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8696
Angiogenesis Antibody Sampler Kit
Primary Antibodies
Antibody Sampler Kit

Angiogenesis Antibody Sampler Kit #8696

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Angiogenesis Antibody Sampler Kit: Image 1

Confocal immunofluorescent images of HUVE cells untreated (left) or stimulated with Vascular Endothelial Growth Factor (VEGF) #9943 (right) and labeled with Phospho-VEGF Receptor 2 (Tyr1175) (19A10) Rabbit mAb (top, green) and VEGF Receptor 2 (55B11) Rabbit mAb #2479 (bottom, green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

Angiogenesis Antibody Sampler Kit: Image 2

Flow cytometric analysis of Jurkat cells, untreated (green) or treated with LY294002 #9901, Wortmannin #9951, and U0126 #9903 (50 μM, 1 μM, and 10 μM, 2 hr; blue) using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (solid lines) or 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.

Angiogenesis Antibody Sampler Kit: Image 3

Western blot analysis of extracts NIH/3T3 cells, serum-starved or treated with human Platelet-Derived Growth Factor BB hPDGF-BB #8912 (100 ng/ml, 15 min), using Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb (upper) or Src (36D10) Rabbit mAb #2109 (lower).

Angiogenesis Antibody Sampler Kit: Image 4

Western blot analysis of extracts from A549 cells, untreated or treated with calf intestinal phosphatase (CIP), using Phospho-FAK (Tyr397) (D20B1) Rabbit mAb (upper) and FAK Antibody #3285 (lower).

Angiogenesis Antibody Sampler Kit: Image 5

Flow cytometric analysis of Jurkat cells, untreated (blue) or anisomycin-treated (green), using Phospho-p38 MAPK (Thr180/Tyr182) (D3F9) XP® Rabbit mAb compared to a nonspecific negative control antibody (red).

Angiogenesis Antibody Sampler Kit: Image 6

Flow cytometric analysis of human peripheral blood mononuclear cells, untreated (left column) or treated with cross-linked anti-CD3 plus anti-CD28 (10ug/ml each, 15 min; right column), using Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb (top row) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (bottom row), and co-stained with CD3 (UCHT1) Mouse mAb (APC Conjugate) #19881. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.

Angiogenesis Antibody Sampler Kit: Image 7

Flow cytometric analysis of Jurkat cells, treated with U0126 (10 µM, 2 hrs; green) or treated with TPA #4174 (200 nM, 30 min; blue) using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb (solid lines) or 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.

Angiogenesis Antibody Sampler Kit: Image 8

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.

Angiogenesis Antibody Sampler Kit: Image 9

Immunohistochemical analysis of paraffin-embedded HUVE cells, untreated (left) or VEGF treated (right).

Angiogenesis Antibody Sampler Kit: Image 10

Confocal immunofluorescent analysis of C2C12 cells, LY294002-treated (left) or insulin-treated (right), using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin #8953 (red). Blue pseudocolor = DRAQ5®#4084 (fluorescent DNA dye).

Angiogenesis Antibody Sampler Kit: Image 11

Confocal immunofluorescent analysis of COS cells, untreated (left) or anisomycin-treated (right) using Phospho-p38 MAPK (Thr180/Tyr182) (D3F9) XP® Rabbit mAb (green). Actin filaments have been labeled with DY-554 phalloidin (red).

Angiogenesis Antibody Sampler Kit: Image 12

Flow cytometric analysis of Jurkat cells, treated with U0126 #9903 (10uM, 2 hr; blue) or TPA #4174 (200nM, 30 min; green) using Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb (solid lines) or 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) was used as a secondary antibody.

Angiogenesis Antibody Sampler Kit: Image 13

Confocal immunofluorescent analysis of Drosophila egg chambers, untreated (top) or λ phosphatase-treated (bottom), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb #4370 (green) and S6 Ribosomal Protein (54D2) Mouse mAb #2317 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

Angiogenesis Antibody Sampler Kit: Image 14

Phospho-VEGF Receptor 2 (Tyr1175) (19A10) 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 (Tyr1175) (19A10) Rabbit mAb (upper) and Phospho-Tyrosine mAb (P-Tyr-100) #9411 (lower). CKR/PAE cells (lanes 12 and 13) express chimeric receptors containing human CSF-1 extracellular binding domain/mouse VEGF receptor-2 intracellular domain (7). CSF-1 stimulates phosphorylation of Tyr1175 of intracellular VEGF receptor-2 domain (lane 12), which was specifically detected by Phospho-VEGF Receptor-2 (Tyr1175) (19A10) Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 15

Immunohistochemical analysis of paraffin-embedded MDA-MB-468 xenograft using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (left) or PTEN (138G6) Rabbit mAb #9559 (right). Note the presence of P-Akt staining in the PTEN deficient MDA-MB-468 cells.

