Cell Signaling Technology

Product Pathways - Tyrosine Kinase/ Adaptors

EGF Receptor Antibody #2232

Applications Reactivity MW (kDa) Source
W IP IHC-P IF-IC IC F H M R 175 Rabbit

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IF-IC=Immunofluorescence (Immunocytochemistry)  IC=Immunocytochemistry  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat
Species enclosed in parentheses are predicted to react based on 100% sequence homology. Species cross-reactivity is determined by Western blot.

Specificity / Sensitivity

EGF Receptor Antibody detects endogenous levels of total EGF receptor protein. The antibody does not cross-react with other proteins of the ErbB family.

Source / Purification

Polyclonal antibodies are produced by immunizing rabbits with a synthetic peptide (KLH-coupled) corresponding to residues surrounding Tyr1068 of human EGF receptor. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines, using EGF Receptor Antibody. (*A431 cells were treated with 100 ng/ml EGF for 2 minutes.)

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma, showing membrane localization, using EGF Receptor Antibody.

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of A-431 cells, using EGF Receptor antibody (blue) compared to a nonspecific negative control antibody (red).


IF-IC

IF-IC

Confocal immunofluorescent analysis of HT-29 cells using EGF Receptor Antibody (green). Blue pseudocolor = DRAQ5™ (fluorescent DNA dye).

Background

The epidermal growth factor (EGF) receptor is a 170 kDa transmembrane tyrosine kinase that belongs to the HER/ErbB protein family. Ligand binding results in receptor dimerization, autophosphorylation, activation of downstream signaling and lysosomal degradation (1,2). Phosphorylation of EGF receptor (EGFR) at Tyr845 in the kinase domain is implicated in stabilizing the activation loop, maintaining the active state enzyme and providing a binding surface for substrate proteins (3,4). c-Src is involved in phosphorylation of EGFR at Tyr845 (5). The SH2 domain of PLCγ binds at phospho-Tyr992, resulting in activation of PLCγ-mediated downstream signaling (6). Phosphorylation of EGFR at Tyr1045 creates a major docking site for c-Cbl, an adaptor protein that leads to receptor ubiquitination and degradation following EGFR activation (7,8). The GRB2 adaptor protein binds activated EGFR at phospho-Tyr1068 (9). A pair of phosphorylated residues (Tyr1148 and Tyr1173) provides a docking site for the SHC scaffold protein, with both sites involved in MAP kinase signaling activation (2). Phosphorylation of EGFR at specific serine and threonine residues attenuates EGFR kinase activity. EGFR carboxy-terminal residues Ser1046 and Ser1047 are phosphorylated by CaM kinase II; mutations to either of these serines results in upregulated EGFR tyrosine autokinase activity (10).

  1. Hackel, P.O. et al. (1999) Curr. Opin. Cell Biol. 11, 184-189.
  2. Zwick, E. et al. (1999) Trends Pharmacol. Sci. 20, 408-412.
  3. Cooper, J.A. and Howell, B. (1993) Cell 73, 1051-1054.
  4. Hubbard, S.R. et al. (1994) Nature 372, 746-754.
  5. Biscardi, J.S. et al. (1999) J. Biol. Chem. 274, 8335-8343.
  6. Emlet, D.R. et al. (1997) J. Biol. Chem. 272, 4079-4086.
  7. Levkowitz, G. et al. (1999) Mol. Cell 4, 1029-1040.
  8. Ettenberg, S.A. et al. (1999) Oncogene 18, 1855-1866.
  9. Rojas, M. et al. (1996) J. Biol. Chem. 271, 27456-27461.
  10. Feinmesser, R.L. et al. (1999) J. Biol. Chem. 274, 16168-16173.

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