Mouse IL-15R alpha 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 IL-15 R alpha. 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: IL-15R alpha
IL-15 Ra (Interleukin 15 Receptor alpha), also known as CD215, is a transmembrane glycoprotein that plays an important role in NK and CD8+ memory T cell homeostasis and activation. It transmits IL-15 induced signals and additionally interacts with a complex of IL-2 R beta and the common gamma chain (gammaC) which are also subunits of the IL-2 receptor complex. In transpresentation, IL-15/IL-15 Ra complexes are expressed on the surface of one cell and interact with complexes of IL-2 R beta/gammaC on adjacent cells. IL-15/IL-15 Ra complexes can transmit reverse signaling that promotes adhesion and activation of the IL-15/IL-15 Ra expressing cells. Shed soluble forms of IL-15 Ra retain the ability to bind tightly to IL-15 and can inhibit IL-15 bioactivity.
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 IL-15R alpha 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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CD4 effector T cell differentiation is controlled by IL-15 that is expressed and presented in trans
Authors: AT Waickman, DL Ligons, S Hwang, JY Park, V Lazarevic, N Sato, C Hong, JH Park
Cytokine, 2017-08-12;0(0):.
Species: Mouse
Sample Types: Cell Culture Supernates
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Systemic inflammatory response syndrome-related lymphopenia is associated with adenosine A1 receptor dysfunction
Authors: R Riff, Y Cohen, H Eini-Rider, O Naamani, J Mazar, YS Haviv, C Chaimovitz, A Douvdevani
J. Leukoc. Biol., 2017-05-11;0(0):.
Species: Mouse
Sample Types: Peritoneal Lavage Fluid
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Rescue of nonlytic Newcastle Disease Virus (NDV) expressing IL-15 for cancer immunotherapy
Authors: X Xu, Q Sun, X Yu, L Zhao
Virus Res, 2017-03-07;233(0):35-41.
Species: Mouse
Sample Types: Cell Culture Supernates
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Different NK cell developmental events require different levels of IL-15 trans-presentation.
Authors: Lee GA, Liou YH, Wang SW, Ko KL, Jiang ST, Liao NS
J. Immunol., 2011-06-29;187(3):1212-21.
Species: Mouse
Sample Types: Cell Lysates
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Interleukin-15 combined with an anti-CD40 antibody provides enhanced therapeutic efficacy for murine models of colon cancer.
Authors: Zhang M, Yao Z, Dubois S, Ju W, Muller JR, Waldmann TA
Proc. Natl. Acad. Sci. U.S.A., 2009-04-21;106(18):7513-8.
Species: Mouse
Sample Types: Serum
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IL-15Ralpha chaperones IL-15 to stable dendritic cell membrane complexes that activate NK cells via trans presentation.
Authors: Mortier E, Woo T, Advincula R, Gozalo S, Ma A
J. Exp. Med., 2008-05-05;205(5):1213-25.
Species: Mouse
Sample Types: Cell Lysates
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Homeostatic MyD88-dependent signals cause lethal inflamMation in the absence of A20.
Authors: Turer EE, Tavares RM, Mortier E, Hitotsumatsu O, Advincula R, Lee B, Shifrin N, Malynn BA, Ma A
J. Exp. Med., 2008-02-11;205(2):451-64.
Species: Mouse
Sample Types: Serum
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Soluble Interleukin IL-15Ralpha is generated by alternative splicing or proteolytic cleavage and forms functional complexes with IL-15.
Authors: Bulanova E, Budagian V, Duitman E, Orinska Z, Krause H, Ruckert R, Reiling N, Bulfone-Paus S
J. Biol. Chem., 2007-02-27;282(18):13167-79.
Species: Mouse
Sample Types: Serum
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