Angiogenesis Antibody Sampler Kit: Image 16

Immunohistochemical analysis of paraffin-embedded human colon carcinoma using Phospho-p38 MAPK (Thr180/Tyr182) (D3F9) XP® Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 17

Confocal immunofluorescent analysis of A-431 cells, serum starved (left) or treated with hEGF #8916 (100 ng/mL for 15 min) using Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

Angiogenesis Antibody Sampler Kit: Image 18

Confocal immunofluorescent analysis of HT1080 cells, starved overnight then treated with U0126 #9903 (10 uM, 2 h; left) or PDBu (Phorbol 12,13-Dibutyrate) #12808 (100 nM, 15 m; right) using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb #4370 (green) and β-Actin (8H10D10) Mouse mAb #3700 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

Angiogenesis Antibody Sampler Kit: Image 19

Western blot analysis of extracts from HUVE cells, untreated or stimulated with VEGF (50 ng/ml for 2 minutes), using Phospho-VEGF Receptor 2 (Tyr1175) (19A10) Rabbit mAb (upper) and VEGF Receptor 2 (55B11) Rabbit mAb #2479 (lower).

Angiogenesis Antibody Sampler Kit: Image 20

Immunohistochemical analysis of paraffin-embedded human breast carcinoma comparing SignalStain® Antibody Diluent #8112 (left) to TBST/5% normal goat serum (right) using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060.

Angiogenesis Antibody Sampler Kit: Image 21

Immunohistochemical analysis of paraffin-embedded mouse colon using Phospho-p38 MAPK (Thr180/Tyr182) (D3F9) XP® Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 22

Immunohistochemical analysis of paraffin-embedded human colon (normal adjacent to tumor) using Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb in the presence of control peptide (left) or antigen-specific peptide (right).

Angiogenesis Antibody Sampler Kit: Image 23

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 24

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 25

Immunohistochemical analysis of paraffin-embedded 293T cell pellets, untreated (left) or UV-treated (right), using Phospho-p38 MAPK (Thr180/Tyr182) (D3F9) XP® Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 26

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 27

Immunohistochemical analysis of paraffin-embedded human lung carcinoma, untreated (left) or λ phosphatase-treated (right), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 28

Immunohistochemical analysis using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb on SignalSlide® Phospho-Akt (Ser473) IHC Controls #8101 (paraffin-embedded LNCaP cells, untreated (left) or LY294002-treated (right)).

Angiogenesis Antibody Sampler Kit: Image 29

Western blot analysis of extracts from COS and 293 cells, untreated or UV-treated, using Phospho-p38 MAPK (Thr180/Tyr182) (D3F9) XP® Rabbit mAb (upper) or p38 MAPK Antibody #9212 (lower).

Angiogenesis Antibody Sampler Kit: Image 30

Immunohistochemical analysis of SignalSlide® Phospho-EGF Receptor IHC Controls #8102 [paraffin-embedded KYSE450 cell pellets untreated (left) or EGF-treated (right)] using Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 31

Immunohistochemical analysis using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb on SignalSlide™ Phospho-p44/42 MAPK (Thr202/Tyr204) IHC Controls #8103 (paraffin-embedded NIH/3T3 cells, treated with U0126 #9903 (left) or TPA #4174 (right).

Angiogenesis Antibody Sampler Kit: Image 32

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 33

Immunoprecipitation (IP)/Western blot analysis of extracts from serum-starved HeLa cells, untreated (-) or treated (+) with TPA #4174 (100 nM, 15 min) prior to lysis in SDS (lanes 1 and 2) or IP lysis buffer (lane 3, TPA-treated only). IP Lysates were then subjected to immunoprecipitation with Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb (lane 4), PLCγ1 (D9H10) XP® Rabbit mAb #5690 (lane 5), or Normal Rabbit IgG #2729 (lane 6). The western blot was probed using Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb. Lane 3 represents 10% input.

Angiogenesis Antibody Sampler Kit: Image 34

Western blot analysis of extracts from COS cells, untreated or treated with either U0126 #9903 (10 µM for 1h) or TPA #4174 (200 nM for 10 m), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb #4370 and p44/42 MAPK (Erk1/2) (3A7) Mouse mAb #9107.

