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7231
PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit
ELISA Kits
ELISA Kit

PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit #7231

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PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit: Image 1

Figure 1. Treatment of H526 cells with SCF stimulates tyrosine phosphorylation of c-Kit, detected by PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit #7231, but does not affect the level of total c-Kit protein detected by PathScan® Total c-Kit Sandwich ELISA Kit #7197. The absorbance readings at 450 nm are shown in the top figure, while the corresponding Western blot using Phospho-c-Kit (Tyr719) Antibody #3391 (right panel) or c-Kit Antibody #3392 (left panel), is shown in the bottom figure.

PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit: Image 2

Figure 2. The relationship between protein concentration of lysates from untreated and SCF-treated H526 lysates and the absorbance at 450 nm is shown. Cells (0.5x106 cells/ml) were serum starved overnight and then treated with Human Stem Cell Factor (hSCF) #8925 (100 ng/ml) for 5 min at 37°C, and then lysed.

PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit: Image 3

Figure 3. Kit specificity as demonstrated by Western analysis of the ELISA microwell captured protein. Lysates were prepared from untreated and SCF-treated H526 cells and incubated in microwells coated with the c-Kit capture antibody. Wells were washed and the captured protein was solubilized in SDS gel loading buffer. Western analysis of untreated and SCF-treated H526 cell starting lysates (lanes 1 & 2) and the captured protein (lanes 3 & 4) was carried out using Phospho-Tyrosine Mouse mAb (P-Tyr-100) (Biotinylated) #9417. The major band detected in the captured material corresponds to the phospho-c-Kit protein (lane 4).

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Important Ordering Details

Custom Ordering Details: When ordering five or more kits, please contact us for processing time and pricing at [email protected].

Supporting Data

REACTIVITY

Application Key:

  • W-Western
  • IP-Immunoprecipitation
  • IHC-Immunohistochemistry
  • ChIP-Chromatin Immunoprecipitation
  • IF-Immunofluorescence
  • F-Flow Cytometry
  • E-P-ELISA-Peptide

Species Cross-Reactivity Key:

  • H-Human
  • M-Mouse
  • R-Rat
  • Hm-Hamster
  • Mk-Monkey
  • 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
  • All-All Species Expected

Product Description

The PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of c-Kit when tyrosine phosphorylated. A c-Kit Mouse mAb has been coated onto the microwells. After incubation with cell lysates, c-Kit (phospho and nonphospho) is captured by the coated antibody. Following extensive washing, a Biotinylated Phospho-Tyrosine Mouse Detection Antibody is added to detect tyrosine phosphorylation of the captured c-Kit protein. HRP-linked Strepavidin is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of c-Kit phosphorylated on tyrosine.

Antibodies in kit are custom formulations specific to kit.

Specificity / Sensitivity

CST's PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit #7231 detects c-Kit when tyrosine phosphorylated. As shown in Figure 1, a significant induction of c-Kit tyrosine phosphorylation can be detected in H526 cells following treatment with SCF using the c-Kit (panTyr) Sandwich ELISA Kit #7231. The level of total c-Kit (phospho and nonphospho) remains unchanged as shown by Western analysis and by PathScan® Total c-Kit Sandwich ELISA Kit #7197. In Figure 3, Western blot analysis of protein captured in the c-Kit antibody coated microwell shows major band corresponding to the phospho-c-Kit protein. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

Background

c-Kit is a member of the subfamily of receptor tyrosine kinases that includes PDGF, CSF-1, and FLT3/flk-2 receptors (1,2). It plays a critical role in activation and growth in a number of cell types including hematopoietic stem cells, mast cells, melanocytes, and germ cells (3). Upon binding with its stem cell factor (SCF) ligand, c-Kit undergoes dimerization/oligomerization and autophosphorylation. Activation of c-Kit results in the recruitment and tyrosine phosphorylation of downstream SH2-containing signaling components including PLCγ, the p85 subunit of PI3 kinase, SHP2, and CrkL (4). Molecular lesions that impair the kinase activity of c-Kit are associated with a variety of developmental disorders (5), and mutations that constitutively activate c-Kit can lead to pathogenesis of mastocytosis and gastrointestinal stromal tumors (6). Tyr719 is located in the kinase insert region of the catalytic domain. c-Kit phosphorylated at Tyr719 binds to the p85 subunit of PI3 kinase in vitro and in vivo (7).

  1. Martin, F.H. et al. (1990) Cell 63, 203-11.
  2. Yarden, Y. et al. (1987) EMBO J 6, 3341-51.
  3. Gommerman, J.L. et al. (1997) J Biol Chem 272, 30519-25.
  4. Sattler, M. et al. (1997) J Biol Chem 272, 10248-53.
  5. Nocka, K. et al. (1990) EMBO J 9, 1805-13.
  6. Hirota, S. et al. (1998) Science 279, 577-80.
  7. Blume-Jensen, P. et al. (2000) Nat Genet 24, 157-62.

Pathways & Proteins

Explore pathways + proteins related to this product.

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's 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's 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.

For Research Use Only. Not For Use In Diagnostic Procedures.
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PathScan is a trademark of Cell Signaling Technology, Inc.