Mouse IL-1ra/IL-1F3 Quantikine ELISA Kit

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MRA00
Mouse IL-1ra/IL-1F3 ELISA Standard Curve
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Mouse IL-1ra/IL-1F3 Quantikine ELISA Kit Summary

Assay Type
Solid Phase Sandwich ELISA
Format
96-well strip plate
Assay Length
4.5 hours
Sample Type & Volume Required Per Well
Cell Culture Supernates (50 uL), Serum (50 uL), EDTA Plasma (50 uL), Heparin Plasma (50 uL)
Sensitivity
13 pg/mL
Assay Range
31.2 - 2,000 pg/mL (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma)
Specificity
Natural and recombinant mouse IL-1ra
Cross-reactivity
< 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.
Interference
Interference observed with 1 or more available related molecules.

Product Summary

The Quantikine Mouse IL-1ra Immunoassay is a 4.5 hour solid-phase ELISA designed to measure mouse IL-1ra in cell culture supernates, serum, and plasma. It contains E. coli-expressed recombinant mouse IL-1ra and antibodies raised against the recombinant factor. This immunoassay has been shown to accurately quantitate the recombinant factor. Results obtained using natural mouse IL-1ra showed linear curves that were parallel to the standard curves obtained using the Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for naturally occurring mouse IL-1ra.

Precision

Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested on one plate to assess intra-assay precision
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in separate assays to assess inter-assay precision

Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma

Intra-Assay Precision Inter-Assay Precision
Sample 1 2 3 1 2 3
n 20 20 20 26 26 27
Mean 63 227 1226 66 225 1233
Standard Deviation 5.1 12.2 47.2 6.1 12.5 29.9
CV% 8.1 5.4 3.8 9.2 5.6 2.4

Recovery

The recovery of mouse IL-1ra spiked to three levels throughout the range of the assay in various matrices was evaluated.

Sample Type Average % Recovery Range %
Cell Culture Supernates (n=5) 110 95-120
EDTA Plasma (n=4) 92 82-101
Heparin Plasma (n=4) 91 83-101
Serum (n=4) 95 88-104

Linearity

To assess the linearity of the assay, samples containing or spiked with high concentrations of mouse IL-1ra in each matrix were diluted with Calibrator Diluent and then assayed.
Mouse IL-1ra/IL-1F3 ELISA Linearity

Data Examples

Mouse IL-1ra/IL-1F3 ELISA Standard Curve

Product Datasheets

Preparation and Storage

Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: IL-1ra/IL-1F3

Mouse Interleukin-1 receptor antagonist (IL-1ra) is a 22-25 kDa glycoprotein produced by a variety of cell types that antagonizes IL-1 activity (1-3). It is a member of the IL-1 family of proteins that includes IL-1 alpha and IL-1 beta. Although there is little amino acid (aa) identity (< 30%) among the three IL-1 family members, all molecules bind to the same receptors, all show a beta -trefoil structure, all are physically placed on mouse chromosome # 2, and all are believed to have evolved from a common ancestral gene (1-4). Mouse IL-1ra is synthesized as a 178 aa precursor that contains a 26 aa signal sequence plus a 152 aa mature region. There is one intrachain disulfide bond and one potential N-linked glycosylation site (3, 5, 6). Mature mouse IL-1ra shares 90%, 77%, 80%, 80%, and 78% aa sequence identity to mature rat (4), human (7), porcine (8), canine (9), and equine (10) IL-1ra, respectively. In humans, three non-secreted IL-1ra isoforms have also been identified (11-14). These result from the use of alternate start sites or exon splicing. In mice, only one of the three human intracellular isoforms has been isolated. This mouse molecule is the ortholog of the 159 aa human intracellular isoform # 1 (6, 12). The mouse intracellular form differs from the secreted precursor by only three amino acids. Cells known to secrete IL-1ra include dermal fibroblasts (15), vascular smooth muscle cells (16), intestinal columnar epithelium (17), chondrocytes (18), macrophages (19), non-keratinized oral stratified squamous epithelium (20), mast cells (21), neutrophils and monocytes (22), Sertoli cells (23), and hepatocytes (24). 

