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42344
Receptor Tyrosine Kinase Antibody Sampler Kit
Primary Antibodies

Receptor Tyrosine Kinase Antibody Sampler Kit #42344

Western Blotting Image 1

Western blot analysis of extracts from A-431 cells, untreated (-) or treated with Human Epidermal Growth Factor (hEGF) #8916 (100 ng/ml, 5 min; +), using Phospho-Tyrosine (P-Tyr-1000) MultiMab™ Rabbit mAb mix. Western blot image was obtained using the Odyssey® Infrared Imaging System (LI-COR® Biotechnology).

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Western Blotting Image 2

Western blot analysis of extracts from HT-29 (Met+), SK-BR-3 (Met-), and T-47D (Met-) cells using Met (D1C2) XP® Rabbit mAb (upper) or β-Actin Antibody #4967 (lower).

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IHC-Leica® Bond™ Image 3

Immunohistochemical analysis of paraffin-embedded human colon adenocarcinoma using Met (D1C2) XP® Rabbit mAb performed on the Leica® Bond™ Rx.

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Western Blotting Image 4

Western blot analysis of extracts from A-431, BxPC3 and HeLa cells using EGF Receptor (D38B1) XP® Rabbit mAb.

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Western Blotting Image 5

Western blot analysis of extracts from NIH/3T3 and human skeletal muscle cells (SKMC), untreated or treated with PDGF-BB, using PDGF Receptor α (D1E1E) XP® Rabbit mAb.

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Western Blotting Image 6

Western blot analysis of extracts from various cell lines, using PDGF Receptor β (28E1) Rabbit mAb.

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Western Blotting Image 7

Western blot analysis of extracts from A-204 (FGFR1 positive), KG-1a (FGFR1 oncogenic partner-FGFR1 fusion), A172 (FGFR1 low), and HT-29 (FGFR1 negative) cells using FGF Receptor 1 (D8E4) XP® Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).

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Flow Cytometry Image 8

Flow cytometric analysis of A-204 cells using FGF Receptor 1 (D8E4) XP® Rabbit mAb (blue) compared to concentration matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red).

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Western Blotting Image 9

Western blot analysis of extracts from Baf3/FLT3 transfected cells and SEM leukemia cells, using FLT3 (8F2) Rabbit mAb.

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Western Blotting Image 10

Western blot analysis of extracts from SK-BR-3 and MCF7 cells using HER2/ErbB2 (D8F12) XP® Rabbit mAb.

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Western Blotting Image 11

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.

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IP Image 12

Immunoprecipitation of phospho-tyrosine proteins from A-431 cell extracts, untreated (-) or treated with Human Epidermal Growth Factor (hEGF) #8916 (100 ng/ml, 5 min; +) (lanes 3 and 4), using Phospho-Tyrosine (P-Tyr-1000) MultiMab™ Rabbit mAb mix. Western blot analysis was performed using the same antibody. Lanes 1 and 2 are 10% input.

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IHC-P (paraffin) Image 13

Immunohistochemical analysis of paraffin-embedded human metastatic lung carcinoma using Met (D1C2) XP® Rabbit mAb.

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IHC-P (paraffin) Image 14

Immunohistochemical analysis of paraffin-embedded MDA-MB-468 (amplified EGFR, left), HT-29 (low EGFR, middle) and CAMA-1 (EGFR negative, right) cells using EGF Receptor (D38B1) XP® Rabbit mAb.

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IHC-P (paraffin) Image 15

Immunohistochemical analysis of paraffin-embedded human glioblastoma using PDGR Receptor α (D1E1E) XP® Rabbit mAb.

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IHC-P (paraffin) Image 16

Immunohistochemical analysis of paraffin-embedded human colon carcinoma using PDGF Receptor β (28E1) Rabbit mAb.

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IHC-P (paraffin) Image 17

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using FGF Receptor 1 (D8E4) XP® Rabbit mAb.

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IHC-P (paraffin) Image 18

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using HER2/ErbB2 (D8F12) XP® Rabbit mAb.

