The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration.
1/500. Detects a band of approximately 175 kDa (predicted molecular weight: 134 kDa).
1/100 - 1/250.
Is unsuitable for Flow Cyt,IHC-P or IP.
Receptor tyrosine kinase binding ligands of the EGF family and activating several signaling cascades to convert extracellular cues into appropriate cellular responses. Known ligands include EGF, TGFA/TGF-alpha, amphiregulin, epigen/EPGN, BTC/betacellulin, epiregulin/EREG and HBEGF/heparin-binding EGF. Ligand binding triggers receptor homo- and/or heterodimerization and autophosphorylation on key cytoplasmic residues. The phosphorylated receptor recruits adapter proteins like GRB2 which in turn activates complex downstream signaling cascades. Activates at least 4 major downstream signaling cascades including the RAS-RAF-MEK-ERK, PI3 kinase-AKT, PLCgamma-PKC and STATs modules. May also activate the NF-kappa-B signaling cascade. Also directly phosphorylates other proteins like RGS16, activating its GTPase activity and probably coupling the EGF receptor signaling to the G protein-coupled receptor signaling. Also phosphorylates MUC1 and increases its interaction with SRC and CTNNB1/beta-catenin. Isoform 2 may act as an antagonist of EGF action.
Ubiquitously expressed. Isoform 2 is also expressed in ovarian cancers.
Involvement in disease
Lung cancer Inflammatory skin and bowel disease, neonatal, 2
Belongs to the protein kinase superfamily. Tyr protein kinase family. EGF receptor subfamily. Contains 1 protein kinase domain.
Phosphorylation at Ser-695 is partial and occurs only if Thr-693 is phosphorylated. Phosphorylation at Thr-678 and Thr-693 by PRKD1 inhibits EGF-induced MAPK8/JNK1 activation. Dephosphorylation by PTPRJ prevents endocytosis and stabilizes the receptor at the plasma membrane. Autophosphorylation at Tyr-1197 is stimulated by methylation at Arg-1199 and enhances interaction with PTPN6. Autophosphorylation at Tyr-1092 and/or Tyr-1110 recruits STAT3. Dephosphorylated by PTPN1 and PTPN2. Monoubiquitinated and polyubiquitinated upon EGF stimulation; which does not affect tyrosine kinase activity or signaling capacity but may play a role in lysosomal targeting. Polyubiquitin linkage is mainly through 'Lys-63', but linkage through 'Lys-48', 'Lys-11' and 'Lys-29' also occurs. Deubiquitination by OTUD7B prevents degradation. Ubiquitinated by RNF115 and RNF126. Methylated. Methylation at Arg-1199 by PRMT5 stimulates phosphorylation at Tyr-1197.
Secreted and Cell membrane. Endoplasmic reticulum membrane. Golgi apparatus membrane. Nucleus membrane. Endosome. Endosome membrane. Nucleus. In response to EGF, translocated from the cell membrane to the nucleus via Golgi and ER. Endocytosed upon activation by ligand. Colocalized with GPER1 in the nucleus of estrogen agonist-induced cancer-associated fibroblasts (CAF).
Immunofluorescent analysis of 4% paraformaldehyde-fixed,0.1% Triton X-100 permeabilized A431 (Human epidermoid carcinoma cell line) cell lines labeling EGFR with ab97613 at 1/50 dilution,followed by Goat anti-Rabbit IgG H & L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution
Cells were counterstained with ab195889 (Anti-alpha Tubulin antibody [DM1A] ) - Microtubule Marker (Alexa Fluor® 594)
The nuclear counterstain is DAPI (blue).
The green staining on the membrane was increased in the EGF (100ng/ml, 10min) treated A431 cells when compared with A431 cells without treatment. After LP treatment, the green signaling was obviously decreased.
For the pan antibody, there was no great difference after EGF (100ng/ml, 10min) or EGF (100ng/ml, 10min) + LP treatment. The data showed mostly membranous staining.
Dot Blot - Anti-EGFR (phospho Y845) antibody [EPR2149Y] (ab97613)
Dot Blot analysis of Lane 1: EGFR (pY845) phospho peptide and Lane 2: EGFR non-phospho peptide labeling EGFR (phospho Y845) with ab97613 at 1/1000 dilution. 5% NFDM/TBST was used as the diluting and blocking buffer. ab97051 Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated was used as the secondary antibody at 1/100000 dilution. Exposure time: 3 minutes.
Western blot - Anti-EGFR (phospho Y845) antibody [EPR2149Y] (ab97613)
All lanes : Anti-EGFR (phospho Y845) antibody [EPR2149Y] (ab97613) at 1/500 dilution
Lane 1 : A431 cell lysates, untreated Lane 2 : A431 cell lysates, treated with EGF
Saghizadeh M et al. Adenovirus-driven overexpression of proteinases in organ-cultured normal human corneas leads to diabetic-like changes. Brain Res Bull81:262-72 (2010).
Read more (PubMed: 19828126) »
Saghizadeh M et al. Normalization of wound healing and diabetic markers in organ cultured human diabetic corneas by adenoviral delivery of c-Met gene. Invest Ophthalmol Vis Sci51:1970-80 (2010).
Read more (PubMed: 19933191) »