Revision 7

#7113Store at +4C

1 Kit

(96 assays)

Species Cross Reactivity

H

UniProt ID:

#P42229, #P51692

Entrez-Gene Id:

#6776, #6777

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Color Storage Temp
Phospho-Stat5 (Tyr694) Mouse mAb Coated Microwells 94647 96 tests +4C
Stat5 Rabbit Detection mAb 90829 1 ea Green (Lyophilized) +4C
Anti-rabbit IgG, HRP-linked Antibody (ELISA Formulated) 13272 1 ea Red (Lyophilized) +4C
Detection Antibody Diluent 13339 11 ml Green +4C
HRP Diluent 13515 11 ml Red +4C
TMB Substrate 7004 11 ml +4C
STOP Solution 7002 11 ml +4C
Sealing Tape 54503 2 ea +4C
ELISA Wash Buffer (20X) 9801 25 ml +4C
ELISA Sample Diluent 11083 25 ml Blue +4C
Cell Lysis Buffer (10X) 9803 15 ml -20C

*The microwell plate is supplied as 12 8-well modules - Each module is designed to break apart for 8 tests.

Description

CST’s PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Stat5a when phosphorylated at Tyr694 and Stat5b at Tyr699. A Phospho-Stat5 Mouse Antibody has been coated onto the microwells. After incubation with cell lysates, phospho-Stat5a (Tyr694) and phospho-Stat5b (Tyr699) proteins are captured by the coated antibody. Following extensive washing, a Stat5 Rabbit Detection Antibody is added to detect the captured phospho-Stat5a and phospho-Stat5b protein. Anti-rabbit IgG, HRP-linked antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of absorbance for this developed color is proportional to the quantity of Stat5a phosphorylated at Tyr694 and Stat5b phosphorylated at Tyr699.

*Antibodies in kit are custom formulations specific to kit.

Specificity/Sensitivity

CST’s PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit #7113 detects endogenous levels of phospho-Stat5a when phosphorylated at Tyr694 and Stat5b at Tyr699. As shown in Figure 1, a significant induction of Stat5 phosphorylation can be detected in EGF-treated HeLa cells using the Phospho-Stat5 (Tyr694) Sandwich ELISA Kit #7113. The level of total Stat5 detected by western analysis remains unchanged. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

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).

  1. Gouilleux, F. et al. (1994) EMBO J 13, 4361-9.
  2. Wakao, H. et al. (1994) EMBO J 13, 2182-91.
  3. Okutani, Y. et al. (2001) Oncogene 20, 6643-50.
  4. Demoulin, J.B. et al. (1999) J Biol Chem 274, 25855-61.
  5. Dentelli, P. et al. (1999) J Immunol 163, 2151-9.
  6. Meinke, A. et al. (1996) Mol Cell Biol 16, 6937-44.

Background References

    Cross-Reactivity Key

    H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    PathScan is a registered trademark of Cell Signaling Technology, Inc.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

    Limited Uses

    Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

    Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

    Revision 7
    #7113

    PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit

    PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit: Image 1 Expand Image
    Figure 1. Treatment of HeLa cells with EGF stimulates phosphorylation of Stat5 at Tyr694, detected by the PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit #7113, but does not affect the level of total Stat5 protein detected by Western analysis. HeLa cells (80-90% confluent) were starved overnight and treated with 100 ng/mL EGF for 40 minutes at 37o C. The absorbance readings at 450 nm are shown in the top figure, while the corresponding Western blots, using Stat5 (3H7) Rabbit mAb #9358 (left panel) or Phospho-Stat5 (Tyr694) (14H2) Mouse mAb #9356 (right panel), are shown in the bottom figure.
    PathScan® Phospho-Stat5 (Tyr694) Sandwich ELISA Kit: Image 2 Expand Image
    Figure 2. The relationship between the protein concentration of the lysate from untreated and EGF-treated HeLa cells and the absorbance at 450 nm is shown.