Mouse HGF DuoSet ELISA Summary
* 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 ELISA Development kit contains the basic components required for the development of sandwich ELISAs to measure natural and recombinant mouse HGF. The suggested diluent is suitable for the analysis of most cell culture supernate samples. Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet.
Product Features
- Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
- Development protocols are provided to guide further assay optimization
- Assay can be customized to your specific needs
- Economical alternative to complete kits
Kit Content
- Capture Antibody
- Detection Antibody
- Recombinant Standard
- Streptavidin conjugated to horseradish-peroxidase (Streptavidin-HRP)
Other Reagents Required
DuoSet Ancillary Reagent Kit 2 (5 plates): (Catalog # DY008) containing 96 well microplates, plate sealers, substrate solution, stop solution, plate coating buffer (PBS), wash buffer, and Reagent Diluent Concentrate 2.
Normal Goat Serum: (Catalog # DY005)
The components listed above may be purchased separately:
PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2PO4, pH 7.2 - 7.4, 0.2 µm filtered
Wash Buffer: (Catalog # WA126), or 0.05% Tween® 20 in PBS, pH 7.2-7.4
Reagent Diluent: (Catalog # DY995), or 1% BSA in PBS, pH 7.2-7.4, 0.2 µm filtered
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)
Plate Sealers: ELISA Plate Sealers (Catalog # DY992)
Normal Goat Serum: (Catalog # DY005)
Scientific Data
Product Datasheets
Preparation and Storage
Background: HGF
HGF (Hepatocyte Growth Factor, Scatter Factor) induces the proliferation and migration of epithelial cells, hepatocytes, chondrocytes, keratinocytes, melanocytes, endothelial cells, and many tumor cells. During organogenesis and tissue repair, HGF promotes epithelial/endothelial morphogenesis by inducing cell scattering and branching tubulogenesis. It also supports insulin production by pancreatic beta cells, neuronal survival, and immune tolerance. HGF is secreted as a propeptide that is activated by uPA or HGF Activator at sites of tissue damage. Its signaling through the receptor HGF R/c-MET is enhanced by its prior binding to heparan sulfate proteoglycans. The serum levels of HGF are elevated in a wide range of pathologies including liver damage, acute kidney failure, myocardial infarction, type 1 diabetes, obesity, and cancer, as well as in the synovial fluid of rheumatoid arthritis patients.
Assay Procedure
GENERAL ELISA PROTOCOL
Plate Preparation
- Dilute the Capture Antibody to the working concentration in PBS without carrier protein. Immediately coat a 96-well microplate with 100 μL per well of the diluted Capture Antibody. Seal the plate and incubate overnight at room temperature.
- Aspirate each well and wash with Wash Buffer, repeating the process two times for a total of three washes. Wash by filling each well with Wash Buffer (400 μL) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each step is essential for good performance. After the last wash, remove any remaining Wash Buffer by aspirating or by inverting the plate and blotting it against clean paper towels.
- Block plates by adding 300 μL Reagent Diluent to each well. Incubate at room temperature for a minimum of 1 hour.
- Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.
Assay Procedure
- Add 100 μL of sample or standards in Reagent Diluent, or an appropriate diluent, per well. Cover with an adhesive strip and incubate 2 hours at room temperature.
- Repeat the aspiration/wash as in step 2 of Plate Preparation.
- Add 100 μL of the Detection Antibody, diluted in Reagent Diluent with NGS, to each well. Cover with a new adhesive strip and incubate 2 hours at room temperature.
- Repeat the aspiration/wash as in step 2 of Plate Preparation.
- Add 100 μL of the working dilution of Streptavidin-HRP to each well. Cover the plate and incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
- Repeat the aspiration/wash as in step 2.
- Add 100 μL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
- Add 50 μL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
- Determine the optical density of each well immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate. Readings made directly at 450 nm without correction may be higher and less accurate.
Citations for Mouse HGF DuoSet 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.
