Human IL-36 alpha/IL-1F6 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.
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.
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)
Scientific Data
Product Datasheets
Preparation and Storage
Background: IL-36 alpha/IL-1F6
Interleukin 36 alpha (IL-36 alpha), also known as IL-1F6 and FIL1 epsilon, is a member of the IL-1 family of proteins. Cells reported to express IL-36 alpha include monocytes, B cells, and T cells. Notably, IL-36 alpha is the only novel IL-1 family member expressed on T-cells. The receptor for IL-36 alpha is reported to be a combination of IL-1 R6/IL-1 R rp2 and IL-1 R3/IL-1 R AcP. Recombinant IL-36 alpha has been shown to activate pathways involving NF-kappa B and MAPK in an IL-1 R6/IL-1 R rp2-dependent manner.
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 of Block Buffer 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, 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 IL-36 alpha/IL-1F6 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 - 8
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Interleukin-36alpha is elevated in diffuse systemic sclerosis and may potentiate fibrosis
Authors: S O'Reilly
Cytokine, 2022-06-03;156(0):155921.
Species: Human
Sample Types: Serum
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IL-36alpha/IL-36RA/IL-38 signaling mediates inflammation and barrier disruption in human corneal epithelial cells under hyperosmotic stress
Authors: JM Li, R Lu, Y Zhang, J Lin, X Hua, SC Pflugfelde, DQ Li
The ocular surface, 2021-08-21;22(0):163-171.
Species: Human
Sample Types: Cell Culture Supernates
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Interleukin-36&alpha suppresses growth of non-small cell lung cancer in vitro by reducing angiogenesis
Authors: X Xie, H Hu, J He, Y Liu, F Guo, F Luo, M Jiang, L Wang
FEBS Open Bio, 2021-05-02;11(5):1353-1363.
Species: Human
Sample Types: Cell Culture Supernates
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Interleukin-38 ameliorates poly(I:C) induced lung inflammation: therapeutic implications in respiratory viral infections
Authors: X Gao, PKS Chan, GCY Lui, DSC Hui, IM Chu, X Sun, MS Tsang, BCL Chan, CW Lam, CK Wong
Cell Death & Disease, 2021-01-07;12(1):53.
Species: Human
Sample Types: Serum
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Increased serum IL-36gamma levels are associated with disease severity in myasthenia gravis patients
Authors: QX Zhang, Y Li, SM Jiang, LJ Zhang, M Yi, J Wang, Y Qi, L Yang, CS Yang
BMC Neurol, 2020-08-19;20(1):307.
Species: Human
Sample Types: Serum
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Interleukin (IL)-17/IL-36 axis participates to the crosstalk between endothelial cells and keratinocytes during inflammatory skin responses
Authors: L Mercurio, CM Failla, L Capriotti, C Scarponi, F Facchiano, M Morelli, S Rossi, G Pagnanelli, C Albanesi, A Cavani, S Madonna
PLoS ONE, 2020-04-30;15(4):e0222969.
Species: Human
Sample Types: Cell Culture Supernates
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Interleukin-36 cytokines alter the intestinal microbiome and can protect against obesity and metabolic dysfunction
Authors: E Giannoudak, YE Hernandez-, K Mulfaul, SL Doyle, E Hams, PG Fallon, A Mat, D O'Shea, M Kopf, AE Hogan, PT Walsh
Nat Commun, 2019-09-05;10(1):4003.
Species: Human
Sample Types: Serum
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The role of IL-36? and its regulation in eosinophilic inflammation in allergic rhinitis
Authors: X Qin, M Liu, S Zhang, C Wang, T Zhang
Cytokine, 2019-03-01;117(0):84-90.
Species: Human
Sample Types: Serum
FAQs
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How soon after reconstitution should standards in the Human IL-36 alpha/IL-1F6 DuoSet ELISA Kit, Catalog # DY1078-05, be used, or aliquotted and stored?
Standards in this kit are very sensitive and should not be allowed to sit longer than about 15-20 minutes before they are aliquotted and stored, undiluted, at -80C. Standards left at room temperature for longer periods could lead to a significant drop in standard signal on subsequent use. Standards should be used soon after reconstitution or after a new aliquot is thawed.
Reviews for Human IL-36 alpha/IL-1F6 DuoSet ELISA
Average Rating: 3 (Based on 1 Review)
Have you used Human IL-36 alpha/IL-1F6 DuoSet ELISA?
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We used this kit for the quantification of IL-36 alpha in human serum and plasma. Unfortunately, we could not find any IL-36 in our samples. Nevertheless, the standard curve works very well.
