Caspase 8 antibody (AA 411-482)

Catalog No. ABIN724205

Product Details anti-Caspase 8 Antibody

Target: Caspase 8 (CASP8)

Binding Specificity: AA 411-482

Reactivity: Human, Mouse, Rat, Monkey, Pig, Sheep

Host: Rabbit

Clonality: Polyclonal

Conjugate: This Caspase 8 antibody is un-conjugated

Application: Western Blotting (WB), ELISA, Immunohistochemistry (Paraffin-embedded Sections) (IHC (p)), Flow Cytometry (FACS), Immunofluorescence (Cultured Cells) (IF (cc)), Immunofluorescence (Paraffin-embedded Sections) (IF (p)), Immunohistochemistry (Frozen Sections) (IHC (fro))

Cross-Reactivity: Human, Monkey, Mouse, Pig, Rat, Sheep

Predicted Reactivity: Dog,Cow,Chicken

Purification: Purified by Protein A.

Immunogen: KLH conjugated synthetic peptide derived from rat CASP8 subunit p10

Isotype: IgG

Application Details

Application Notes: WB 1:300-5000
ELISA 1:500-1000
FCM 1:20-100
IHC-P 1:200-400
IHC-F 1:100-500
IF(IHC-P) 1:50-200
IF(IHC-F) 1:50-200
IF(ICC) 1:50-200

Restrictions: For Research Use only

Independent Validation (WB)

Validation #029759 (Western Blotting)

by: Alamo Laboratories Inc

No.: #029759

Date: 07/03/2014

Lot Number: 131127

Method validated: Western Blotting

Positive Control: HeLa cells

Negative Control: c6/36 mosquito cells (non-reactive species)

Notes: A strong band was observed at the correct molecular weight in the positive control sample. No major bands were observed in the negative sample.

Validation Images

Figure 1: Western blot of lysates from HeLa cells (Lane 1), and c6/36 cells (Lane 2) probed with anti-CASP8 (upper panel) or with anti-Actin for loading control (lower panel).

Full Methods

Primary Antibody:

• Antigen: Caspase 8, Apoptosis-Related Cysteine Peptidase (CASP8) (1:200 dilution)
• Catalog number: ABIN724205
• Lot number: 131127

Secondary Antibody:

• Antigen: Goat Anti-Rabbit IgG (H + L)-HRP Conjugate (1:20,000 dilution)
• Lot number: L170-6515

Full Protocol:

• 1. Total protein extracts were boiled in 1X SDS Sample Buffer containing 1% SDS and 1.25% β-mercaptoethanol at 95°C for 5 min prior to loading.
• 2. 20 μg of boiled extracts were loaded and resolved on 8-16% SDS-polyacrylamide gel.
• 3. The Thermo Scientific - Spectra Multicolor Broad Range (Cat # 26634) were used as molecular mass markers.
• 4. Proteins were then transferred onto PVDF membrane by wet transfer and protein transfer was confirmed with Ponceau-S staining.
• 5. The PVDF membrane was incubated with 25 mL of blocking buffer [Tris Buffered Saline, pH 7.4 plus 0.1% TW20 (TBST)] containing 5% (W/V) BSA at room temperature for 1 h.
• 6. The membrane was rinsed with TBST once.
• 7. The membrane was immersed with the protein side up in the primary antibody solution (CASP8; 1:200) in TBST containing 5% (W/V) BSA and incubated for 16 h at 4°C.
• 8. The membrane was rinsed in TBST thrice for 5 min each.
• 9. The membrane was incubated in the HRP-conjugated secondary antibody solution (Goat anti-rabbit IgG-HRP; 1:20,000) in TBST containing 5% (W/V) BSA and incubated for 1 hour at room temperature (~26°C) with gentle agitation.
• 10. The membrane was rinsed thrice TBST thrice for 5 min each.
• 11. The membrane was rinsed in TBS twice for 30 s each.
• 12. Signals were detected with ECL-2 Substrate. The blot was scanned for 300 s.
• 13. The membrane was rinsed three times TBST.
• 14. Incubated in Acidic Glycine Stripping Buffer at room temperature with gentle agitation for 3 times, 10 min each.
• 15. The membrane was washed in TBST 2 times for 10 min each.
• 16. Repeated Steps 5-12 with the loading control antibody (anti-Actin; 1:6,000) and its matching secondary antibody (Goat anti-rabbit IgG-HRP; 1:20,000).

Experimental Notes: - Nothing to note.

Handling

Format: Liquid

Concentration: 1 μg/μL

Buffer: 0.01M TBS( pH 7.4) with 1 % BSA, 0.02 % Proclin300 and 50 % Glycerol.

Preservative: ProClin

Precaution of Use: This product contains ProClin: a POISONOUS AND HAZARDOUS SUBSTANCE, which should be handled by trained staff only.

Storage: 4 °C,-20 °C

Storage Comment: Shipped at 4°C. Store at -20°C for one year. Avoid repeated freeze/thaw cycles.

Expiry Date: 12 months

References for anti-Caspase 8 antibody (ABIN724205)

Zhang, Zhao, Zhang, Hao, Yu, Min, Li, Ma, Chen, Yi, Tang, Meng, Liu, Wang, Shen, Zhang: "Decrease in male mouse fertility by hydrogen sulfide and/or ammonia can Be inheritable." in: Chemosphere, Vol. 194, pp. 147-157, (2018) (PubMed).