There are two type I transmembrane glycoprotein receptors for IL-1ra: the bioactive 80 kDa type I IL-1 receptor (IL-1 RI), and the inert (decoy) 65 kDa type II IL-1 receptor (IL-1 RII). IL-1ra binding to IL-1 RI competitively blocks IL-1 ( alpha or beta ) binding to the same receptor. Unlike the IL-1/IL-1 RI complex, the IL-1ra/IL-1 RI complex cannot recruit the IL-1 receptor accessory protein that is required for signal transduction. This results in receptor ligation without cell activation (1, 25). IL-1ra also competitively blocks IL-1 binding to the decoy IL-1 RII. In this case, IL-1ra may actually potentiate IL-1 activity by cancelling the functions of both antagonists. 
All activities attributed to IL-1ra are explained by its role as a competitive inhibitor of IL-1 binding to IL-1 RI (1, 2, 26, 27). In general, the ratio of IL-1ra to IL-1 beta is close to 1 in both health and disease (26, 28). To achieve total abrogation of IL-1 activity, more than 95% of all IL-1 RI receptors apparently need to be occupied by IL-1ra (29). Thus, local concentrations of secreted IL-1ra may be the critical determinants of IL-1 activity. The function of intracellular IL-1ra is less clear. While it would seem to be a simple competitor of IL-1 activity, its role may be limited to that of downmodulating non-specific inflammation associated with cell debris and death (14, 27). It has no action intracellularly; only when released through cell death would it then become a functional IL-1 antagonist.

Long Name:
Interleukin 1 Receptor Antagonist
Entrez Gene IDs:
3557 (Human); 16181 (Mouse); 60582 (Rat); 397499 (Porcine)
Alternate Names:
DIRA; ICIL-1ra; IL1F3; IL-1F3; IL1ra; IL-1ra; IL-1ra3; IL1RN; IL-1RN; interleukin 1 receptor antagonist; IRAP; MVCD4
⚠ WARNING: This product can expose you to chemicals including N,N-Dimethylforamide, which is known to the State of California to cause cancer. For more information, go to www.P65Warnings.ca.gov.

Assay Procedure

Refer to the product for complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.
  1.   Prepare all reagents, standard dilutions, and samples as directed in the product insert.
  2.   Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

  3. 50 µL Assay Diluent
  4.   Add 50 µL of Assay Diluent to each well.

  5. 50 µL Standard, Control, or Sample
  6.   Add 50 µL of Standard, Control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours on a horizontal orbital microplate shaker.
  7.   Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.

  8. 100 µL Conjugate
  9.   Add 100 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours on the shaker.
  10.   Aspirate and wash 5 times.

  11. 100 µL Substrate Solution
  12.   Add 100 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes on the benchtop. PROTECT FROM LIGHT.

  13. 100 µL Stop Solution
  14.   Add 100 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.

Citations for Mouse IL-1ra/IL-1F3 Quantikine ELISA Kit

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.

