Human EGFR (pY1068) + total EGFR ELISA Kit (ab126439)
- Datasheet
- References
- Protocols
Overview
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Product name
Human EGFR (pY1068) + total EGFR ELISA Kit
See all EGFR kits -
Detection method
Colorimetric -
Sample type
Cell Lysate -
Assay type
Semi-quantitative -
Assay time
5h 00m -
Assay duration
Multiple steps standard assay -
Species reactivity
Reacts with: Human -
Product overview
ab126439 is a very rapid, convenient and sensitive assay kit that can monitor the activation or function of important biological pathways in cell lysates. By determining phosphorylated EGFR protein in your experimental model system, you can verify pathway activation in your cell lysates. You can simultaneously measure numerous different cell lysates without spending excess time and effort in performing a Western Blot analysis.
This Sandwich ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of human phospho-EGFR (Tyr 1068) and total EGFR (help normalize the results of phospho-EGFR from different cell lysate being compared). An anti-EGFR antibody has been coated onto a 96-well plate. Samples are pipetted into the wells and phosphorylated and total EGFR present in a sample is bound to the wells by the immobilized antibody. The wells are washed and anti-phosphorylated EGFR (Tyr 1068) or anti-total-EGFR antibody is used to detect phosphorylated or non-phosphorylated EGFR. After washing away unbound antibody, HRP-conjugated anti-Rabbit IgG or HRP-Streptavidin is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of EGFR (Tyr 1068) or total EGFR bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.
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Tested applications
Suitable for: Sandwich ELISAmore details -
Platform
Microplate
Properties
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Storage instructions
Store at -20°C. Please refer to protocols. -
Components 1 x 96 tests 20X Wash Buffer Concentrate 1 x 25ml 2X Cell Lysis Buffer 1 x 5ml 5X Assay Diluent 1 x 15ml 600X HRP-Streptavidin Concentrate 1 x 200µl Anti-phospho EGFR (Y1068) 1 x 11µl Biotinylated Anti-EGFR: goat anti-human EGFR 1 vial EGFR Microplate (12 strips x 8 wells) coated with monoclonal anti-EGFR antibody 1 unit 500X HRP-conjugated anti-rabbit IgG 1 x 25µl Positive Control: lyophilized powder from A431 cell lysate 1 vial Stop Solution 1 x 8ml TMB One-Step Substrate Reagent 1 x 12ml -
Research areas
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Function
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. -
Tissue specificity
Ubiquitously expressed. Isoform 2 is also expressed in ovarian cancers. -
Involvement in disease
Lung cancer
Inflammatory skin and bowel disease, neonatal, 2 -
Sequence similarities
Belongs to the protein kinase superfamily. Tyr protein kinase family. EGF receptor subfamily.
Contains 1 protein kinase domain. -
Post-translational
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 localization
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). - Information by UniProt
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Alternative names
- Avian erythroblastic leukemia viral (v erb b) oncogene homolog
- Cell growth inhibiting protein 40
- Cell proliferation inducing protein 61
see all -
Database links
- Entrez Gene: 1956 Human
- Omim: 131550 Human
- SwissProt: P00533 Human
- Unigene: 488293 Human
- Unigene: 605083 Human
Applications
Our Abpromise guarantee covers the use of ab126439 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Abreviews | Notes |
|---|---|---|
| Sandwich ELISA | Use at an assay dependent concentration. |
Images
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Western blot - EGFR (pY1068) + total EGFR Human ELISA Kit (ab126439)A431 cells were treated or untreated with 100 ng/ml recombinant human EGF for 10 min. Cell lysates were analyzed by Western Blot. -
Sandwich ELISA - EGFR (pY1068) + total EGFR Human ELISA Kit (ab126439)A431 cells were treated or untreated with 100 ng/ml recombinant human EGF for 10 min. Cell lysates were analyzed using ab126439. -
Sandwich ELISA - EGFR (pY1068) + total EGFR Human ELISA Kit (ab126439)A431 cells were treated with recombinant human EGF at 37°C for 10 min. Solubilize cells at 4 x 107 cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed using ab126439.
Protocols
References
ab126439 has not yet been referenced specifically in any publications.
