Mouse IL-15 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. 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.
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-15
IL-15 (Interleukin 15) is a cytokine that induces or enhances the differentiation, maintenance, or activation of multiple T cell subsets including NK, NKT, Th17, Treg, and CD8+ memory cells. It also induces dendritic cell differentiation and inflammatory activation, exhibits anti-tumor activity, and inhibits the deposition of lipid in adipocytes. Complexes of IL-15 with cell surface IL-15 R alpha interact with complexes of IL-2 R beta and the Common gamma Chain on adjacent cells. This transpresentation mechanism enables cells to respond to IL-15 even if they do not express IL-15 R alpha. Ligation of membrane-associated IL-15/IL-15 R alpha complexes also induces reverse signaling that promotes activation of the IL-15/IL-15 R alpha expressing cells. The activity of circulating IL-15 is limited by its association with soluble IL-15 R alpha.
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, 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-15 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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Construction and utilization of a new generation of bacteriophage-based particles, or TPA, for guided systemic delivery of nucleic acids to tumors
Authors: Gay, L;Suwan, K;Hajitou, A;
Nature protocols
Species: Mouse
Sample Types: Cell Culture Supernates
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Tumor-induced natural killer cell dysfunction is a rapid and reversible process uncoupled from the expression of immune checkpoints
Authors: Pouxvielh, K;Marotel, M;Drouillard, A;Villard, M;Moreews, M;Bossan, A;Poiget, M;Khoryati, L;Benezech, S;Fallone, L;Hamada, S;Rousseaux, N;Picq, L;Rocca, Y;Berton, A;Teixeira, M;Mathieu, AL;Ainouze, M;Hasan, U;Fournier, A;Thaunat, O;Marçais, A;Walzer, T;
Science advances
Species: Mouse
Sample Types: Cell Culture Supernates
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Interferon-epsilon is a novel regulator of NK cell responses in the uterus
Authors: Mayall, JR;Horvat, JC;Mangan, NE;Chevalier, A;McCarthy, H;Hampsey, D;Donovan, C;Brown, AC;Matthews, AY;de Weerd, NA;de Geus, ED;Starkey, MR;Kim, RY;Daly, K;Goggins, BJ;Keely, S;Maltby, S;Baldwin, R;Foster, PS;Boyle, MJ;Tanwar, PS;Huntington, ND;Hertzog, PJ;Hansbro, PM;
EMBO molecular medicine
Species: Mouse
Sample Types: Uterine Lavage Fluid
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Systemically targeted cancer immunotherapy and gene delivery using transmorphic particles
Authors: P Asavarut, S Waramit, K Suwan, GJK Marais, A Chongchai, S Benjathumm, M Al-Bahrani, P Vila-Gomez, M Williams, P Kongtawele, T Yata, A Hajitou
Oncogene, 2022-06-27;0(0):e15418.
Species: Mouse
Sample Types: Cell Culture Supernates
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A Gut-Ex-Vivo System to Study Gut Inflammation Associated to Inflammatory Bowel Disease (IBD)
Authors: M Gagliardi, R Monzani, N Clemente, L Fusaro, V Saverio, G Grieco, E Pa?czyszyn, N Yissachar, F Boccafosch, M Corazzari
Biology, 2021-06-30;10(7):.
Species: Mouse
Sample Types: Tissue Homogenates
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Probiotics Supplements Reduce ER Stress and Gut Inflammation Associated with Gliadin Intake in a Mouse Model of Gluten Sensitivity
Authors: E Ferrari, R Monzani, V Saverio, M Gagliardi, E Pa?czyszyn, V Raia, VR Villella, G Bona, M Pane, A Amoruso, M Corazzari
Nutrients, 2021-04-07;13(4):.
Species: Mouse
Sample Types: Tissue Homogenates
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DeSUMOylase SENP7-Mediated Epithelial Signaling Triggers Intestinal Inflammation via Expansion of Gamma-Delta T Cells
Authors: A Suhail, ZA Rizvi, P Mujagond, SA Ali, P Gaur, M Singh, V Ahuja, A Awasthi, CV Srikanth
Cell Rep, 2019-12-10;29(11):3522-3538.e7.
Species: Mouse
Sample Types: Cell Culture Supernates
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Co-expression of IL-15 enhances anti-neuroblastoma effectivity of a tyrosine hydroxylase-directed DNA vaccination in mice
Authors: M Marx, M Zumpe, S Troschke-M, D Shah, HN Lode, N Siebert
PLoS ONE, 2018-11-19;13(11):e0207320.
Species: Chinese Hamster
Sample Types: Cell Culture Supernates
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Neuroprotective effects of ellagic acid on cuprizone-induced acute demyelination through limitation of microgliosis, adjustment of CXCL12/IL-17/IL-11 axis and restriction of mature oligodendrocytes apoptosis
Authors: N Sanadgol, F Golab, Z Tashakkor, N Taki, S Moradi Kou, A Mostafaie, M Mehdizadeh, M Abdollahi, G Taghizadeh, M Sharifzade
Pharm Biol, 2017-12-01;55(1):1679-1687.
