: Tissue Homogenates
    Applications: Western Blot
  7. Ellipticine derivative NSC 338258 represents a potential new antineoplastic agent for the treatment of multiple myeloma.
    Authors: Tian E, Landowski TH, Stephens OW, Yaccoby S, Barlogie B, Shaughnessy JD
    Mol. Cancer Ther., 2008-03-04;7(3):500-9.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  8. The B30.2 domain of pyrin, the familial Mediterranean fever protein, interacts directly with caspase-1 to modulate IL-1beta production.
    Authors: Chae JJ, Wood G, Masters SL, Richard K, Park G, Smith BJ, Kastner DL
    Proc. Natl. Acad. Sci. U.S.A., 2006-06-19;103(26):9982-7.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Western Blot
  9. Caspase Inhibition Modulates Monocyte-Derived Macrophage Polarization in Damaged Tissues
    Authors: S Solier, M Mondini, L Meziani, A Jacquel, C Lacout, TV Berghe, Y Julé, JC Martinou, G Pierron, J Rivière, M Deloger, C Dupuy, A Slama-Schw, N Droin, P Vandenabee, P Auberger, E Deutsch, J El-Benna, PM Dang, E Solary
    International Journal of Molecular Sciences, 2023-02-19;24(4):.

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