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Flow Cytometry Image 19

Flow cytometric analysis of K-562 cells, untreated (green) or Gleevec®-treated (blue), using Phospho-Tyrosine (P-Tyr-1000) MultiMab™ Rabbit mAb mix.

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IHC-P (paraffin) Image 20

Immunohistochemical analysis of paraffin-embedded human hepatocellular carcinoma using Met (D1C2) XP® Rabbit mAb.

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IHC-P (paraffin) Image 21

Immunohistochemical analysis of paraffin-embedded human hepatocellular carcinoma using EGF Receptor (D38B1) XP® Rabbit mAb.

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IHC-P (paraffin) Image 22

Immunohistochemical analysis of paraffin-embedded human colon using PDGR Receptor α (D1E1E) XP® Rabbit mAb.

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IHC-P (paraffin) Image 23

Immunohistochemical analysis of paraffin-embedded human glioblastoma using PDGF Receptor β (28E1) Rabbit mAb.

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IHC-P (paraffin) Image 24

Immunohistochemical analysis of paraffin-embedded human kidney using FGF Receptor 1 (D8E4) XP® Rabbit mAb.

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IHC-P (paraffin) Image 25

Immunohistochemical analysis of paraffin-embedded SK-BR-3 (Her2 high, left) and MCF7 cell pellets (Her2 low, right) using HER2/ErbB2 (D8F12) XP® Rabbit mAb.

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IF-IC Image 26

Confocal immunofluorescent analysis of C2C12 cells, serum-starved (left), treated with H2O2 (2 mM, 10 min; middle), or treated with H2O2 followed by λ phosphatase (right), using Phospho-Tyrosine (P-Tyr-1000) MultiMab™ Rabbit mAb mix (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IHC-P (paraffin) Image 27

Immunohistochemical analysis of paraffin-embedded human papillary renal cell carcinoma using Met (D1C2) XP® Rabbit mAb.

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IHC-P (paraffin) Image 28

Immunohistochemical analysis of paraffin-embedded human placenta using EGF Receptor (D38B1) XP® Rabbit mAb.

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IHC-P (paraffin) Image 29

Immunohistochemical analysis of paraffin-embedded U-118 MG xenograft using PDGF Receptor α (D1E1E) XP® Rabbit mAb in the presence of control peptide (left) or antigen specific peptide (right).

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IHC-P (paraffin) Image 30

Immunohistochemical analysis of paraffin-embedded U-87MG cells, showing membrane localization, using PDGF Receptor β (28E1) Rabbit mAb.

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IHC-P (paraffin) Image 31

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using FGF Receptor 1 (D8E4) XP® Rabbit mAb.

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IHC-P (paraffin) Image 32

Immunohistochemical analysis of paraffin-embedded cell pellets, MKN-45 (left) and T-47D (right), using Met (D1C2) XP® Rabbit mAb.

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IHC-P (paraffin) Image 33

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using EGF Receptor (D38B1) XP® Rabbit mAb.

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IHC-P (paraffin) Image 34

Immunohistochemical analysis of paraffin-embedded HCC827 xenograft using PDGF Receptor α (D1E1E) XP® Rabbit mAb.

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IHC-F (frozen) Image 35

Immunohistochemical analysis of frozen U-87MG xenograft using PDGF Receptor beta (28E1) Rabbit mAb.

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IHC-F (frozen) Image 36

Immunohistochemical analysis of frozen MKN-45 xenograft using Met (D1C2) XP® Rabbit mAb.

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Flow Cytometry Image 37

Flow cytometric analysis of Jurkat cells (red) and Kyse70 cells (blue), using EGF Receptor (D38B1) XP® Rabbit mAb.

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Flow Cytometry Image 38

Flow cytometric analysis of U-87 MG cells (blue) and H1703 cells (green) using PDGF Receptor α (D1E1E) XP® Rabbit mAb.

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IF-IC Image 39

Confocal immunofluorescent analysis of NIH/3T3 cells, serum-starved (left) or PDGF-treated (right), using PDGF Receptor beta (28E1) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IF-IC Image 40

Confocal immunofluorescent analysis of A204 cells (positive, left), KG-1 cells (positive, middle) and A172 cells (weak expression, right) using FGF Receptor 1 (D8E4) XP® Rabbit mAb (green). Blue pseudocolor= DRAQ5® #4084 (fluorescent DNA dye).

