Anti-EGFR 抗体 [E114] (ab32562)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [E114] to EGFR
- Suitable for: IHC-P, WB, IP, ICC/IF, Flow Cyt (Intra)
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
製品の概要
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製品名
Anti-EGFR antibody [E114]
EGFR 一次抗体 製品一覧 -
製品の詳細
Rabbit monoclonal [E114] to EGFR -
由来種
Rabbit -
特異性
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.
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アプリケーション
適用あり: IHC-P, WB, IP, ICC/IF, Flow Cyt (Intra)more details -
種交差性
交差種: Mouse, Rat, Human -
免疫原
Synthetic peptide within Human EGFR (C terminal). The exact sequence is proprietary.
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エピトープ
ab32562 reacts with an epitope located in the C terminal region of EGF receptor. -
ポジティブ・コントロール
- WB: HeLa and A431 cell lysates. IHC-P: human cervical carcinoma, mouse skin, rat skin ICC/IF: A431 cells. IP: HeLa cell lysate.
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特記事項
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.
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
製品の特性
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製品の状態
Liquid -
保存方法
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle. -
バッファー
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA -
Concentration information loading...
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精製度
Protein A purified -
ポリ/モノ
モノクローナル -
クローン名
E114 -
アイソタイプ
IgG -
研究分野
関連製品
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Positive Controls
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Recombinant Protein
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Related Products
アプリケーション
The Abpromise guarantee
Abpromise保証は、 次のテスト済みアプリケーションにおけるab32562の使用に適用されます
アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。
アプリケーション | Abreviews | 特記事項 |
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IHC-P |
1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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WB |
1/1000 - 1/10000. Predicted molecular weight: 170 kDa.
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. |
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IP |
1/10 - 1/20.
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ICC/IF |
1/10 - 1/100.
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
特記事項 |
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IHC-P
1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. |
WB
1/1000 - 1/10000. Predicted molecular weight: 170 kDa. 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. |
IP
1/10 - 1/20. |
ICC/IF
1/10 - 1/100. |
Flow Cyt (Intra)
Use at an assay dependent concentration. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
ターゲット情報
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機能
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. -
組織特異性
Ubiquitously expressed. Isoform 2 is also expressed in ovarian cancers. -
関連疾患
Lung cancer
Inflammatory skin and bowel disease, neonatal, 2 -
配列類似性
Belongs to the protein kinase superfamily. Tyr protein kinase family. EGF receptor subfamily.
Contains 1 protein kinase domain. -
翻訳後修飾
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. -
細胞内局在
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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参照データベース
- Entrez Gene: 1956 Human
- Entrez Gene: 13649 Mouse
- Entrez Gene: 24329 Rat
- Omim: 131550 Human
- SwissProt: P00533 Human
- SwissProt: Q01279 Mouse
- Unigene: 488293 Human
- Unigene: 605083 Human
see all -
別名
- Avian erythroblastic leukemia viral (v erb b) oncogene homolog antibody
- Cell growth inhibiting protein 40 antibody
- Cell proliferation inducing protein 61 antibody
see all
画像
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervical carcinoma tissue labelling EGFR with purified ab32562 at a dilution of 1/100. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
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Anti-EGFR antibody [E114] (ab32562) at 1/10000 dilution + A431 at 20 µg
Performed under reducing conditions.
Predicted band size: 170 kDaThis blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour before being incubated with ab32562 overnight at 4°C in the presence of loading control ab18058 Mouse monoclonal [SPM227] to Vinculin diluted 1:10000). Antibody binding was detected using IR-labelled goat anti-Rabbit Ab at a 1:10,000 dilution for one hour at room temperature before imaging.
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.
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Immunocytochemistry/Immunofluorescence analysis of A431 cells labelling EGFR with purified ab32562 at 1/100. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain. ab7291, a mouse anti-tubulin (1/1000) and ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000) were also used.
Control 1: primary antibody (1/100) and secondary antibody, ab150120, an Alexa Fluor® 594-conjugated goat anti-mouse IgG (1/1000).
Control 2: ab7291 (1/1000) and secondary antibody, ab150077, an Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000).
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ab32562 (purified) at 1/20 immunoprecipitating EGFR in HeLa whole cell lysate.
Lane 1 (input): HeLa whole cell lysate (10µg)
Lane 2 (+): ab32562 + HeLa whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32562 in HeLa whole cell lysate.
For western blotting, ab131366 VeriBlot for IP (HRP) was used for detection (1/10000).
Blocking buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM /TBST.
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ab32562 staining EGFRin the human cell line A431 (human epidermoid carcinoma) by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde and the sample was incubated with the primary antibody at a dilution of 1/20. A goat anti rabbit IgG (Alexa Fluor® 488) at a dilution of 1/2000 was used as the secondary antibody.
Isoytype control: Rabbit monoclonal IgG (Black)
Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue)
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IHC image of EGFR staining in human nomal cervix formalin fixed paraffin embedded tissue section*, performed on a Leica Bond™ system using the standard protocol F. 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 ab32562, 1in50 dilution, for 15 mins at room temperature and detected using an HRP conjugated compact polymer 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.
*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre
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IHC image of EGFR staining in mouse skin formalin fixed paraffin embedded 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 ab32562, 1in50 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.
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IHC image of EGFR staining in rat skin formalin fixed paraffin embedded tissue section, performed on a Leica Bond™ system using the standard protocol F. 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 ab32562, 1in50 dilution, for 15 mins at room temperature and detected using an HRP conjugated compact polymer 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.
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Anti-EGFR antibody [E114] (ab32562) at 1/10000 dilution (purified) + A431 whole cell lysate at 10 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 170 kDa
Observed band size: 170 kDaBlocking and dilution buffer: 5% NFDM /TBST.
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Anti-EGFR antibody [E114] (ab32562) at 1/10000 dilution (purified) + HeLa whole cell lysate at 10 µg
Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 170 kDa
Observed band size: 170 kDaBlocking and dilution buffer: 5% NFDM /TBST.
プロトコール
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
データシートおよび資料
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SDS download
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Datasheet download
参考文献 (12)
ab32562 は 12 報の論文で使用されています。
- Cheng CC et al. Overexpression of N-acetylglucosaminyltransferase V promotes human parotid gland acinar cell immortalization via the epidermal receptor activation. J Cell Physiol 237:1780-1789 (2022). PubMed: 34806177
- Shin HY et al. Tumor Spheroids of an Aggressive Form of Central Neurocytoma Have Transit-Amplifying Progenitor Characteristics with Enhanced EGFR and Tumor Stem Cell Signaling. Exp Neurobiol 30:120-143 (2021). PubMed: 33972466
- Yen CH et al. Cell detachment ratio on pH-responsive chitosan: A useful biometric for prognostic judgment and drug efficacy assessment in oncology. Carbohydr Polym 261:117911 (2021). PubMed: 33766385
- Scott ML et al. Human Papillomavirus 16 E5 Inhibits Interferon Signaling and Supports Episomal Viral Maintenance. J Virol 94:N/A (2020). PubMed: 31666385
- Huang P et al. Circular RNA cSMARCA5 regulates the progression of cervical cancer by acting as a microRNA-432 sponge. Mol Med Rep 21:1217-1223 (2020). PubMed: 31922210