Receptor Tyrosine Kinase Antibody Sampler Kit #42344

Kit Usage Information

Protocols

• 3169: Western Blotting, Immunoprecipitation (Magnetic), Immunohistochemistry (Paraffin), Immunofluorescence
• 3174: Western Blotting, Immunoprecipitation (Agarose), Immunohistochemistry (Paraffin), Immunofluorescence, Flow
• 3462: Western Blotting, Immunoprecipitation (Agarose)
• 4267: Western Blotting, Immunoprecipitation (Agarose), Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin), Immunofluorescence, Flow
• 4290: Western Blotting, Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin)
• 7074: Western Blotting
• 8198: Western Blotting, Immunoprecipitation (Agarose), Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin), Immunofluorescence, Flow
• 8954: Western Blotting, Immunoprecipitation (Magnetic), Immunofluorescence, Flow
• 9740: Western Blotting, Immunoprecipitation (Magnetic), Immunohistochemistry (Paraffin), Immunofluorescence, Flow

Product Description

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.

Specificity / Sensitivity

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.

Source / Purification

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.

Background

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).

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40. Kwon, Y.K. et al. (1997) J Neurosci 17, 8293-9.

Pathways & Proteins

Explore pathways + proteins related to this product.

Select Your Pathway/Protein Angiogenesis ESC Pluripotency and Differentiation ErbB/HER Signaling IL6 Signaling Phospholipase Signaling Tyrosine Kinases & Substrates Protein: P00533 Protein: P04626 Protein: P08581 Protein: P09619 Protein: P11362 Protein: P16234 Protein: P36888

Database Links

UniProt ID: P16234 , P00533 , P08581 , P11362 , P04626 , P09619 , P36888

Entrez-Gene Id: 5156 , 1956 , 4233 , 2260 , 2064 , 5159 , 2322

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