Revision 1

#12599Store at -20C

1 Kit

(5 x 20 microliters)

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

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3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Mol. Wt Isotype/Source
Phospho-VEGF Receptor 2 (Tyr951) (15D2) Rabbit mAb 4991 20 µl 230 kDa Rabbit IgG
Phospho-VEGF Receptor 2 (Tyr996) Antibody 2474 20 µl 230 kDa Rabbit 
Phospho-VEGF Receptor 2 (Tyr1059) (D5A6) Rabbit mAb 3817 20 µl 230 kDa Rabbit IgG
Phospho-VEGF Receptor 2 (Tyr1175) (D5B11) Rabbit mAb 3770 20 µl 230 kDa Rabbit IgG
VEGF Receptor 2 (D5B1) Rabbit mAb 9698 20 µl 210, 230 kDa Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl Goat 

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

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.

Storage

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.

Background

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).

  1. Meyer, M. et al. (1999) EMBO J 18, 363-74.
  2. Dougher-Vermazen, M. et al. (1994) Biochem Biophys Res Commun 205, 728-38.
  3. Kroll, J. and Waltenberger, J. (1997) J Biol Chem 272, 32521-7.
  4. Takahashi, T. et al. (2001) EMBO J 20, 2768-78.
  5. Holmqvist, K. et al. (2004) J Biol Chem 279, 22267-75.
  6. Karkkainen, M.J. and Petrova, T.V. (2000) Oncogene 19, 5598-605.
  7. Rahimi, N. et al. (2000) J Biol Chem 275, 16986-92.
  8. Claesson-Welsh, L. (2003) Biochem Soc Trans 31, 20-4.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

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    Revision 1
    #12599

    Phospho-VEGF Receptor 2 Antibody Sampler Kit

    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 1 Expand Image
    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).
    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 2 Expand Image
    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.
    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 3 Expand Image
    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.
    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 4 Expand Image
    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 Antibody Sampler Kit: Image 5 Expand Image
    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.
    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 6 Expand Image
    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 Antibody Sampler Kit: Image 7 Expand Image
    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.
    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 8 Expand Image
    Immunohistochemical analysis of paraffin-embedded human angiosarcoma using VEGF Receptor 2 (D5B1) Rabbit mAb.
    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 9 Expand Image
    Immunohistochemical analysis of paraffin-embedded mouse liver using VEGF Receptor 2 (D5B1) Rabbit mAb.
    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 10 Expand Image
    Immunohistochemical analysis of paraffin-embedded HCC827 xenograft using VEGFR2 (D5B1) Rabbit mAb.
    Phospho-VEGF Receptor 2 Antibody Sampler Kit: Image 11 Expand Image
    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 Antibody Sampler Kit: Image 12 Expand Image
    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.