BP7223 STAT1 alpha antibody

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Rabbit anti Human, Mouse STAT1 alpha

Product Description for STAT1 alpha

Rabbit anti Human, Mouse STAT1 alpha.
Presentation: Aff - Purified
Product is tested for Western blot / Immunoblot.

Properties for STAT1 alpha

Product Category Primary Antibodies
Quantity 0.1 mg
Synonyms DKFZP686B04100, ISGF-3, P84, P91, STAT1-alpha, STAT1a, STAT91, Signal Transducer And Activator Of Transcription 1 91kDa, Transcription Factor ISGF-3 Components P91/P84
Presentation Aff - Purified
Reactivity Hu, Ms
Applications WB
Clonality Polyclonal
Host Rabbit
Shipping to Worldwide
PDF datasheet View Datasheet
Manufacturer Acris Antibodies GmbH
Material safety datasheet MSDS for Polyclonal Antibodies (de)

Datasheet Extract

Chemically synthesized peptide derived from the C-terminal region of mouse STAT1α
Application Western blot (0.1 - 1.0 µg/ml).
Positive Control Used: NIH3T3 cells; 3T3-L1 adipocytes.
Background The signal transducer and activator of transcription (STAT) family of transcription factors is composed of seven family members (STATs 1, 2, 3, 4, 5A, 5B and 6) that are involved in many cell processes (apoptosis, anti-apoptosis, cell differentiation and proliferation), depending upon the cell type. STATs are activated by ligand binding to cytokine receptors and other receptors associating with Janus kinase (JAK) family members that phosphorylate STAT proteins, causing them to assemble and translocate to the nucleus. In the nucleus, phosphorylated STAT protein complexes bind to DNA and regulate the transcription of particular genes. STAT1α (91 kDa) participates in a signaling pathway which is initiated by IFN-γ. The STAT1α antibody does not recognize STAT1β, which is missing the C-terminal 38 amino acids present in STAT1α. STAT1α is useful as a positive control for measuring total STAT1α protein levels.
Protocols Western Blotting Procedure

1. Lyse approximately 10e7 cells in 0.5 mL of ice cold Cell Lysis Buffer (formulation provided below). This buffer, a modified RIPA buffer, is suitable for recovery of most proteins, including membrane receptors, cytoskeletal-associated proteins, and soluble proteins. Other cell lysis buffer formulations, such as Laemmli sample buffer and Triton-X 100 buffer, are also compatible with this procedure. Additional optimization of the cell stimulation protocol and cell lysis procedure may be required for each specific application.
2. Remove the cellular debris by centrifuging the lysates at 14,000 x g for 10 minutes. Alternatively, lysates may be ultracentrifugedat 100,000 x g for 30 minutes for greater clarification.
3. Carefully decant the clarified cell lysates into clean tubes and determine the protein concentration using a suitable method, such as the Bradford assay. Polypropylene tubes are recommended for storing cell lysates.
4. React an aliquot of the lysate with an equal volume of 2x Laemmli Sample Buffer (125 mM Tris, pH 6.8, 10% glycerol, 10% SDS, 0.006% bromophenol blue, and 130 mM dithiothreitol [DTT]) and boil the mixture for 90 seconds at 100°C.
5. Load 10-30 µg of the cell lysate into the wells of an appropriate single percentage or gradient minigel and resolve the proteins by SDS-PAGE.
6. In preparation for the Western transfer, cut a piece of PVDF membrane slightly larger than the gel. Soak the membrane in methanol for 1 minute, then rinse with ddH2O for 5 minutes. Alternatively, nitrocellulose may be used.
7. Soak the membrane, 2 pieces of Whatman paper, and Western apparatus sponges in transfer buffer (formulation provided below) for 2 minutes.
8. Assemble the gel and membrane into the sandwich apparatus.
9. Transfer the proteins at 140 mA for 60-90 minutes at room temperature.
10. Following the transfer, rinse the membrane with Tris buffered saline for 2 minutes.
11. Block the membrane with blocking buffer (formulation provided below) for one hour at room temperature or overnight at 4°C.
12. Incubate the blocked blot with primary antibody at 0.1-1.0 µg/mL in Tris buffered saline supplemented with 3% non-fat dried milk and 0.1% Tween 20 overnight at 4°C or for two hours at room temperature.
13. Wash the blot with several changes of Tris buffered saline supplemented with 0.1% Tween 20.
14. Detect the antibody band using an appropriate secondary antibody, such as goat F(ab)2 anti-rabbit IgG alkaline phosphatase conjugate or goat F(ab)2 anti-rabbit IgG horseradish peroxidase conjugate in conjunction with your chemiluminescence reagents and instrumentation.