Tulsulkar, Glueck, Hinds, Shah: "Ginkgo biloba Extract Prevents Female Mice from Ischemic Brain Damage and the Mechanism Is Independent of the HO1/Wnt Pathway." in: Translational stroke research, Vol. 7, Issue 2, pp. 120-31, (2016) (PubMed).

Akinrinde, Oyagbemi, Omobowale, Asenuga, Ajibade: "Alterations in blood pressure, antioxidant status and caspase 8 expression in cobalt chloride-induced cardio-renal dysfunction are reversed by Ocimum gratissimum and gallic acid in Wistar rats." in: Journal of trace elements in medicine and biology : organ of the Society for Minerals and Trace Elements (GMS), Vol. 36, pp. 27-37, (2016) (PubMed).

Gao, Liu, Li, Wu, Peng, Jing: "Hispidulin induces mitochondrial apoptosis in acute myeloid leukemia cells by targeting extracellular matrix metalloproteinase inducer." in: American journal of translational research, Vol. 8, Issue 2, pp. 1115-32, (2016) (PubMed).

Daverey, Agrawal: "Curcumin alleviates oxidative stress and mitochondrial dysfunction in astrocytes." in: Neuroscience, Vol. 333, pp. 92-103, (2016) (PubMed).

Wang, Zhao, Yu, Feng, Zhang, Kou, Chu, Cui, Li, Zhang, Shen, Min: "Regulation of steroid hormones and energy status with cysteamine and its effect on spermatogenesis." in: Toxicology and applied pharmacology, Vol. 313, pp. 149-158, (2016) (PubMed).

Zhao, Zhang, Liu, Zhang, Hao, Li, Chen, Shen, Tang, Min, Meng, Wang, Yi, Zhang: "Hydrogen Sulfide and/or Ammonia Reduces Spermatozoa Motility through AMPK/AKT Related Pathways." in: Scientific reports, Vol. 6, pp. 37884, (2016) (PubMed).

Pang, Fan: "Cytotoxic effect and possible mechanisms of Tetracaine on human corneal epithelial cells in vitro." in: International journal of ophthalmology, Vol. 9, Issue 4, pp. 497-504, (2016) (PubMed).

Yuan, Wen, Zhang, Fan: "Cytotoxicity of pilocarpine to human corneal stromal cells and its underlying cytotoxic mechanisms." in: International journal of ophthalmology, Vol. 9, Issue 4, pp. 505-11, (2016) (PubMed).

Peng, Fu, Guo, Zhang, Di, Jiang, Li: "Effects and Mechanism of Baicalin on Apoptosis of Cervical Cancer HeLa Cells I n -v itro." in: Iranian journal of pharmaceutical research : IJPR, Vol. 14, Issue 1, pp. 251-61, (2015) (PubMed).

Fang, Zhang, Qi, Fan, Luo, Liu, Tan: "Evodiamine induces G2/M arrest and apoptosis via mitochondrial and endoplasmic reticulum pathways in H446 and H1688 human small-cell lung cancer cells." in: PLoS ONE, Vol. 9, Issue 12, pp. e115204, (2014) (PubMed).

Wang, Han, Zhou, Wu, Wang, Jia, Lv, Cheng, Pan, Liu, Zhou, Sun: "Inhibition of hydrogen sulfide synthesis provides protection for severe acute pancreatitis rats via apoptosis pathway." in: Apoptosis : an international journal on programmed cell death, Vol. 18, Issue 1, pp. 28-42, (2013) (PubMed).

Wu, Tang, Jiang, Li, Jiang, Liu: "PCSK9 siRNA inhibits HUVEC apoptosis induced by ox-LDL via Bcl/Bax-caspase9-caspase3 pathway." in: Molecular and cellular biochemistry, Vol. 359, Issue 1-2, pp. 347-58, (2011) (PubMed).

Jeng, Dyson: "Direct measurement of the aspartic acid 26 pKa for reduced Escherichia coli thioredoxin by 13C NMR." in: Biochemistry, Vol. 35, Issue 1, pp. 1-6, (1996) (PubMed).

Target Details for Caspase 8

Target: Caspase 8 (CASP8)

Alternative Name: Caspase 8 (CASP8 Products)

Synonyms: casp8-A antibody, xCaspase-8 antibody, casp8 antibody, CASP8 antibody, CASP-8 antibody, FLICE antibody, MACH antibody, Mch5 antibody, ALPS2B antibody, CAP4 antibody, Casp-8 antibody, MCH5 antibody, caspase-8 antibody, zgc:92075 antibody, caspase 8 L homeolog antibody, death related ced-3/Nedd2-like protein antibody, caspase 8 antibody, caspase-8 antibody, caspase-8-like antibody, caspase 8, apoptosis-related cysteine peptidase antibody, casp8.L antibody, LOC661095 antibody, CASP8 antibody, LOC5563818 antibody, casp8 antibody, LOC100222284 antibody, Casp8 antibody

Background:

Synonyms: MACH, Mch5, FLICE, CASP-8, Caspase-8, Casp8

Background: 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 CASP1. 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.

Gene ID: 12370

UniProt: Q9JHX4

Pathways: Apoptosis, Caspase Cascade in Apoptosis, TLR Signaling, Activation of Innate immune Response, Tube Formation, Positive Regulation of Endopeptidase Activity, Toll-Like Receptors Cascades