Species: Mouse
Sample Types: Tissue Homogenates
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STAT1 Represses Cytokine-Producing Group 2 and Group 3 Innate Lymphoid Cells during Viral Infection
Authors: MT Stier, K Goleniewsk, JY Cephus, DC Newcomb, TP Sherrill, KL Boyd, MH Bloodworth, ML Moore, K Chen, JK Kolls, RS Peebles
J. Immunol., 2017-06-02;0(0):.
Species: Mouse
Sample Types: Tissue Homogenates
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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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Bone marrow produces sufficient alloreactive natural killer (NK) cells in vivo to cure mice from subcutaneously and intravascularly injected 4T1 breast cancer
Authors: Michel van Gelder
Breast Cancer Res. Treat, 2016-12-03;0(0):.
Species: Mouse
Sample Types: Plasma
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PGI2 Controls Pulmonary NK Cells That Prevent Airway Sensitization to House Dust Mite Allergen
Authors: Bryan Simons
J. Immunol, 2016-11-28;0(0):.
Species: Mouse
Sample Types: Cell Culture Supernates
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Mast Cells Limit the Exacerbation of Chronic Allergic Contact Dermatitis in Response to Repeated Allergen Exposure
Authors: Marcus Maurer
J. Immunol., 2016-11-02;0(0):.
Species: Mouse
Sample Types: Peritoneal Lavage Fluid
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Hydrodynamic delivery of interleukin 15 gene promotes resistance to high fat diet-induced obesity, fatty liver and improves glucose homeostasis.
Authors: Sun H, Liu D
Gene Ther, 2014-12-11;22(4):341-7.
Species: Mouse
Sample Types: Serum
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Armed oncolytic virus enhances immune functions of chimeric antigen receptor-modified T cells in solid tumors.
Authors: Nishio N, Diaconu I, Liu H, Cerullo V, Caruana I, Hoyos V, Bouchier-Hayes L, Savoldo B, Dotti G
Cancer Res, 2014-07-24;74(18):5195-205.
Species: Mouse
Sample Types: Serum
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BAFF suppresses IL-15 expression in B cells.
Authors: Ma, Ning, Xing, Chen, Xiao, He, He, Youdi, Han, Gencheng, Chen, Guojiang, Hou, Chunmei, Marrero, Bernadet, Wang, Yujuan, Zhang, Shengqua, Shen, Beifen, Li, Yan, Wang, Renxi
J Immunol, 2014-03-26;192(9):4192-201.
Species: Mouse
Sample Types: Serum
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IL-15 complexes induce NK- and T-cell responses independent of type I IFN signaling during rhinovirus infection.
Authors: Jayaraman A, Jackson D, Message S, Pearson R, Aniscenko J, Caramori G, Mallia P, Papi A, Shamji B, Edwards M, Westwick J, Hansel T, Stanciu L, Johnston S, Bartlett N
Mucosal Immunol, 2014-01-29;7(5):1151-64.
Species: Mouse
Sample Types: Tissue Homogenates
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Systemic inhibition of transforming growth factor-beta in glioma-bearing mice improves the therapeutic efficacy of glioma-associated antigen peptide vaccines.
Authors: Ueda R, Fujita M, Zhu X, Sasaki K, Kastenhuber ER, Kohanbash G, McDonald HA, Harper J, Lonning S, Okada H
Clin. Cancer Res., 2009-10-27;15(21):6551-9.
Species: Mouse
Sample Types: Plasma
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Immunological mechanisms and clinical implications of regulatory T cell deficiency in a systemic autoimmune disorder: roles of IL-2 versus IL-15.
Authors: Yang CH, Tian L, Ling GS, Trendell-Smith NJ, Ma L, Lo CK, Stott DI, Liew FY, Huang FP
Eur. J. Immunol., 2008-06-01;38(6):1664-76.
Species: Mouse
Sample Types: Serum
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Mouse fetal and embryonic liver cells differentiate human umbilical cord blood progenitors into CD56-negative natural killer cell precursors in the absence of interleukin-15.
Authors: McCullar V, Oostendorp R, Panoskaltsis-Mortari A, Yun G, Lutz CT, Wagner JE, Miller JS
Exp. Hematol., 2008-03-04;36(5):598-608.
Species: Mouse
Sample Types: Cell Culture Supernates
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IFN-alpha-induced murine B16 melanoma cancer vaccine cells: induction and accumulation of cell-associated IL-15.
Authors: Wu TG, Rose WA, Albrecht TB, Knutson EP, Konig R, Perdigao JR, Nguyen AP, Fleischmann WR
J. Interferon Cytokine Res., 2007-01-01;27(1):13-22.
Species: Mouse
Sample Types: Cell Culture Supernates
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A GMCSF and IL-15 fusokine leads to paradoxical immunosuppression in vivo via asymmetrical JAK/STAT signaling through the IL-15 receptor complex.
Authors: Rafei M, Wu JH, Annabi B, Lejeune L, Francois M, Galipeau J
Blood, 2006-11-02;109(5):2234-42.
Species: Mouse
Sample Types: Cell Culture Supernates
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This kit worked well with in vitro (cell culture supernatants) and in vivo (organ macerates) samples.
Optimal color development was achieved within 15 minutes of incubation (commercial TMB reagent) with minimal background.
ELISAs were performed on supernatants of various cell clones - nice range of secretion levels of Il-15
Some important reagents were not provided such as blocking buffer, and substrate solution. Directions did not include sufficient information about diluting the antibodies to the appropriate working concentration.
From R&D Systems:
The information required to properly dilute antibodies to working concentrations can be found in the documentation provided with your DuoSet development kit. If misplaced, the C of A can also be found for download on the product webpage or by contacting Bio-Techne Technical Service at techsupport@bio-techne.com. Technical Service is always available for any questions or assistance you may need to ensure a successful assay.
