Knockout Tested Rabbit Recombinant Monoclonal EGFR antibody. Carrier free. Suitable for IHC-P, IP, WB, ICC/IF, Flow Cyt (Intra) and reacts with Mouse, Human, Rat samples. Cited in 3 publications.
IgG
Rabbit
pH: 7.2 - 7.4
Constituents: PBS
Liquid
Monoclonal
IHC-P | IP | WB | ICC/IF | Flow Cyt (Intra) | |
---|---|---|---|---|---|
Human | Expected | Tested | Tested | Tested | Tested |
Mouse | Tested | Expected | Tested | Expected | Expected |
Rat | Expected | Expected | Tested | Expected | Expected |
Species | Dilution info | Notes |
---|---|---|
Species Mouse | Dilution info - | Notes Perform heat-mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
Species | Dilution info | Notes |
---|---|---|
Species Human, Rat | Dilution info Use at an assay dependent concentration. | Notes - |
Species | Dilution info | Notes |
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Species Human | Dilution info - | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Mouse, Rat | Dilution info Use at an assay dependent concentration. | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Human | Dilution info - | Notes This product yielded a strong signal in western blot using A431 (human squamous carcinoma) lysate which naturally overexpresses the EGFR protein. Western blot conditions may need to be optimised for cell lines and tissues that express lower levels of endogenous EGFR. |
Species Mouse | Dilution info - | Notes This product yielded a strong signal in western blot using A431 (human squamous carcinoma) lysate which naturally overexpresses the EGFR protein. Western blot conditions may need to be optimised for cell lines and tissues that express lower levels of endogenous EGFR. |
Species Rat | Dilution info - | Notes This product yielded a strong signal in western blot using A431 (human squamous carcinoma) lysate which naturally overexpresses the EGFR protein. Western blot conditions may need to be optimised for cell lines and tissues that express lower levels of endogenous EGFR. |
Species | Dilution info | Notes |
---|---|---|
Species Human | Dilution info - | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Mouse, Rat | Dilution info Use at an assay dependent concentration. | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Human | Dilution info - | Notes ab199376- Rabbit monoclonal IgG (Low endotoxin, Azide free), is suitable for use as an isotype control with this antibody. |
Species | Dilution info | Notes |
---|---|---|
Species Mouse, Rat | Dilution info Use at an assay dependent concentration. | Notes - |
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Receptor tyrosine kinase binding ligands of the EGF family and activating several signaling cascades to convert extracellular cues into appropriate cellular responses (PubMed:10805725, PubMed:27153536, PubMed:2790960, PubMed:35538033). Known ligands include EGF, TGFA/TGF-alpha, AREG, epigen/EPGN, BTC/betacellulin, epiregulin/EREG and HBEGF/heparin-binding EGF (PubMed:12297049, PubMed:15611079, PubMed:17909029, PubMed:20837704, PubMed:27153536, PubMed:2790960, PubMed:7679104, PubMed:8144591, PubMed:9419975). 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 (PubMed:27153536). May also activate the NF-kappa-B signaling cascade (PubMed:11116146). 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 (PubMed:11602604). Also phosphorylates MUC1 and increases its interaction with SRC and CTNNB1/beta-catenin (PubMed:11483589). Positively regulates cell migration via interaction with CCDC88A/GIV which retains EGFR at the cell membrane following ligand stimulation, promoting EGFR signaling which triggers cell migration (PubMed:20462955). Plays a role in enhancing learning and memory performance (By similarity). Plays a role in mammalian pain signaling (long-lasting hypersensitivity) (By similarity).Isoform 2 may act as an antagonist of EGF action.(Microbial infection) Acts as a receptor for hepatitis C virus (HCV) in hepatocytes and facilitates its cell entry. Mediates HCV entry by promoting the formation of the CD81-CLDN1 receptor complexes that are essential for HCV entry and by enhancing membrane fusion of cells expressing HCV envelope glycoproteins.
ERBB, ERBB1, HER1, EGFR, ERBB, ERBB1, HER1, Epidermal growth factor receptor, Proto-oncogene c-ErbB-1, Receptor tyrosine-protein kinase erbB-1
Knockout Tested Rabbit Recombinant Monoclonal EGFR antibody. Carrier free. Suitable for IHC-P, IP, WB, ICC/IF, Flow Cyt (Intra) and reacts with Mouse, Human, Rat samples. Cited in 3 publications.
IgG
Rabbit
pH: 7.2 - 7.4
Constituents: PBS
Liquid
Monoclonal
Yes
E235
Affinity purification Protein A
This antibody detects Epidermal growth factor receptor (EGFR). It does not cross react with other ERBB family members. This product yielded a strong signal in western blot using A431 (human squamous carcinoma) lysate which naturally overexpresses the EGFR protein. Western blot conditions may need to be optimised for cell lines and tissues that express lower levels of endogenous EGFR.
Blue Ice
+4°C
+4°C
Do Not Freeze
ab227459 is the carrier-free version of Anti-EGFR antibody [E235] ab32077.
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
This product is a recombinant monoclonal antibody, which offers several advantages including:
For more information, read more on recombinant antibodies.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with 1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
This supplementary information is collated from multiple sources and compiled automatically.
