Western blot analysis of extracts from CKR/PAE cells expressing chimeric receptors containing human CSF-1 extracellular binding domain/mouse VEGF receptor 2 intracellular domains (Rahimi, N. et al. (2000) J. Biol. Chem. 275, 16986-16992), using Phospho-VEGF Receptor 2 (Tyr996) Antibody (upper) or VEGF receptor 2 antibody (lower).
Western blot analysis of PAE/CKR cells, untreated or stimulated with CSF-1, using Phospho-VEGF Receptor 2 (Tyr1059) (D5A6) Rabbit mAb (upper) and VEGF Receptor 2 (55B11) Rabbit mAb #2479 (lower). PAE/CKR cells express a chimeric receptor made up of human CSF-1 receptor extracellular domain and mouse VEGF receptor 2 transmembrane and intracellular domains.
Western blot analysis of extracts from HUVEC, untreated or stimulated with VEGF #9943, using Phospho-VEGF Receptor 2 (Tyr1175) (D5B11) Rabbit mAb (upper) and VEGF Receptor 2 (55B11) Rabbit mAb (lower) #2479.
Flow cytometric analysis of fixed and permeabilized HeLa cells (blue, negative) and HUVEC cells (green, positive) using VEGF Receptor 2 (D5B1) Rabbit mAb (solid lines) or a concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
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).
Confocal immunofluorescent analysis of rat pancreas using VEGF Receptor 2 (D5B1) Rabbit mAb (green) and β-Catenin (L54E2) Mouse mAb (IF Preferred) #2677 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
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 human angiosarcoma using VEGF Receptor 2 (D5B1) Rabbit mAb.
Immunohistochemical analysis of paraffin-embedded mouse liver using VEGF Receptor 2 (D5B1) Rabbit mAb.
Immunohistochemical analysis of paraffin-embedded HCC827 xenograft using VEGFR2 (D5B1) Rabbit mAb.
Western blot analysis of extracts from HUVE cells, HeLa cells and mouse heart using VEGF Receptor 2 (D5B1) Rabbit mAb for VEGFR2 expression (upper) and β-Actin (D6A8) Rabbit mAb (#8457) for loading control. (The 80 kDa bands represent partial degradation product).
|Phospho-VEGF Receptor 2 (Tyr951) (15D2) Rabbit mAb 4991||20 µl||
||H M||230||Rabbit IgG|
|Phospho-VEGF Receptor 2 (Tyr996) Antibody 2474||20 µl||
|Phospho-VEGF Receptor 2 (Tyr1059) (D5A6) Rabbit mAb 3817||20 µl||
||H M||230||Rabbit IgG|
|Phospho-VEGF Receptor 2 (Tyr1175) (D5B11) Rabbit mAb 3770||20 µl||
||H M||230||Rabbit IgG|
|VEGF Receptor 2 (D5B1) Rabbit mAb 9698||20 µl||
||H M R||210, 230||Rabbit IgG|
|Anti-rabbit IgG, HRP-linked Antibody 7074||100 µl||
The Phospho-VEGF Receptor 2 Antibody Sampler Kit provides an economical means of evaluating the VEGFR2 tyrosine kinase and several phosphorylation sites that are involved in its activation. The kit includes enough antibody to perform two western blot experiments with each primary antibody.
The phospho-VEGF Receptor 2 (Tyr1175) (D5B11) Rabbit mAb detects endogenous VEGF Receptor 2 at tyrosine 1175 and may cross-react with other tyrosine-phosphorylated receptor tyrosine kinases, including VEGFR1 and FLT3. The phospho-VEGF Receptor 2 (Tyr1059) (D5A6) Rabbit mAb
detects endogenous levels of VEGF Receptor 2 at tyrosine 1059 and can detect VEGF receptors 1 and 3 when phosphorylated at corresponding tyrosine residues due to identical sequences within the epitope region. The phospho-VEGF Receptor 2 (Tyr951) (15D2) Rabbit mAb detects endogenous VEGF Receptor 2 at tyrosine 951 and may slightly cross-react with activated VEGF receptor 1, but not with other related tyrosine phosphorylated tyrosine kinases. The phospho-VEGF Receptor 2 (Tyr996) Antibody detects transfected VEGF Receptor 2 only when phosphorylated at tyrosine 996. The antibody cross-reacts with tyrosine-phosphorylated PDGF receptor. The VEGF Receptor 2 (D5B1) Rabbit mAb detects endogenous total VEGF Receptor 2.
Polyclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Tyr996 of human VEGFR2. Polyclonal antibodies are purified by protein A and peptide affinity chromatography. Monoclonal antibodies are produced by immunizing animals with synthetic phosphopeptides corresponding to residues surrounding Tyr951, Tyr1059 or Tyr1175. VEGFR2 monoclonal antibody is produced by immunizing animals with a recombinant protein containing the carboxy-terminal 150 residues of human VEGFR2.
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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