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Flow Cytometry Image 41

Flow cytometric analysis of T-47D cells (blue) and HT-29 cells (green) using Met (D1C2) XP® Rabbit mAb.

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IF-IC Image 42

Confocal immunofluorescent analysis of A549 cells, untreated (left) or treated with human epidermal growth factor (right), using EGF Receptor (D38B1) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IF-IC Image 43

Confocal immunofluorescent analysis of A-204 (left) and U-87 MG cells (right) using PDGF Receptor α (D1E1E) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IF-IC Image 44

Confocal immunofluorescent analysis of HT-29 and T-47D cells using Met (D1C2) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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Product Includes Quantity Applications Reactivity MW(kDa) Isotype
Phospho-Tyrosine (P-Tyr-1000) MultiMab™ Rabbit mAb mix 8954 20 µl
  • WB
  • IP
  • IF
  • F
All N/A Rabbit IgG
Met (D1C2) XP® Rabbit mAb 8198 20 µl
  • WB
  • IP
  • IHC
  • IF
  • F
H 140, 170 Rabbit IgG
EGF Receptor (D38B1) XP® Rabbit mAb 4267 20 µl
  • WB
  • IP
  • IHC
  • IF
  • F
H M Mk 175 Rabbit IgG
PDGF Receptor α (D1E1E) XP® Rabbit mAb 3174 20 µl
  • WB
  • IP
  • IHC
  • IF
  • F
H M 190 Rabbit IgG
PDGF Receptor β (28E1) Rabbit mAb 3169 20 µl
  • WB
  • IP
  • IHC
  • IF
H M R 190 Rabbit IgG
FGF Receptor 1 (D8E4) XP® Rabbit mAb 9740 20 µl
  • WB
  • IP
  • IHC
  • IF
  • F
H M R Mk 92 , 120, 145 Rabbit IgG
FLT3 (8F2) Rabbit mAb 3462 20 µl
  • WB
  • IP
H M 130 nonglycosylated form;160 glycosylated mature form Rabbit IgG
HER2/ErbB2 (D8F12) XP® Rabbit mAb 4290 20 µl
  • WB
  • IHC
H M 185 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

The Receptor Tyrosine Kinase Antibody Sampler Kit provides the means to detect a broad range of common receptor tyrosine kinases, as well as total phospho-tyrosine activity. The kit provides enough antibody to perform two western blot experiments with each primary antibody.

Each of the antibodies in the Receptor Tyrosine Kinase Assay Kit recognizes endogenous levels of the specified protein.

Phospho-Tyrosine (P-Tyr-1000) MultiMab™ rabbit mAb recognizes a broad range of tyrosine-phosphorylated proteins and peptides. This antibody does not cross-react with proteins or peptides containing phospho-Ser or phospho-Thr residues.

EGF Receptor (D38B1) XP® Rabbit mAb does not cross-react with other proteins of the ErbB family. Species cross-reactivity for IHC-P and IF-IC is human only.

PDGF Receptor α (D1E1E) XP® Rabbit mAb may cross-react with PDGFRβ at overexpressed levels. Nuclear staining has been observed with this antibody in certain tissues. The specificity of this staining is unknown.

PDGF Receptor β (28E1) Rabbit mAb may cross-react with PDGF receptor α at overexpressed levels.

FGF Receptor 1 (D8E4) XP® Rabbit mAb may slightly cross-react with overexpressed FGF receptor family members.

FLT3 (8F2) Rabbit mAb does not cross-react with related proteins.

HER2/ErbB2 (D8F12) XP® Rabbit mAb may slightly cross-react with other overexpressed RTKs.

