Caspase 8 抗体 (C-Term)
Quick Overview for Caspase 8 抗体 (C-Term) (ABIN5541755)
抗原
See all Caspase 8 (CASP8) 抗体适用
宿主
克隆类型
标记
应用范围
克隆位点
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抗原表位
- AA 180-480, C-Term
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特异性
- This antibody reacts with caspase-8.
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交叉反应 （详细）
- Does not work with Mouse (WR19L) and Rat (PC12).
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纯化方法
- Protein A agarose
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免疫原
- Recombinant human caspase s-8 corresponding to C-terminal amino acids (180-480 aa)
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亚型
- IgG2b
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anti-Caspase 8 (CASP8) (AA 411-482) antibody
CASP8 适用: 人, 小鼠, 大鼠, 猴, Pig, 绵羊 WB, ELISA, FACS, IHC (p), IF (cc), IF (p), IHC (fro) 宿主: 兔 Polyclonal unconjugated
anti-Caspase 8 (CASP8) (AA 1-479) antibodyCASP8 适用: 人 WB, IHC, IF 宿主: 兔 Polyclonal unconjugated
anti-Caspase 8 (CASP8) (AA 188-280) antibodyCASP8 适用: 人, 小鼠, 大鼠 WB, ELISA, FACS, IHC (p), IF (cc), IF (p), IHC (fro) 宿主: 兔 Polyclonal unconjugated
anti-Caspase 8 (CASP8) (AA 217-384) antibodyCASP8 适用: 人 WB, IHC, ICC, IF 宿主: 兔 Polyclonal unconjugated
anti-Caspase 8 (CASP8) (AA 7-201) antibodyCASP8 适用: 人 WB, IHC, ICC, IF 宿主: 兔 Polyclonal unconjugated
anti-Caspase 8 (CASP8) (AA 385-479) antibodyCASP8 适用: 人 WB, IHC, ICC, IP 宿主: 兔 Polyclonal unconjugated
anti-Caspase 8 (CASP8) antibodyCASP8 适用: 人, 小鼠, 大鼠, 猴 WB, IHC, ELISA, FACS 宿主: 小鼠 Monoclonal 1H11 unconjugated
anti-Caspase 8 (CASP8) (AA 410-449), (N-Term) antibodyCASP8 适用: 人, 小鼠, 大鼠 WB, IHC, ICC, FACS, IHC (fro) 宿主: 兔 Polyclonal unconjugated
anti-Caspase 8 (CASP8) (AA 217-374) antibodyCASP8 适用: 人 WB, IHC, ELISA, IF 宿主: 兔 Polyclonal unconjugated
anti-Caspase 8 (CASP8) antibodyCASP8 适用: 小鼠 WB, IHC, ELISA, ICC, FACS 宿主: 大鼠 Monoclonal 3B10 unconjugated
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应用备注
- Western blot: 1 μg/mL for chemiluminescence detection system. Immunoprecipitation: 2 μg/500 μL of cell extract. For details see protocol below.
