Human sgp130 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 human gp130. 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: gp130
Gp130, the common signal transducing receptor component shared by the functional receptor complexes of the IL-6 family of cytokines, belongs to the classI cytokine receptor family. Binding of IL-6 or IL-11 to either the membrane-anchored or soluble IL-6 R or IL-11 R, respectively, initiates the association of IL-6 R (IL-11 R) with gp130. The complex then undergoes homodimerization and signal transduction. For other IL-6 family cytokines, such as LIF and OSM, signal transduction is similarly triggered by the heterodimerization of gp130 with LIFR or OSM R.
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 Human sgp130 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.
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Citations: Showing 1 - 10
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Targeting IL-6 trans-signalling by sgp130Fc attenuates severity in SARS-CoV-2 -infected mice and reduces endotheliopathy
Authors: Rodríguez-Hernández, MÁ;Baena-Bustos, M;Carneros, D;Zurita-Palomo, C;Muñoz-Pinillos, P;Millán, J;Padillo, FJ;Smerdou, C;von Kobbe, C;Rose-John, S;Bustos, M;
EBioMedicine
Species: Transgenic Mouse
Sample Types: Serum
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Transgenic inhibition of interleukin-6 trans-signaling does not prevent skeletal pathologies in mucolipidosis type II mice
Authors: LM Westermann, A Baranowsky, G Di Lorenzo, T Danyukova, J Soul, JM Schwartz, G Hendrickx, M Amling, S Rose-John, C Garbers, T Schinke, S Pohl
Scientific Reports, 2021-02-11;11(1):3556.
Species: Mouse
Sample Types: Serum
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IL-6 Trans-Signaling in the Brain Influences the Metabolic Phenotype of the 3xTg-AD Mouse Model of Alzheimer's Disease
Authors: A Escrig, A Molinero, B Méndez, M Giralt, G Comes, P Sanchis, O Fernández-, L Giménez-Ll, C Becker-Pau, S Rose-John, J Hidalgo
Cells, 2020-07-02;9(7):.
Species: Mouse
Sample Types: Tissue Homogenates
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Trans IL-6 signaling does not appear to play a role in renal scarring after urinary tract infection
Authors: S Gupta, GY Junquera, L Nicassio, B Becknell, CB Ching
J Pediatr Urol, 2020-05-29;0(0):.
Species: Human
Sample Types: Urine
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IL-6 exhibits both cis and trans signaling in osteocytes and osteoblasts, but only�trans�signaling promotes bone formation and osteoclastogenesis
Authors: NE McGregor, M Murat, J Elango, IJ Poulton, EC Walker, B Crimeen-Ir, PWM Ho, JH Gooi, TJ Martin, NA Sims
J. Biol. Chem., 2019-03-28;0(0):.
Species: Mouse
Sample Types: Serum
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The effect of rosuvastatin on inflammation, matrix turnover and left ventricular remodeling in dilated cardiomyopathy: a randomized, controlled trial.
Authors: Broch K, Askevold E, Gjertsen E, Ueland T, Yndestad A, Godang K, Stueflotten W, Andreassen J, Svendsmark R, Smith H, Aakhus S, Aukrust P, Gullestad L
PLoS ONE, 2014-02-25;9(2):e89732.
Species: Human
Sample Types: Whole Blood
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Altered interleukin-6 receptor, IL-6R and gp130, production and expression and decreased SOCS-3 expression in placentas from women with pre-eclampsia.
Authors: Zhao S, Gu Y, Dong Q, Fan R, Wang Y
Placenta, 2008-11-05;29(12):1024-8.
Species: Human
Sample Types: Cell Culture Supernates
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Soluble gp130, an antagonist of IL-6 transsignaling, is elevated in uveitis aqueous humor.
Authors: Simon D, Denniston AK, Tomlins PJ, Wallace GR, Rauz S, Salmon M, Murray PI, Curnow SJ
Invest. Ophthalmol. Vis. Sci., 2008-05-09;49(9):3988-91.
Species: Human
Sample Types: Serum
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Suspension microarrays for the identification of the response patterns in hyperinflammatory diseases.
Authors: Hsu HY, Wittemann S, Schneider EM, Weiss M, Joos TO
Med Eng Phys, 2008-03-03;30(8):976-83.
Species: Human
Sample Types: Plasma
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Transgenic blockade of interleukin 6 transsignaling abrogates inflammation.
Authors: Rabe B, Chalaris A, May U, Waetzig GH, Seegert D, Williams AS, Jones SA, Rose-John S, Scheller J
Blood, 2007-11-07;111(3):1021-8.
Species: Transgenic Mouse
Sample Types: Air Pouch Exudate
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