Western blot analysis of recombinant human GST-VEGF Receptor 2 (Val789-Val1356), untreated or λ phosphatase-treated, using Phospho-VEGF Receptor 2 (Tyr951) (15D2) Rabbit mAb (upper) and VEGF Receptor 2 Antibody # 2472 (lower).
Phospho-VEGF Receptor 2 (Tyr951) (15D2) Rabbit mAb specifically binds to phosphorylated VEGFR2, but not other phosphorylated tyrosine kinases. Western blot analysis of extracts from cells expressing different activated tyrosine kinase proteins, using Phospho-VEGF Receptor-2 (Tyr951) (15D2) Rabbit mAb (upper) or Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 (lower). CKR/PAE cells (lanes 13 and 14) express chimeric receptors containing human CSF-1 extracellular binding domain/mouse VEGF receptor-2 intracellular domain (5). CSF-1 stimulates phosphorylation of Tyr951 of intracellular VEGF receptor-2 domain (lane 13) , which was specifically detected by Phospho-VEGF Receptor-2 (Tyr951) (15D2) Rabbit mAb.
Immunohistochemical analysis of paraffin-embedded HUVEC cells, untreated (left) or VEGF-treated (right), using Phospho-VEGF Receptor-2 (Tyr951) (15D2) Rabbit mAb.
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
NOTE: Do not allow slides to dry at any time during this procedure.
For EDTA: Heat slides in a microwave submersed in 1X EDTA unmasking solution until boiling is initiated; follow with 15 min at a sub-boiling temperature (95°-98°C). No cooling is necessary.
posted June 2005
revised March 2016
Protocol Id: 309
Phospho-VEGF Receptor 2 (Tyr951) (15D2) Rabbit Monoclonal Antibody detects endogenous levels of VEGF receptor 2 only when phosphorylated at Tyr951. The antibody may slightly cross-react with activated VEGF receptor 1, but not with other related tyrosine phosphorylated tyrosine kinases.
Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Tyr951 of human VEGF receptor 2.
Vascular endothelial growth factor receptor 2 (VEGFR2, KDR, Flk-1) is a major receptor for VEGF-induced signaling in endothelial cells. Upon ligand binding, VEGFR2 undergoes autophosphorylation and becomes activated (1). Major autophosphorylation sites of VEGFR2 are located in the kinase insert domain (Tyr951/996) and in the tyrosine kinase catalytic domain (Tyr1054/1059) (2). Activation of the receptor leads to rapid recruitment of adaptor proteins, including Shc, GRB2, PI3 kinase, NCK, and the protein tyrosine phosphatases SHP-1 and SHP-2 (3). Phosphorylation at Tyr1212 provides a docking site for GRB2 binding and phospho-Tyr1175 binds the p85 subunit of PI3 kinase and PLCγ, as well as Shb (1,4,5). Signaling from VEGFR2 is necessary for the execution of VEGF-stimulated proliferation, chemotaxis and sprouting, as well as survival of cultured endothelial cells in vitro and angiogenesis in vivo (6-8).
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