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Human Phospho-EGFR DuoSet IC ELISA
Catalog # DYC1095B-2 | R&D Systems, Inc. a Bio-Techne Brand
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Key Product Details
| Assay Type: | Solid Phase Sandwich ELISA |
| Sample Types: | Cell Lysates |
| Human Phospho-EGFR DuoSet IC ELISA Features: |
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Figure 1: The Human Phospho-EGF R DuoSet IC ELISA is more sensitive than immunoprecipitation (IP)-Western blot analysis The human epidermoid carcinoma cell line, A431, was treated with 25 ng/mL recombinant human EGF (R&D Systems Catalog #236-EG) for 5 minutes to induce tyrosine phosphorylation of EGF R. Lysates were serially diluted and analyzed by (A) IP-Western blot and (B) this DuoSet IC ELISA. IPs were done using an anti-EGF R monoclonal antibody and anti-mouse IgG agarose. Immunoblots were incubated with an HRP-conjugated anti-phospho-tyrosine monoclonal antibody (R&D Systems, Catalog #HAM1676) to detect phosphor-EGF R. Bands were visualized by chemiluminescent detection. Human Phospho-EGF R can be detected in this DuoSet IC ELISA by using approximately 2-4 times less lysate than is needed for a conventional IP-Western blot. |
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Figure 2: The Human Phospho-EGF R DuoSet IC ELISA detects ligand-induced EGF R tyrosine phosphorylation A431 cells were untreated or treated with 25 ng/mL recombinant human EGF for 5 minutes. ELISA and IP-Western blot (inset) analyses were done using 25 and 50 μg of lysate, respectively. IP-Western blots for phospho-EGF R (p-EGF R) were done as described in Figure 1. Blots were stripped and total EGF R was detected using a biotinylated anti-EGF R polyclonal antibody (R&D Systems, Catalog #AF231). |
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Figure 3. The quantification of Human Phospho-EGF R DuoSet IC ELISA is demonstrated by using cells pretreated with the inhibitor PD168393 A431 cells were incubated with no additions or treated with 25 ng/mL recombinant human EGF for 5 minutes, after either being treated with or without 1 nM, 10 nM, or 100 nM PD168393. ELISA and IP-Western blot (inset) analyses were done using 25 and 50 μg of lysate, respectively. IP-Western blots for phospho-EGF R (p-EGF R) were done as described in Figure 1. Total EGF R blots were done as described in Figure 2. The DuoSet IC ELISA results correlate well with the relative amount of phosphorylated EGF R detected by IP-Western blot. |
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Figure 4. The specificity of Human Phospho-EGF R DuoSet IC ELISA is confirmed by receptor competition A431 cells were treated with 25 ng/mL recombinant human EGF for 5 minutes. The indicated amounts of recombinant extracellular domains of human EGF R (R&D Systems, Catalog #1095-ER), human ErbB2/Fc Chimera (R&D Systems, Catalog #1129-ER), human ErbB3/Fc Chimera (R&D Systems, Catalog #348-RB), or human ErbB4/Fc Chimera (R&D Systems, Catalog #1131-ER) were added to 25 μg of lysate and analyzed using this DuoSet IC ELISA. Competition was observed only with recombinant human EGF R. |
Citations for Human Phospho-EGFR DuoSet IC ELISA (14)
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Citations are publications that use Bio-Techne products. Selected citations for Human Phospho-EGFR DuoSet IC ELISA include:
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Species: Human
Sample Types: Tissue Homogenates
N Floc'h et al. (2018-02-26), Anti-tumor activity of osimertinib, an irreversible mutant-selective EGFR tyrosine kinase inhibitor, in NSCLC harboring EGFR Exon 20 Insertions Mol. Cancer Ther., 2018-00(0).
PMID: 29483211 -
Species: Human
Sample Types: Cell Culture Supernates
M Su et al. (2017-11-23), Andrographolide enhanced 5-fluorouracil-induced antitumor effect in colorectal cancer via inhibition of c-MET pathway Drug Des Devel Ther, 2017-111(0):3333-3341.
