Interferon gamma Rabbit Recombinant mAb
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使用信息
| 抗体应用 | WB ,ELISA | ||
|---|---|---|---|
| 稀释比例 |
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| 反应性 | Human Mouse Rat | ||
| MW (kDa) | 25kDa | ||
| 抗体类型 | Rabbit | ||
| 浓度 | 1mg/ml | ||
| 储存液配方 | 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. | ||
| 储存条件 (自收到货起) |
Store at –20°C. |
生物描述
| 特异性 | Interferon gamma Rabbit Recombinant mAb detects endogenous level of total Interferon gamma. |
|---|---|
| 背景 | The IFN family of cytokines is divided into type I and type II IFNs. IFN-γ is at present the only type II interferon identified and activates a distinct type II receptor. IFN-γ has a multitude of functions, displaying direct antiviral activity as well as a variety of immunomodulatory and inflammatory roles. Specific actions of IFN-γ signaling include: stimulation of antigen presentation through induced expression of Class I and II major histocompatibility complex (MHC) molecules on the surface of macrophages and T-lymphocytes; antigen processing; control of Th1 and Th2 balance; activation of macrophages, T-lymphocytes and NK cells; stimulation of cytokine production in target cells; and recruitment of cells to the site of injury through increased expression of chemokines and adhesion molecules. IFN-γ also influences cellular state including regulation of the rate of proliferation, differentiation and apoptosis. Interferon-γ is a master checkpoint regulator of cytokine-induced differentiation. It decreases cytokine-induced activation of STAT3 while increasing STAT1 signaling. IFN-γ induces internalization of the common cytokine receptor gp130 by activating p38 signaling. |
实验方法
| WB |
Western Blotting Sample preparation 1. Aspirate media from cultures and Wash the cells with 1X PBS. 2. Lyse cells by adding 1X SDS sample buffer and transfer the extract to a microcentrifuge tube. Keep onice. 3. Sonicate for 10–15 sec to complete cell lysis and shear DNA. 4. Heat a 20 µl sample to 95–100°C for 5 min, then cool on ice. 5. Centrifuge for 5 min (with Microcentrifuge). 6. Load appropriate volumes of samples onto SDS-PAGE gel (loading quantity of protein sample depends on the concentration of extracted proteins). NOTE: At the same time, please load the pre-stained molecular weight markers to determine molecular weights and verify electrotransfer. 7. Electrotransfer to nitrocellulose/PVDF membrane. Membrane Blocking and Antibody Incubations a. Blocking 1. (Optional) After transfer, wash the transferred membrane with TBS for 5 min at room temperature. 2. Incubate the membrane in the blocking buffer for 1 hr at room temperature. 3. Wash three times for 5 min each with TBST. b. Antibodies Incubation 1. Incubate membrane and primary antibody (at the appropriate dilution and diluent recommended) in a primary antibody dilution buffer with gentle agitation overnight at 4°C. 2. Wash three times for 5 min each with TBST. 3. Incubate membrane with an appropriate second antibodydissolved in the blocking buffer with gentle agitation for 1 hr at room temperature. 4. Wash three times for 5 min each with TBST. 5. Proceed with detection. Detection of Proteins 1. After antibodies incubation, Wash membrane three times for 5 minutes in TBST. 2. PrepareECL Reagent (or other chromogenic agents/substrate according to your second antibody). Mix well. 3. Incubate substrate with membrane for 1 minute, remove excess solution (membrane remains wet), wrap in plastic and expose to X-ray film. |
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