Human Phospho-HGFR/c-MET DuoSet IC ELISA

Catalog # Availability Size / Price Qty
DYC2480E
DYC2480-5
DYC2480-2
Ancillary Products Available
Figure 1.  The Human Phospho-HGF R DuoSet IC ELISA is more sensitive than immunoprecipitation (IP)-Western analysis
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Product Details
Citations (5)
FAQs
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Human Phospho-HGFR/c-MET DuoSet IC ELISA Summary

Assay Type
Solid Phase Sandwich ELISA
Format
96-well strip plate
Sample Volume Required
Cell lysates (100 µL)
Sufficient Materials
Kits available for two, five, or fifteen 96-well plates*
Specificity
Please see the product datasheet

* Provided that the recommended microplates, buffers, diluents, substrates and solutions are used, and the assay is run as summarized in the Assay Procedure provided.

This DuoSet IC ELISA contains the basic components required for the development of sandwich ELISAs to measure tyrosine-phosphorylated human HGF R / c-MET in cell lysates. An immobilized capture antibody specific for HGF R / c-MET binds both phosphorylated and unphosphorylated HGF R / c-MET. After washing away unbound material, a biotinylated detection antibody is used to detect only phosphorylated receptor, utilizing a standard HRP format.

Product Features

  • Optimized capture and detection antibody pairings and recommended concentrations save lengthy development time
  • Development protocols are provided to guide further assay optimization
  • Assay can be customized to your specific needs
  • Available in 2, 5, and 15-(96-well) plate pack sizes
  • Economical alternative to Western blot

Kit Content

  • Capture Antibody
  • Conjugated Detection Antibody
  • Calibrated Immunoassay Standard or Control

Other Reagents Required


PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2O4, pH 7.2 - 7.4, 0.2 µm filtered

Wash Buffer: (Catalog # WA126), or equivalent

Lysis Buffer*

IC Diluent*

Blocking Buffer*


Substrate Solution: 1:1 mixture of Color Reagent A (H2O2) and Color Reagent B (Tetramethylbenzidine) (Catalog # DY999)

Stop Solution: 2 N H2SO4 (Catalog # DY994)

Microplates: R&D Systems (Catalog # DY990), or equivalent

Plate Sealers: ELISA Plate Sealers (Catalog # DY992), or equivalent

*For the Lysis Buffer, IC Diluent, and Blocking BUffer recommended for a specific DuoSet ELISA Development Kit, please see the product

Scientific Data

Figure 1. The Human Phospho-HGF R DuoSet IC ELISA is more sensitive than immunoprecipitation (IP)-Western analysis The human epidermoid carcinoma cell line, A431, was treated with 100 ng/mL recombinant human HGF for five minutes to induce tyrosine phosphorylation of HGF R. Serial dilutions of lysates were analyzed by (A) IP-Western blot and (B) this ELISA. IPs were done using an anti-HGF R polyclonal antibody and Protein G agarose. Immunoblots were incubated with a biotinylated anti-phosphotyrosine monoclonal antibody (Catalog # BAM1676) to detect phospho-HGF R (p-HGF R). Bands were visualized with Streptavidin-HRP (Cat # DY998) followed by chemiluminescent detection using WesternGloTM Chemiluminescent Detection Substrate (Catalog # AR004).

Figure 2. The Human Phospho-HGF RDuoSet IC ELISA detects ligand-induced HGF R tyrosine phosphorylation A431 cells were untreated or treated with 100 ng/mL recombinant human HGF for five minutes. ELISA and IP-Western blot (inset) analyses were done using 50 μg and 500 μg of lysate, respectively. IP-Western blots for phospho-HGF R (p-HGF R) were done as described in Figure 1. Blots were stripped and total HGF R was detected using a biotinylated anti-HGF R polyclonal antibody (Catalog # BAF358).

