Caspase 3 Antikörper

Produktnummer ABIN5541794

Dieses Ratte Monoklonal-Antikörper erkennt spezifisch Caspase 3 in WB. Er zeigt eine Reaktivität gegenüber Human.

Kurzübersicht für Caspase 3 Antikörper (ABIN5541794)

Target: Caspase 3 (CASP3)

Reaktivität: Human

Wirt: Ratte

Klonalität: Monoklonal

Konjugat: Dieser Caspase 3 Antikörper ist unkonjugiert

Applikation: Western Blotting (WB)

Klon: 7D12

Produktdetails anti-Caspase 3 Antikörper

Spezifität: This antibody reacts with caspase-3 (32 kDa).

Kreuzreaktivität (Details): Does not react with Mouse (NIH/3T3, WR19L) and Rat (Rat1, PC12).

Aufreinigung: Protein G agarose

Immunogen: Recombinant human caspase-3

Isotyp: IgG2b

Anwendungsinformationen

Applikationshinweise: Western blot: 1 μg/mL for chemiluminescence detection system. For details see protocol below. Not recommended for Immunoprecipitation and Immunohistochemistry.

Protokoll: 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 3 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 manufact ure'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 3 times). 9) Incubate the membrane with the 1:10,000 HRP-conjugated anti-rat IgG diluted with 1 % skimmed milk (in PBS, pH 7.2) for 1 hour at room temperature. 10) Wash the membrane with PBS-T (10 minutes x 3 times). 11) Wipe excess buffer on the membrane, then incubate it with appropriate chemiluminescence reagent for 1 minute. 12) Remove extra reagent from the membrane by dabbing with paper towel, and seal it in plastic wrap. 13) Expose to an X-ray film in a dark room for 3 minutes. 14) Develop the film as usual. The condition for exposure and development may vary. (Positive controls for Western blotting HeLa, Raji, Jurkat, HL60, A431, HPB-ALL, KG-1)

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Format: Liquid

Buffer: PBS containing 50 % glycerol, pH 7.2. No preservative is contained.

Konservierungsmittel: Azide free

Lagerung: -20 °C

Informationen zur Lagerung: Upon receipt, store undiluted (in aliquots) at -20°C. Avoid repeated freezing and thawing. Shelf life: One year from despatch.

Details zu Caspase 3

Target: Caspase 3 (CASP3)

Andere Bezeichnung: caspase-3

Hintergrund: Caspase-3 (also known as CPP32, Yama, apopain) is a key member of the caspase family of cysteine proteases. Caspase-3 exists in cells as an inactive 32 kDa proenzyme. During apoptosis procaspase-3 is processed at aspartate residues by self-proteolysis and/or cleavage by upstream caspases, such as caspase-6, -8, or -9. The processed form of caspase-3 consists of large (17 kDa) and small (12 kDa) subunits which associate to form the active tetrameric enzyme tetramer (a pair of heterodimers). The active caspase-3 proteolyti cally cleaves and activates other caspases, as well as relevant targets in the cells (e.g., PARP, SREBPs, and DFF). Activation of procaspase-3 stands at a point of convergence for the two major types of apoptosis signaling pathways-those linked to cell surface death receptors and those linked to mitochondrial release of cytochrome c.

UniProt: P42574

Pathways: Apoptose, Caspase Kaskade in der Apoptose, Sensory Perception of Sound, ER-Nucleus Signaling, Positive Regulation of Endopeptidase Activity, Activated T Cell Proliferation