EGFR or Epidermal Growth Factor Receptor is a transmembrane glycoprotein that acts as a receptor for members of the epidermal growth factor family. Known alternatively as ErbB1 or HER1 this receptor has an approximate molecular weight of 170 kDa. EGFR is expressed in various cell types notably on epithelial cells and can influence multiple cellular processes through its kinase activity. It participates in the regulation of cell growth multiplication and survival by activating its kinase domain upon ligand binding.
The EGFR protein plays an important role in cellular communication and signaling processes. EGFR pairs with other receptor family members to form active dimers or even higher-order complexes which in turn initiate intracellular signaling cascades. Through these complexes EGFR influences many processes including cell differentiation and repair. This function of EGFR makes it an integral part of mammalian biology affecting how cells respond to their environment by mediating changes in gene expression.
EGFR is a central player in the MAPK and PI3K/Akt signaling pathways. Alongside other protein partners like KRAS and PI3 kinase it contributes to transmitting signals from the cell surface to the nucleus affecting gene transcription and cell behavior. These pathways are important for normal cell growth and division and aberrations in these pathways can lead to excessive or insufficient cell proliferation.
EGFR is pertinent to cancer biology including non-small cell lung cancer and glioblastoma where mutations or overexpression of the receptor frequently occur. It connects to proteins such as PTEN and BRAF which influence tumor progression and response to targeted therapies. EGFR's involvement in these disorders highlights its significance as a therapeutic target since it can be manipulated to alter disease progression.
We have tested this species and application combination and it works. It is covered by our product promise.
We have not tested this specific species and application combination in-house, but expect it will work. It is covered by our product promise.
This species and application combination has not been tested, but we predict it will work based on strong homology. However, this combination is not covered by our product promise.
We do not recommend this combination. It is not covered by our product promise.
We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.
In the unlikely event of one of our products not working as expected, you are covered by our product promise.
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False colour image of Western blot: Anti-EGFR antibody [E235] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-EGFR antibody [E235] ab32077 was shown to bind specifically to EGFR. A band was observed at 180 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in Egfr knockout cell line Human EGFR knockout HCT116 cell line ab281597 (knockout cell lysate ab282949). To generate this image, wild-type and Egfr knockout HCT 116 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/20000 dilution.
All lanes: Western blot - Anti-EGFR antibody [E235] (Anti-EGFR antibody [E235] ab32077) at 1/1000 dilution
Lane 1: Wild-type HCT 116 cell lysate at 20 µg
Lane 2: EGFR knockout HCT 116 cell lysate at 20 µg
Lane 3: HeLa cell lysate at 20 µg
Lane 4: Caco-2 cell lysate at 20 µg
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 134 kDa
Observed band size: 180 kDa
Purified Anti-EGFR antibody [E235] ab32077 at 1/50 dilution (2μg) immunoprecipitating EGFR in A431 whole cell lysate.
Lane 1 (input): A431 (Human epidermoid carcinoma epithelial cell) whole cell lysate 10μg
Lane 2 (+): Anti-EGFR antibody [E235] ab32077 + A431 whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of Anti-EGFR antibody [E235] ab32077 in A431 whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: 180 kDa
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-EGFR antibody [E235] ab32077).
All lanes: Immunoprecipitation - Anti-EGFR antibody [E235] (Anti-EGFR antibody [E235] ab32077)
Predicted band size: 134 kDa
Immunocytochemistry/Immunofluorescence analysis of A431 cells labelling EGFR with Anti-EGFR antibody [E235] ab32077 at 1/500. Cells were fixed with 100% methanol. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody.
Control: PBS only.
Nuclear counter stain: DAPI.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-EGFR antibody [E235] ab32077).
Overlay histogram showing A431 cells stained with Anti-EGFR antibody [E235] ab32077 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (Anti-EGFR antibody [E235] ab32077, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (Goat Anti-Rabbit IgG H&L (DyLight® 488) preadsorbed ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-EGFR antibody [E235] ab32077).
This IHC data was generated using the same anti-EGFR antibody clone, E235, in a different buffer formulation (cat# Anti-EGFR antibody [E235] ab32077).
IHC image of EGFR staining in a formalin fixed, paraffin embedded mouse normal skin tissue section, performed on a Leica Bond™ system using the standard protocol B. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with Anti-EGFR antibody [E235] ab32077 at 1/200 dilution for 15 mins at room temperature. A goat anti-rabbit biotinylated secondary antibody was used to detect the primary, and visualized using an HRP conjugated ABC system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Western blot: Anti-EGFR antibody [E235] (Anti-EGFR antibody [E235] ab32077) staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-EGFR antibody [E235] ab32077 was shown to bind specifically to EGFR. A band was observed at 160 kDa in wild-type cell lysates with no signal observed at this size in EGFR knockout cell lines. To generate this image, wild-type and EGFR knockout A549 (Human EGFR knockout A549 cell line ab286394) and HeLa (Human EGFR knockout HeLa cell line ab255385) cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1% Tween ® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-EGFR antibody [E235] ab32077).
All lanes: Western blot - Anti-EGFR antibody [E235] (Anti-EGFR antibody [E235] ab32077) at 1/1000 dilution
Lane 1: Wild-type A549 cell lysate at 20 µg
Lane 2: EGFR knockout A549 cell lysate at 20 µg
Lane 3: Wild-type HeLa cell lysate at 20 µg
Lane 4: EGFR knockout HeLa cell lysate at 20 µg
Performed under reducing conditions.
Observed band size: 160 kDa
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