Cell Lysis Buffer Formulation:
10 mM Tris, pH 7.4
100 mM NaCl
1 mM NaF
20 mM Na4P2O7
2 mM Na3VO4
0.1% SDS
0.5% sodium deoxycholate
1% Triton-X 100
10% glycerol
1 mM PMSF (made from a 0.3 M stock in DMSO)
or 1 mM AEBSF (water soluble version of PMSF)
60 µg/mL aprotinin
10 µg/mL leupeptin
1 µg/mL pepstatin
(alternatively, protease inhibitor cocktail such as Sigma Cat. # P2714 may be used)

Transfer Buffer Formulation:
2.4 gm Tris base
14.2 gm glycine
200 mL methanol
Q.S. to 1 liter, then add 1 mL 10% SDS.
Cool to 4°C prior to use.

Tris Buffered Saline Formulation:
20 mM Tris-HCl, pH 7.4
0.9% NaCl

Blocking Buffer Formulation:
100 mL Tris buffered saline
5 gm BSA
0.1 mL Tween 20
General Readings Choudhury, G.C. (2004) A linear signal transduction pathway involving phosphatidylinositol 3-kinase, protein kinase Cepsilon, and MAPK in mesangeal cells regulates interferon-gamma-induced STAT1alpha transcriptional activation. J. Biol. Chem. 279(26):27399-27409.
Burysek, L., et al. (2002) The serine protease plasmin triggers expression of MCP-1 and CD40 in human primary monocytes via activation of p38 MAPK and Janus kinase (JAK)/STAT signaling pathways. J. Biol. Chem. 277(36):33509-33517.
Chen, G., et al. (2001) Expression of the transcription factor STAT-1α in insulinoma cells protects against cytotoxic effects of multiple cytokines. J. Biol. Chem. 276(1):766-772.
Subramaniam, P.S., et al. (2000) The COOH-terminal nuclear localization sequence of interferon γ regulates STAT1α nuclear translocation at an intracellular site. J. Cell Sci. 113:2771-2781.
Singh, K., et al. (1996) Regulation of cytokine-inducible nitric oxide synthase in cardiac myocytes and microvascular endothelial cells. Role of extracellular signal-regulated kinases 1 and 2 (ERK1/ERK2) and STAT1α. J. Biol. Chem. 271(2):1111-1117.
Hague, S.J., et al. (1995) Roles of protein-tyrosine phosphatases in STAT1α-mediated cell signaling. J. Biol. Chem. 270(43):25709-25714.
Storage Store at 2 - 8 °C up to one week or (in aliquots) at -20 °C for longer. Avoid repeated freezing and thawing.
Centrifuge vial before opening.
Shelf life: one year from despatch.
Epitope-specific affinity chromatography
Buffer System:
Dulbecco's phosphate buffered saline (without Mg2+ and Ca2+), pH 7.3
(+/- 0.1), 50% glycerol with 1.0 mg/mL BSA (IgG, protease free) as a carrier, containing
0.05 % sodium azide
Liquid Ig fraction
Aff - Purified
This antibody detects STATalpha.
Human, Mouse.

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