Cell Signaling Technology

Product Pathways - PathScan ELISA

PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Antibody Pair #7294

Kit Includes Volume Cap Color
c-Kit Mouse Capture Antibody (100X) 0.4 milliliters Pink
Biotinylated Phospho-Tyrosine Mouse Detection Antibody(100X) 0.4 milliliters Blue
HRP-Linked Streptavidin (1000X) 0.04 milliliters Natural
Cell Lysis Buffer (10X) # 9803

Capture and Detection Antibodies are stored at 4°C. HRP-Linked Secondary Antibody is stored at -20°C.

Species Cross-Reactivity

H

Reactivity Key:  H=Human

Description

CST's PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Antibody Pair is being offered as an economical alternative to our PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Kit #7231. Capture and Detection Antibodies (100X stocks) and HRP-Conjugated Streptavidin (1000X stock) are supplied. Sufficient reagents are supplied for 4 x 96 well ELISAs. The c-Kit Mouse Capture Antibody is coated in PBS overnight in a 96 well microplate. After blocking, cell lysates are added followed by Biotinylated Phospho-Tyrosine Mouse Detection Antibody and HRP-conjugated streptavidin. HRP substrate (TMB) is added for color development. The magnitude of the absorbance for this developed color is proportional to the quantity of phospho-c-Kit (panTyr) protein.*Antibodies in this kit are custom formulations specific to the kit.

Sandwich ELISA

Sandwich ELISA

The relationship between protein concentration of lysates from untreated and SCF-treated H526 lysates and the absorbance at 450 nm using PathScan® Phospho-c-Kit (panTyr) Sandwich ELISA Antibody Pair #7294 is shown. Cells (0.5x106 cells/ml) were serum starved overnight and then treated with Human Stem Cell Factor (SCF) #9907 (40 ng/ml) for 5 min at 37?C.

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 such as 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), while 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, 203-211.
  2. Yarden, Y. et al. (1987) EMBO J. 6, 3341-3351.
  3. Gommerman, J.L. et al. (1997) J. Biol. Chem. 272, 30519-30525.
  4. Sattler, M. et al. (1997) J. Biol. Chem. 272, 10248-10253.
  5. Nocka, K. et al. (1990) EMBO J. 9, 1805-1813.
  6. Hirota, S. et al. (1998) Science 279, 577-580.
  7. Blume-Jensen, P. et al. (2000) Nat. Genet. 24, 157-162.

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