11
Citations: Showing 1 - 10
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Opposing roles of hepatic stellate cell subpopulations in hepatocarcinogenesis
Authors: A Filliol, Y Saito, A Nair, DH Dapito, LX Yu, A Ravichandr, S Bhattachar, S Affo, N Fujiwara, H Su, Q Sun, TM Savage, JR Wilson-Kan, JM Caviglia, L Chin, D Chen, X Wang, S Caruso, JK Kang, AD Amin, S Wallace, R Dobie, D Yin, OM Rodriguez-, C Yin, A Mehal, B Izar, RA Friedman, RG Wells, UB Pajvani, Y Hoshida, HE Remotti, N Arpaia, J Zucman-Ros, M Karin, NC Henderson, I Tabas, RF Schwabe
Nature, 2022-10-05;0(0):.
Species: Mouse
Sample Types: Tissue Homogenates
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Evaluation and Characterization of Post-Stroke Lung Damage in a Murine Model of Cerebral Ischemia
Authors: J Faura, L Ramiro, A Simats, F Ma, A Penalba, T Gasull, A Rosell, J Montaner, A Bustamante
International Journal of Molecular Sciences, 2022-07-22;23(15):.
Species: Mouse
Sample Types: Tissue Homogenates
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Lung Fibrosis Is Improved by Extracellular Vesicles from IFNgamma-Primed Mesenchymal Stromal Cells in Murine Systemic Sclerosis
Authors: P Rozier, M Maumus, ATJ Maria, K Toupet, C Jorgensen, P Guilpain, D Noël
Cells, 2021-10-13;10(10):.
Species: Mouse
Sample Types: Cell Lysates
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Heparin-conjugated collagen as a potent growth factor-localizing and stabilizing scaffold for regenerative medicine
Authors: Y Ikegami, H Mizumachi, K Yoshida, H Ijima
Regenerative Therapy, 2020-11-01;15(0):236-242.
Species: Mouse
Sample Types: Cell Culture Supernates
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Tumor-independent host secretomes induced by angiogenesis and immune-checkpoint inhibitors
Authors: M Mastri, CR Lee, A Tracz, RS Kerbel, M Dolan, Y Shi, JML Ebos
Mol. Cancer Ther., 2018-04-25;0(0):.
Species: Mouse
Sample Types: Plasma
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Tpl2 regulates intestinal myofibroblast HGF release to suppress colitis-associated tumorigenesis.
Authors: Koliaraki V, Roulis M, Kollias G
J Clin Invest, 2012-10-15;122(11):4231-42.
Species: Mouse
Sample Types: Cell Culture Supernates
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Tumour hypoxia promotes tolerance and angiogenesis via CCL28 and T(reg) cells.
Authors: Facciabene A, Peng X, Hagemann IS, Balint K, Barchetti A, Wang LP, Gimotty PA, Gilks CB, Lal P, Zhang L, Coukos G
Nature, 2011-07-13;475(7355):226-30.
Species: Mouse
Sample Types: Cell Culture Supernates
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c-Met and its ligand hepatocyte growth factor/scatter factor regulate mature B cell survival in a pathway induced by CD74.
Authors: Gordin M, Tesio M, Cohen S
J. Immunol., 2010-07-16;185(4):2020-31.
Species: Mouse
Sample Types: Cell Culture Supernates
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CCL3-CCR5 axis regulates intratumoral accumulation of leukocytes and fibroblasts and promotes angiogenesis in murine lung metastasis process.
Authors: Wu Y, Li YY, Matsushima K, Baba T, Mukaida N
J. Immunol., 2008-11-01;181(9):6384-93.
Species: Mouse
Sample Types: Tissue Homogenates
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Stem cell factor and its receptor, c-kit, are important for hepatocyte proliferation in wild-type and tumor necrosis factor receptor-1 knockout mice after 70% hepatectomy.
Authors: Ren X, Hu B, Colletti L
Surgery, 2008-06-01;143(6):790-802.
Species: Mouse
Sample Types: Tissue Homogenates
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The EGF receptor is required for efficient liver regeneration.
Authors: Natarajan A, Wagner B, Sibilia M
Proc. Natl. Acad. Sci. U.S.A., 2007-10-16;104(43):17081-6.
Species: Mouse
Sample Types: Serum
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