MultiMab™ rabbit monoclonal mix antibodies are prepared by combining individual rabbit monoclonal clones in optimized ratios for the approved applications. Each antibody in the mix is carefully selected based on motif recognition and performance in multiple assays. Each mix is engineered to yield the broadest possible coverage of the modification being studied while ensuring a high degree of specificity for the modification or motif. Total monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human Met, the carboxy terminus of human PDGFRα, Ser740 of human FLT3, the amino terminus of human HER2/ErbB2 protein, with a fusion protein containing the cytoplasmic domain of human EGF receptor, with a GST fusion protein containing a carboxy-terminal fragment of human PDGF receptor β, or with a recombinant protein specific to the carboxy terminus of human FGF 1 receptor protein.

Tyrosine phosphorylation plays a key role in cellular signaling (1). In cancer studies, unregulated tyrosine kinase activity can drive malignancy and tumor formation by generating inappropriate proliferation and survival signals (2). Antibodies specific for phospho-tyrosine have been invaluable reagents in these studies (3,4).

Met, a tyrosine kinase receptor for hepatocyte growth factor (HGF), is a heterodimer made of α- and β-subunits (5,6). The cytoplasmic region of the β-chain is essential for tyrosine kinase activity. Interaction of Met with HGF results in autophosphorylation at multiple tyrosines (Tyr1003, 1234/1235, 1349) which recruit downstream signaling components, including Gab1, c-Cbl, and PI3 kinase (7-9). Altered Met levels and/or tyrosine kinase activities are found in several types of tumors, including renal, colon, and breast (10,11).

The epidermal growth factor (EGF) receptor is a transmembrane tyrosine kinase that belongs to the HER/ErbB protein family. Ligand binding results in receptor dimerization, autophosphorylation, activation of downstream signaling, internalization, and lysosomal degradation (12,13). c-Src mediated phosphorylation of EGF receptor (EGFR) at Tyr845 provides a binding surface for substrate proteins (14-16). The SH2 domain of PLCγ binds at phospho-Tyr992, activating PLCγ-mediated downstream signaling (17). Adaptor protein c-Cbl binds at phospho-Tyr1045, leading to receptor ubiquitination and degradation (18,19). The GRB2 adaptor protein binds activated EGFR at phospho-Tyr1068 (20), while phospho-Tyr1148 and -Tyr1173 provide a docking site for the Shc scaffold protein, playing a role in MAP kinase signaling (13).

Platelet derived growth factor (PDGF) family proteins bind to two closely related receptor tyrosine kinases, PDGF receptor α (PDGFRα) and PDGF receptor β (PDGFRβ) (21). PDGFRα and PDGFRβ can each form heterodimers with EGFR, which is also activated by PDGF (22). Ligand binding induces receptor dimerization and autophosphorylation, followed by binding and activation of signal transduction molecules such as GRB2, Src, GAP, PI3 kinase, PLCγ, and NCK. Signaling pathways initiated by activated PDGF receptors lead to control of cell growth, actin reorganization, migration, and differentiation (23). Tyr751 and Tyr740 of PDGFRβ regulate binding and activation of PI3 kinase (24,25).

Fibroblast growth factors (FGFs) produce mitogenic and angiogenic effects in target cells by signaling through cell surface receptor tyrosine kinases, after ligand binding and dimerization (26,27). Tyr653 and Tyr654 are important for catalytic activity of activated FGFR and are essential for signaling (28). The other phosphorylated tyrosine residues (Tyr463, 583, 585, 730, and 766) may provide docking sites for downstream signaling components such as Crk and PLCγ (29,30).

FMS-related tyrosine kinase 3 (FLT3), a member of the type III receptor tyrosine kinase family, is expressed on early hematopoietic progenitor cells and supports growth and differentiation within the hematopoietic system (31,32). FLT3 is activated after binding with its ligand FL, which results in a cascade of tyrosine autophosphorylation and tyrosine phosphorylation of downstream targets (33). The p85 subunit of PI3 kinase, SHP2, GRB2 and Shc are associated with FLT3 after FL stimulation (34-36). Tyr589/591 may play an important role in regulation of FLT3 tyrosine kinase activity (37).

The ErbB2 (HER2) proto-oncogene encodes a transmembrane, receptor-like glycoprotein with tyrosine kinase activity (38). ErbB2 kinase activity can be activated in the absence of a ligand when overexpressed and through associations with other ErbB family members (39). Phosphorylation at Tyr877 may be involved in regulating ErbB2 activity. Autophosphorylation of ErbB2 at Tyr1248 and Tyr1221/1222 couples ErbB2 to the Ras-Raf-MAP kinase signal transduction pathway (38,40).

