Product Pathways - Jak/Stat Pathway
Phospho-Stat5 (Tyr694) Antibody #9351
| Applications | Reactivity | Sensitivity | MW (kDa) | Source |
|---|---|---|---|---|
| W F ChIP | H M (R) (B) | Endogenous | 90 | Rabbit |
Applications Key:
W=Western Blotting
F=Flow Cytometry
ChIP=Chromatin IP
Reactivity Key:
H=Human
M=Mouse
R=Rat
B=Bovine
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.
Protocols
- 9351:
- ChIP Agarose, ChIP Magnetic, Flow, Western Blotting
Specificity / Sensitivity
Phospho-Stat5 (Tyr694) Antibody detects endogenous levelsof Stat5a only when phosphorylated at Tyr694 and Stat5b when phosphorylated at Tyr699. The antibody does not cross-react with the corresponding phospho-tyrosine residues of other Stat proteins, but does cross-react with EGFR.
Source / Purification
Polyclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Tyr694 of mouse Stat5a. Antibodies are purified by protein A and peptide affinity chromatography.
Western Blotting
Western blot analysis of extracts from TF-1 cells, treated with GM-CSF (25 ng/ml) for the indicated times, using Phospho-Stat5 (Tyr694) Antibody (upper) or Stat5 antibody (lower).
Flow Cytometry
Flow cytometric analysis of K562 cells, untreated (green) or STI571-treated (blue), using Phospho-Stat5 (Tyr694) 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 Phospho-Stat5 (Tyr694) 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 SOCS-3 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.
Background
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).
- Gouilleux, F. et al. (1994) EMBO J. 13, 4361-4369.
- Wakao, H. et al. (1994) EMBO J. 13, 2182-2191.
- Okutani, Y. et al. (2001) Oncogene 20, 6643-6650.
- Demoulin, J.B. et al. (1999) J. Biol. Chem. 274, 25855-25861.
- Dentelli, P. et al. (1999) J. Immunol. 163, 2151-2159.
- Meinke, A. et al. (1996) Mol. Cell. Biol. 16, 6937-6944.
Application References
- Fata, J. E. et al. (2000) The osteoclast differentiation factor osteoprotegerin-ligand is essential for mammary gland development. Cell 103, 41-50. Applications: IHC-P (paraffin)
- Kapur, R. and Zhang, L. (2000) A novel mechanism of cooperation between c-Kit and erythropoietin receptor: Stem cell factor induces the expression of Stat5 and erythropoietin receptor resulting in efficient proliferation and survival by erythropoietin. J. Biol. Chem. 276, 1099-1106. Applications: Western Blotting
- Siren, A. L. et al. (2001) Erythropoietin prevents neuronal apoptosis after cerebral ischemia and metabolic stress. Proc. Natl. Acad. Sci. USA 98, 4044-4049. Applications: Western Blotting
- Reuther, G. W. et al. (2000) Identification and characterization of an activating TrkA deletion mutation in acute myeloid leukemia. Mol. Cell. Biol. 20, 8655-8666. Applications: Western Blotting
- Bergonzini, V. et al. (2004) HIV-Tat promotes cellular proliferation and inhibits NGF-induced differentiation through mechanisms involving Id1 regulation. Oncogene 23, 7701-7711. Applications: Western Blotting
- Brizzi, M. F. et al. (2002) Diabetic LDL inhibits cell-cycle progression via STAT5B and p21waf. J. Clin. Invest. 109 (1), 111. Applications: IC-IF IHC-P (paraffin) Western Blotting
- Hutt, J. A. et al. (2000) Signal transducer and activator of transcription 3 activates CCAAT enhancer-binding protein delta gene transcription in G0 growth-arrested mouse mammary epithelial cells and in involuting mouse mammary gland. J. Biol. Chem. 275, 29123-29131. Applications: Western Blotting
- Weisberg, E. et al. (2011) PLoS One 6, e25351. Applications: Western Blotting
Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!
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- 7074 Anti-rabbit IgG, HRP-linked Antibody
- 7720 Prestained Protein Marker, Broad Range (Premixed Format)
- 7727 Biotinylated Protein Ladder Detection Pack
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- 9363 Stat5 Antibody
This product is intended for research purposes only. The product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