PMID: 29200829 -
Species: Human
Sample Types: Cell Lysates
Adam Jedlinski (2017-01-28), Cetuximab sensitivity of head and neck squamous cell carcinoma xenografts is associated with treatment-induced reduction of EGFR, pEGFR, and pSrc J. Oral Pathol. Med, 2017-00(0).
PMID: 28036101 -
Applications: ChIP(2016-09-01), Stand-Sit Microchip for High-Throughput, Multiplexed Analysis of Single Cancer Cells Sci Rep, 2016-06(0):32505.
PMID: 27581736 -
Species: Human
Sample Types: Cell Lysates
Ekerljung L et al. (2012-11-14), The HER2-binding affibody molecule (Z(HER2ratio342))(2) increases radiosensitivity in SKBR-3 cells. PLoS ONE, 2012-17(11):e49579.
PMID: 23166716 -
Species: Human
Sample Types: Cell Lysates
Brave SR et al. (2011-04-29), Vandetanib inhibits both VEGFR-2 and EGFR signalling at clinically relevant drug levels in preclinical models of human cancer. Int. J. Oncol., 2011-039(1):271-8.
PMID: 21537841 -
Species: Human
Sample Types: Cell Lysates
Giorgio C et al. (2011-03-30), Lithocholic acid is an Eph-ephrin ligand interfering with Eph-kinase activation. PLoS ONE, 2011-06(3):e18128.
PMID: 21479221 -
Species: Human
Sample Types: Cell Lysates
Kim S et al. (2011-02-07), CCL20/CCR6 feedback exaggerates epidermal growth factor receptor-dependent MUC5AC mucin production in human airway epithelial (NCI-H292) cells. J. Immunol., 2011-0186(6):3392-400.
PMID: 21300824 -
Species: Human
Sample Types: Cell Lysates
O'Brien NA et al. (2010-05-25), Activated phosphoinositide 3-kinase/AKT signaling confers resistance to trastuzumab but not lapatinib. Mol. Cancer Ther., 2010-09(6):1489-502.
PMID: 20501798 -
Species: Human
Sample Types: Cell Lysates
Hickinson DM et al. (2010-02-09), AZD8931, an equipotent, reversible inhibitor of signaling by epidermal growth factor receptor, ERBB2 (HER2), and ERBB3: a unique agent for simultaneous ERBB receptor blockade in cancer. Clin. Cancer Res., 2010-016(4):1159-69.
PMID: 20145185 -
Species: Human
Sample Types: Cell Lysates
Kim S et al. (2009-05-08), Fibrinogen binding to ICAM-1 promotes EGFR-dependent mucin production in human airway epithelial cells. Am. J. Physiol. Lung Cell Mol. Physiol., 2009-0297(1):L174-83.
PMID: 19429776 -
Species: Human
Sample Types: Cell Lysates
Shankaran H et al. (2008-04-18), Quantifying the effects of co-expressing EGFR and HER2 on HER activation and trafficking. Biochem. Biophys. Res. Commun., 2008-0371(2):220-4.
PMID: 18424261 -
Species: Human
Sample Types: Cell Lysates
Liu M et al. (2007-06-28), EGFR signaling is required for TGF-beta 1 mediated COX-2 induction in human bronchial epithelial cells. Am. J. Respir. Cell Mol. Biol., 2007-037(5):578-88.
PMID: 17600311 -
Species: Human
Sample Types: Cell Lysates
Sakai K et al. (2006-08-01), In-frame deletion in the EGF receptor alters kinase inhibition by gefitinib. Biochem. J., 2006-0397(3):537-43.
PMID: 16623663
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FAQs for Human Phospho-EGFR DuoSet IC ELISA
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Which phosphorylated sites are recognized in this assay?
This assay utilizes an anti-phosphorylated tyrosine monoclonal detection antibody, and it recognizes all phosphorylated tyrosines.