  1. Schlessinger, J. (2000) Cell 103, 211-25.
  2. Cooper, C.S. et al. (1984) Nature 311, 29-33.
  3. Hackel, P.O. et al. (1999) Curr Opin Cell Biol 11, 184-9.
  4. Deuel, T.F. et al. (1988) Biofactors 1, 213-7.
  5. Powers, C.J. et al. (2000) Endocr Relat Cancer 7, 165-97.
  6. Shurin, M.R. et al. (1998) Cytokine Growth Factor Rev 9, 37-48.
  7. Muthuswamy, S.K. et al. (1999) Mol Cell Biol 19, 6845-57.
  8. Blume-Jensen, P. and Hunter, T. (2001) Nature 411, 355-65.
  9. Bottaro, D.P. et al. (1991) Science 251, 802-4.
  10. Zwick, E. et al. (1999) Trends Pharmacol Sci 20, 408-12.
  11. Reilly, J.F. et al. (2000) J Biol Chem 275, 7771-8.
  12. Naoe, T. et al. (2001) Cancer Chemother Pharmacol 48 Suppl 1, S27-30.
  13. Qian, X. et al. (1994) Proc Natl Acad Sci U S A 91, 1500-4.
  14. Bardelli, A. et al. (1997) Oncogene 15, 3103-11.
  15. Cooper, J.A. and Howell, B. (1993) Cell 73, 1051-4.
  16. Betsholtz, C. et al. (2001) Bioessays 23, 494-507.
  17. Mohammadi, M. et al. (1996) Mol Cell Biol 16, 977-89.
  18. Namikawa, R. et al. (1996) Stem Cells 14, 388-95.
  19. Ward, S.G. et al. (1992) J Biol Chem 267, 23862-9.
  20. Taher, T.E. et al. (2002) J Immunol 169, 3793-800.
  21. Hubbard, S.R. et al. (1994) Nature 372, 746-54.
  22. Mohammadi, M. et al. (1991) Mol Cell Biol 11, 5068-78.
  23. Beslu, N. et al. (1996) J Biol Chem 271, 20075-81.
  24. Glenney, J.R. et al. (1988) J Immunol Methods 109, 277-85.
  25. Schaeper, U. et al. (2000) J Cell Biol 149, 1419-32.
  26. Biscardi, J.S. et al. (1999) J Biol Chem 274, 8335-43.
  27. Ostman, A. and Heldin, C.H. (2001) Adv Cancer Res 80, 1-38.
  28. Larsson, H. et al. (1999) J Biol Chem 274, 25726-34.
  29. Zhang, S. and Broxmeyer, H.E. (2000) Biochem Biophys Res Commun 277, 195-9.
  30. Kwon, Y.K. et al. (1997) J Neurosci 17, 8293-9.
  31. Eder, J.P. et al. (2009) Clin Cancer Res 15, 2207-14.
  32. Emlet, D.R. et al. (1997) J Biol Chem 272, 4079-86.
  33. Panayotou, G. et al. (1992) EMBO J 11, 4261-72.
  34. Zhang, S. et al. (1999) J Leukoc Biol 65, 372-80.
  35. Sattler, M. and Salgia, R. (2009) Update Cancer Ther 3, 109-118.
  36. Levkowitz, G. et al. (1999) Mol Cell 4, 1029-40.
  37. Mizuki, M. et al. (2000) Blood 96, 3907-14.
  38. Ettenberg, S.A. et al. (1999) Oncogene 18, 1855-66.
  39. Kashishian, A. et al. (1992) EMBO J 11, 1373-82.
  40. Rojas, M. et al. (1996) J Biol Chem 271, 27456-61.
Entrez-Gene Id
1956 , 2260 , 2322 , 2064 , 4233 , 5156 , 5159
Swiss-Prot Acc.
P00533 , P11362 , P36888 , P04626 , P08581 , P16234 , P09619
For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
MultiMab is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.

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