Figure 3. The specificity of the Human Phospho-HGF RDuoSet IC ELISA is confirmed by receptor competition A431 cells were treated with 100 ng/mL recombinant human HGF for five minutes. The indicated amounts of recombinant extracellular domains of human HGF R (Catalog #358-MT), human EGF R (Catalog #1095-ER) or human IGF-I sR (Catalog #305-GR) were added to 50 μg lysate and analyzed using this ELISA. Competition was observed only with recombinant HGF R.

Product Datasheets

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Preparation and Storage

Shipping
The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: HGFR/c-MET

HGF receptor, a product of the proto-oncogene c-met, is a heterodimeric transmembrane glycoprotein that is a receptor-type tyrosine kinase. c-MET is synthesized as a single-chain precursor (pr170) which undergoes post-translational glycosylation and proteolytic cleavage to give rise to the heterodimeric mature form.

Long Name:
Hepatocyte Growth Factor Receptor
Entrez Gene IDs:
4233 (Human); 17295 (Mouse)
Alternate Names:
AUTS9; cMET; c-MET; EC 2.7.10; EC 2.7.10.1; hepatocyte growth factor receptor; HGF R; HGF receptor; HGF/SF receptor; HGFR; Met (c-Met); met proto-oncogene (hepatocyte growth factor receptor); met proto-oncogene tyrosine kinase; MET; oncogene MET; Proto-oncogene c-Met; RCCP2; Scatter factor receptor; SF receptor; Tyrosine-protein kinase Met

Citations for Human Phospho-HGFR/c-MET DuoSet IC ELISA

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

5 Citations: Showing 1 - 5
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  1. Andrographolide enhanced 5-fluorouracil-induced antitumor effect in colorectal cancer via inhibition of c-MET pathway
    Authors: M Su, B Qin, F Liu, Y Chen, R Zhang
    Drug Des Devel Ther, 2017;11(0):3333-3341.
    Species: Human
    Sample Types: Cell Culture Supernates
  2. Shedding of c-Met ectodomain correlates with c-Met expression in non-small cell lung cancer.
    Authors: Fu, Le, Guo, Wei, Liu, Bingshan, Sun, Linlin, Bi, Zhenghon, Zhu, Li, Wang, Xinyan, Liu, Bin, Xie, Qian, Li, Ke
    Biomarkers, 2013;18(2):126-35.
    Species: Human
    Sample Types: Cell Culture Supernates
  3. Imbalance in the pro-hepatocyte growth factor activation system in bleomycin-induced lung fibrosis in mice.
    Authors: Phin S, Marchand-Adam S, Fabre A, Marchal-Somme J, Bantsimba-Malanda C, Kataoka H, Soler P, Crestani B
    Am. J. Respir. Cell Mol. Biol., 2010;42(3):286-93.
    Species: Human
    Sample Types: Cell Lysates
  4. Epidermal growth factor receptor regulates MET levels and invasiveness through hypoxia-inducible factor-1alpha in non-small cell lung cancer cells.
    Authors: Xu L, Nilsson MB, Saintigny P, Cascone T, Herynk MH, Du Z, Nikolinakos PG, Yang Y, Prudkin L, Liu D, Lee JJ, Johnson FM, Wong KK, Girard L, Gazdar AF, Minna JD, Kurie JM, Wistuba II, Heymach JV
    Oncogene, 2010;29(18):2616-27.
    Species: Human
    Sample Types: Cell Lysates
  5. Use of IHC and newly designed matriptase inhibitors to elucidate the role of matriptase in pancreatic ductal adenocarcinoma.
    Authors: Uhland K, Siphos B, Arkona C, Schuster M, Petri B, Steinmetzer P, Mueller F, Schweinitz A, Steinmetzer T, Van De Locht A
    Int. J. Oncol., 2009;35(2):347-57.
    Species: Human
    Sample Types: Cell Lysates

FAQs

  1. Which phosphorylated sites are recognized by this assay?

    • This assay utilizes an anti-phosphorylated tyrosine monoclonal detection antibody, and it recognizes all phosphorylated tyrosine residues.

View all ELISA FAQs

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