9363:

ChIP Agarose, ChIP Magnetic, Immunoprecipitation, Western Blotting

Stat5 Antibody detects endogenous levels of total Stat5α and β protein.

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to an amino-terminal region within Stat5. Antibodies were purified by protein A and peptide affinity chromatography.

Western Blotting

Western blot analysis of extracts from various cell lines using the Stat5 Antibody.

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 10⁶ BaF3 cells starved of IL-3 for 6 hours followed by induction with IL-3 for 45 minutes, and either 20 μl of Stat5 Antibody or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by Real-Time PCR using SimpleChIP® Mouse CIS Intron 1 Primers #5131, mouse SOCS3 promoter primers, and SimpleChIP® Mouse RPL30 Intron 2 Primers #7015. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.

Stat5 is activated in response to a wide variety of ligands including IL-2, GM-CSF, growth hormone and prolactin. Phosphorylation at Tyr694 is obligatory for Stat5 activation (1,2). This phosphorylation is mediated by Src upon erythropoietin stimulation (3). Stat5 is constitutively active in some leukemic cell types (4). Phosphorylated Stat5 is found in some endothelial cells treated with IL-3, which suggests its involvement in angiogenesis and cell motility (5). Stat5a and Stat5b are independently regulated and activated in various cell types. For instance, interferon treatment predominantly activates Stat5a in U-937 cells and Stat5b in HeLa cells (6).

1. Gouilleux, F. et al. (1994) EMBO J. 13, 4361-4369.
2. Wakao, H. et al. (1994) EMBO J. 13, 2182-2191.
3. Okutani, Y. et al. (2001) Oncogene 20, 6643-6650.
4. Demoulin, J.B. et al. (1999) J. Biol. Chem. 274, 25855-25861.
5. Dentelli, P. et al. (1999) J. Immunol. 163, 2151-2159.
6. Meinke, A. et al. (1996) Mol. Cell. Biol. 16, 6937-6944.

• Mandal, M. et al. (2011) Nat Immunol 12, 1212-20. Applications: Western Blotting

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• 9351 Phospho-Stat5 (Tyr694) Antibody
• 9359 Phospho-Stat5 (Tyr694) (C11C5) Rabbit mAb
• 9314 Phospho-Stat5 (Tyr694) (C71E5) Rabbit mAb
• 9356 Phospho-Stat5 (Tyr694) (14H2) Mouse mAb
• 9358 Stat5 (3H7) Rabbit mAb
• 9171 Phospho-Stat1 (Tyr701) Antibody
• 9131 Phospho-Stat3 (Tyr705) Antibody
• 9361 Phospho-Stat6 (Tyr641) Antibody
• 9914 Phospho-Stat Antibody Sampler Kit
• 7071 Phototope^®-HRP Western Blot Detection System, Anti-rabbit IgG, HRP-linked Antibody
• 7074 Anti-rabbit IgG, HRP-linked Antibody
• 7720 Prestained Protein Marker, Broad Range (Premixed Format)
• 7727 Biotinylated Protein Ladder Detection Pack
• 7003 20X LumiGLO^® Reagent and 20X Peroxide
• 9806 RIPA Buffer (10X)

For Research Use Only. Not For Use In Diagnostic Procedures.