Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EP2255Y] to EGFR (phospho T654)
- Suitable for: WB, Dot blot
- Reacts with: Human
Product nameAnti-EGFR (phospho T654) antibody [EP2255Y]
See all EGFR primary antibodies
DescriptionRabbit monoclonal [EP2255Y] to EGFR (phospho T654)
SpecificityRecognises EGFR phosphorylated on Threonine 654 of the mature human isoform 1 (corresponding to T678 from the precursor form P00533-1/p170)
Tested Applications & Species
Application Species WBHuman
corresponding to Human EGFR (phospho T654).
- WB: EGF treated A431 whole cell lysate.
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.
Storage instructionsShipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid repeated freeze / thaw cycles.
Storage bufferpH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 0.05% BSA, 40% Glycerol (glycerin, glycerine), 59% PBS
Concentration information loading...
PurityProtein A purified
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab75986 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Tested applications are guaranteed to work and covered by our Abpromise guarantee.
Predicted to work for this combination of applications and species but not guaranteed.
Does not work for this combination of applications and species.
1/1000 - 1/50000. Predicted molecular weight: 134 kDa.
1/1000 - 1/50000. Predicted molecular weight: 134 kDa.
FunctionReceptor 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.
Tissue specificityUbiquitously expressed. Isoform 2 is also expressed in ovarian cancers.
Involvement in diseaseLung cancer
Inflammatory skin and bowel disease, neonatal, 2
Sequence similaritiesBelongs to the protein kinase superfamily. Tyr protein kinase family. EGF receptor subfamily.
Contains 1 protein kinase domain.
modificationsPhosphorylation 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.
Cellular localizationSecreted 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).
- Information by UniProt
- Avian erythroblastic leukemia viral (v erb b) oncogene homolog antibody
- Cell growth inhibiting protein 40 antibody
- Cell proliferation inducing protein 61 antibody
All lanes : Anti-EGFR (phospho T654) antibody [EP2255Y] (ab75986) at 1/1000 dilution (Purified)
Lane 1 : A431 (Human epidermoid carcinoma epithelial cell) whole cell lysate
Lane 2 : A431 (Human epidermoid carcinoma epithelial cell) treated with 100ng/ml EGF for 30 minutes whole cell lysate
Lane 3 : A431 (Human epidermoid carcinoma epithelial cell) treated with 100ng/ml EGF for 30 minutes whole cell lysate, then the membrane treated with Lambda Phosphotase for 1 hour
Lysates/proteins at 15 µg per lane.
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 134 kDa
Observed band size: 170 kDa why is the actual band size different from the predicted?
Blocking buffer: 5% NFDM/TBST
Primary antibody dilution: 1/1000
Secondary antibody: goat anti-rabbit IgG, (H+L), peroxidase conjugated
Secondary antibody dilution: 1/2500
Blocking & dilution buffer: 5% NFDM/TBST
Lane 1 sample: EGFR (pT654) phospho peptide
Lane 2 sample: EGFR non-phospho peptide
Exposure time: 3 minutes
ab75986 has not yet been referenced specifically in any publications.