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实验流程
- SDS-PAGE & Western Blotting 1) Wash the cells 3 times with PBS and suspend with 10 volume of cold Lysis buffer (50 mM Tris-HCl, pH 7.2, 250 mM NaCl, 0.1 % NP-40, 2 mM EDTA, 10 % glycerol) containing appropriate protease inhibitors. Incubate it at 4 o C with rotating for 30 minutes, then sonicate briefly (up to 10 seconds). 2) Centrifuge the tube at 12,000 x g for 10 minutes at 4 o C and transfer the supernatant to another tube. Measure the protein concentration of the supernatant and add the cold Lysis buffer to make 8 mg/mL solution. 3) Mix the sample with equal volume of Laemmli's sample buffer. 4) Boil the samples for 2 minutes and centrifuge. Load 10 μ L of the sample per lane in a 1 mm thick SDS-polyacrylamide gel for electrophoresis. 5) Blot the protein to a polyvinylidene difluoride (PVDF) membrane at 1 mA/cm 2 for 1 hour in a semi-dry transfer system (Transfer Buffer: 25 mM Tris, 190 mM glycine, 20 % MeOH). See the manufacture's manual for precise transfer procedure. 6) To reduce nonspecific binding, soak the membrane in 10 % skimmed milk (in PBS, pH 7.2) for 1 hour at room temperature, or overnight at 4 o C. 7) Incubate the membrane with primary antibody diluted with PBS, pH 7.2 containing 1 % skimmed milk as suggest in the APPLICATIONS for 1 hour at room temperature. (The concentration of antibody will depend on condition.) 8) Wash the membrane with PBS-T [0.05 % Tween-20 in PBS] (5 minutes x 6 times). 9) Incubate the membrane with the 1:10,000 HRP-conjugated anti-mouse IgG diluted with 1 % skimmed milk (in PBS, pH 7.2) for 1 hour at room temperature. 10) Wash the membrane with PBS-T (5 minutes x 6 times). 11) Wipe excess buffer on the membrane, then incubate it with appropriate chemilumin escence reagent for 1 minute. Remove extra reagent from the membrane by dabbing with paper towel, and seal it in plastic wrap. 12) Expose to an X-ray film in a dark room for 3 minutes. Develop the film as usual. The condition for exposure and development may vary. (Positive controls for Western blotting Jurkat, Raji, HeLa, U937, MCF7, HEp-G2, ZR-75-1) Immunoprecipitation 1) Wash the cells 3 times with PBS and suspend with 10 volume of cold Lysis buffer (50 mM Tris-HCl pH 7.2, 250 mM NaCl, 0.1 % NP-40, 2 mM EDTA, 10 % glycerol) containing appropriate protease inhibitors. Incubate it at 4 o C with rotating for 30 minutes, then sonicate briefly (up to 10 seconds). 2) Centrifuge the tube at 12,000 x g for 10 minutes at 4 o C and transfer the supernatant to another tube. 3) Add primary antibody as suggest in the APPLICATIONS into 500 μ L of the supernatant. Mix well and incubate with gentle agitation for 30-120 minutes at 4 o C. Add 20 μ L of 50 % protein A agarose beads resuspended in the cold Lysis buffer. Mix well and incubate with gentle agitation for 60 minutes at 4 o C. 4) Wash the beads 3-5 times with the cold Lysis buffer (centrifuge the tube at 2,500 x g for 10 seconds). 5) Resuspend the beads in 20 μ L of Laemmli's sample buffer, boil for 3-5 minutes, and centrifuge for 5 minutes. Use 10 μ L/lane for the SDS-PAGE analysis. (See SDS-PAGE & Western blotting.)
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限制
- 仅限研究用
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状态
- Liquid
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缓冲液
- PBS containing 50 % glycerol, pH 7.2. No preservative is contained.
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储存液
- Azide free
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储存条件
- -20 °C
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储存方法
- Upon receipt, store undiluted (in aliquots) at -20°C. Avoid repeated freezing and thawing. Shelf life: One year from despatch.
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- Caspase 8 (CASP8)
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别名
- caspase-8,flice
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背景
- Caspase-8 (FLICE/MACH/Mch5) is a member of the ICE (interleukin- 1 converting enzyme)/CED-3 family cysteine protease. It is the most upstream protease that recei ves the activation signal from the Fas (APO1/CD95) and TNFR1 (Tumor Necrosis Factor Receptor 1) to initiate the ap optotic protease cascade that leads to activation of ICE/CED-3 family proteases. Caspase-8 has high homologous region to the ICE/CED-3 family in C-terminal and two death effecter domains (DED) in N-terminal. Binding of caspase-8 to FADD (MORT1) through association of their DED, and consequent activation of the caspases by th eir proteolytic cleavage, are thought to be critical steps in the initiation of Fas- and TNFR1-induced apoptosis. Recen tly the inhibitor of Fas- and TNFR1-induced apoptosis is identified, called I-FLICE (FLIP/Casper/FLAME/CASH). I-FLICE has high homology to caspase-8 and it contains two DED, which interacts with caspase-8 and FADD, and potently inhibits Fas- and TNFR1-induced apoptosis.
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UniProt
- Q14790
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途径
- Apoptosis, Caspase Cascade in Apoptosis, TLR signaling, Activation of Innate immune Response, Tube Formation, Positive Regulation of Endopeptidase Activity, Toll-Like Receptors Cascades
抗原
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