Angiogenesis Antibody Sampler Kit: Image 35

Immunohistochemical analysis of paraffin-embedded PTEN heterozygous mutant mouse endometrium using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb. (Tissue section courtesy of Dr. Sabina Signoretti, Brigham and Women's Hospital, Harvard Medical School, Boston, MA.)

Angiogenesis Antibody Sampler Kit: Image 36

Western blot analysis of extracts from serum-starved A-431 and A549 cells, untreated (-) or treated (+) with hEGF #8916 (100 ng/mL, 15 min) or serum-starved NIH/3T3 cells, untreated (-) or treated (+) with hPDGF-BB #8912 (50 ng/mL, 15 min), using Phospho-PLCγ1 (Ser1248) (D25A9) Rabbit mAb (upper) or PLCγ1 (D9H10) XP® Rabbit mAb #5690 (lower).

Angiogenesis Antibody Sampler Kit: Image 37

Western blot analysis of extracts from 293, NIH/3T3 and C6 cells, treated with λ phosphatase or TPA #4174 as indicated, using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb (upper), or p44/42 MAPK (Erk1/2) (137F5) Rabbit mAb #4695 (lower).

Angiogenesis Antibody Sampler Kit: Image 38

Immunohistochemical analysis of paraffin-embedded U-87MG xenograft, untreated (left) or lambda phosphatase-treated (right), using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb.

Angiogenesis Antibody Sampler Kit: Image 39

Immunoprecipitation of phospho-Akt (Ser473) from Jurkat extracts treated with Calyculin A #9902 (100nM, 30 min). Lane 1 is 10% input, lane 2 is Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb, and lane 3 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900. Western blot analysis was performed with Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb. Anti-rabbit IgG, HRP-linked Antibody #7074 was used as a secondary antibody.

Angiogenesis Antibody Sampler Kit: Image 40

Western blot analysis of extracts from PC-3 cells, untreated or LY294002/wortmannin-treated, and NIH/3T3 cells, serum-starved or PDGF-treated, using Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb (upper) or Akt (pan) (C67E7) Rabbit mAb #4691 (lower).

To Purchase # 8696T
Product # Size Price Inventory
8696T
1 Kit  (7 x 20 µl)

Product Description

The Angiogenesis Antibody Sampler Kit provides an economical means to investigate the angiogenic pathway downstream of VEGFR2. The kit contains enough primary antibody to perform two western blots per primary antibody.

Specificity / Sensitivity

Each antibody in the Angiogenesis Antibody Sampler Kit recognizes the phosphorylated form of its specific target. Phospho-FAK (Tyr397) (D20B1) Rabbit mAb may cross-react with overexpressed tyrosine phosphorylated proteins such as EGFR. Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb may cross-react with other Src family members or overexpressed phosphorylated RTKs. Phospho-VEGFR2 (Tyr1175) (19A10) Rabbit mAb may cross-react with VEGFR1.

Source / Purification

Rabbit monoclonal antibodies are produced by immunizing animals with synthetic phosphopeptides corresponding to residues surrounding Ser473 of human Akt protein, Tyr397 of human FAK protein, Thr180/Tyr182 of human p38 MAPK protein, Thr202/Tyr204 of human p44 MAPK protein, Ser1248 of human PLCγ1 protein, Tyr416 of human Src protein, and Tyr1175 of human VEGFR2 protein.

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). Signaling from VEGFR2 is necessary for angiogenesis in vivo (2-4). 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 (5). Phosphorylation of VEGFR2 at Tyr1212 provides a docking site for GRB2 binding and phosphorylation at Tyr1175 binds the p85 subunit of PI3 kinase and PLCγ (1,5,6). Activation of VEGFR2 during angiogenesis leads to signaling through multiple downstream kinase pathways including Akt, Src, FAK, p38, and Erk1/2 (2,7).

  1. Meyer, M. et al. (1999) EMBO J 18, 363-74.
  2. Karkkainen, M.J. and Petrova, T.V. (2000) Oncogene 19, 5598-605.
  3. Rahimi, N. et al. (2000) J Biol Chem 275, 16986-92.
  4. Claesson-Welsh, L. (2003) Biochem Soc Trans 31, 20-4.
  5. Holmqvist, K. et al. (2004) J Biol Chem 279, 22267-75.
  6. Takahashi, T. et al. (2001) EMBO J 20, 2768-78.
  7. Le Boeuf F et al. (2004) J Biol Chem 279, 39175–85

Pathways & Proteins

Explore pathways + proteins related to this product.

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