CST's PathScan^® Phospho-c-Kit (Tyr719) Sandwich ELISA Antibody Pair is offered as an economical alternative to our PathScan^® Phospho-c-Kit (Tyr719) Sandwich ELISA Kit #7298. Capture and Detection Antibodies (100X stocks) and HRP-Conjugated Secondary Antibody (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 Phospho-c-Kit (Tyr719) Rabbit Detection Antibody and Anti-rabbit IgG, HRP-linked Antibody. 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 (Tyr719) protein.
Antibodies in kit are custom formulations specific to kit.

Phosphate Buffered Saline (PBS-20X) #9808
Phosphate Buffered Saline with Tween-20 (PBST-20X) #9809
Cell Lysis Buffer (10X) #9803
TMB Substrate #7004
STOP Solution #7002
Blocking Buffer: 1X PBS/0.5% Tween-20, 1% BSA
96 Well Microplates**
Microplate Reader
** Antibody Pairs have been validated on Corning© 96 Well Clear Polystyrene High Bind Stripwell™ Microplates (#2592).

Notes: Antibody pairs have been optimized using recommended buffers, reagents, plates and the included protocol. Solutions should be made fresh daily.

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).