27 Citations: Showing 1 - 10
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  1. Type I interferon-driven susceptibility to Mycobacterium tuberculosis is mediated by IL-1Ra
    Authors: DX Ji, LH Yamashiro, KJ Chen, N Mukaida, I Kramnik, KH Darwin, RE Vance
    Nat Microbiol, 2019;0(0):.
    Species: Mouse
    Sample Types: Tissue Homogenates
  2. Diet-Induced Obese Mice and Leptin-Deficient Lepob/ob Mice Exhibit Increased Circulating GIP Levels Produced by Different Mechanisms
    Authors: E Lee, EL Miedzybrod, X Zhang, R Hatano, J Miyamoto, I Kimura, K Fujimoto, S Uematsu, S Rodriguez-, A Vidal-Puig, FM Gribble, F Reimann, T Miki
    Int J Mol Sci, 2019;20(18):.
    Species: Mouse
    Sample Types: Plasma
  3. miR-100 maintains phenotype of tumor-associated macrophages by targeting mTOR to promote tumor metastasis via Stat5a/IL-1ra pathway in mouse breast cancer
    Authors: W Wang, Y Liu, J Guo, H He, X Mi, C Chen, J Xie, S Wang, P Wu, F Cao, L Bai, Q Si, R Xiang, Y Luo
    Oncogenesis, 2018;7(12):97.
    Species: Mouse
    Sample Types: Tissue Homogenates
  4. Interleukin-1 receptor antagonist-mediated neuroprotection by umbilical cord-derived mesenchymal stromal cells following transplantation into a rodent stroke model
    Authors: SH Oh, C Choi, JE Noh, N Lee, YW Jeong, I Jeon, JM Shin, JH Kim, HJ Kim, JM Lee, HS Kim, OJ Kim, J Song
    Exp. Mol. Med., 2018;50(4):22.
    Species: Mouse
    Sample Types: Cell Culture Supernates
  5. Effect of internal contamination with tritiated water on the neoplastic colonies in the lungs, innate anti-tumour reactions, cytokine profile, and haematopoietic system in radioresistant and radiosensitive mice
    Authors: EM Nowosielsk, A Cheda, R Zdanowski, S Lewicki, BR Scott, MK Janiak
    Radiat Environ Biophys, 2018;0(0):.
    Species: Mouse
    Sample Types: Serum
  6. IL-24 Promotes Pseudomonas aeruginosa Keratitis in C57BL/6 Mouse Corneas
    Authors: BX Ross, N Gao, X Cui, TJ Standiford, J Xu, FX Yu
    J. Immunol, 2017;0(0):.
    Species: Mouse
    Sample Types: Tissue Homogenates
  7. Controlled Release of Interleukin-1 Receptor Antagonist from Hyaluronic Acid-Chitosan Microspheres Attenuates Interleukin-1?-Induced Inflammation and Apoptosis in Chondrocytes
    Biomed Res Int, 2016;2016(0):6290957.
  8. C-type Lectin Mincle Recognizes Glucosyl-diacylglycerol of Streptococcus pneumoniae and Plays a Protective Role in Pneumococcal Pneumonia
    PLoS Pathog, 2016;12(12):e1006038.
    Species: Mouse
    Sample Types: Tissue Homogenates
  9. Sharpin Controls Osteogenic Differentiation of Mesenchymal Bone Marrow Cells.
    Authors: Jeschke A, Catala-Lehnen P, Sieber S, Bickert T, Schweizer M, Koehne T, Wintges K, Marshall R, Mautner A, Duchstein L, Otto B, Horst A, Amling M, Kreienkamp H, Schinke T
    J Immunol, 2015;195(8):3675-84.
    Species: Mouse
    Sample Types: Serum
  10. Prion infection of mouse brain reveals multiple new upregulated genes involved in neuroinflammation or signal transduction.
    Authors: Carroll J, Striebel J, Race B, Phillips K, Chesebro B
    J Virol, 2015;89(4):2388-404.
    Species: Mouse
    Sample Types: Tissue Homogenates
  11. Correction: Yersinia pestis Activates Both IL-1beta and IL-1 Receptor Antagonist to Modulate Lung Inflammation during Pneumonic Plague.
    PLoS Pathog, 2015;11(5):e1004921.
    Species: Mouse
    Sample Types: BALF
  12. Inflammatory bone loss in experimental periodontitis induced by Aggregatibacter actinomycetemcomitans in interleukin-1 receptor antagonist knockout mice.
    Authors: Izawa A, Ishihara Y, Mizutani H, Kobayashi S, Goto H, Okabe E, Takeda H, Ozawa Y, Kamiya Y, Sugita Y, Kubo K, Kamei H, Kikuchi T, Mitani A, Hayashi J, Nishihara T, Maeda H, Noguchi T
    Infect Immun, 2014;82(5):1904-13.
    Species: Mouse
    Sample Types: Cell Culture Supernates
  13. The multifaceted effects of polysaccharides isolated from Dendrobium huoshanense on immune functions with the induction of interleukin-1 receptor antagonist (IL-1ra) in monocytes.
    Authors: Lin J, Chang Y, Yang W, Yu A, Wong C
    PLoS ONE, 2014;9(4):e94040.
    Species: Mouse
    Sample Types: Serum
  14. Impaired bone formation and increased osteoclastogenesis in mice lacking chemokine (C-C motif) ligand 5 (Ccl5).
    Authors: Wintges K, Beil F, Albers J, Jeschke A, Schweizer M, Claass B, Tiegs G, Amling M, Schinke T
    J Bone Miner Res, 2013;28(10):2070-80.
    Species: Mouse
    Sample Types: Cell Culture Supernates
  15. Inflammation induced by MMP-9 enhances tumor regression of experimental breast cancer.
    Authors: Leifler K, Svensson S, Abrahamsson A, Bendrik C, Robertson J, Gauldie J, Olsson A, Dabrosin C
    J Immunol, 2013;190(8):4420-30.
    Species: Mouse
    Sample Types: Tissue Homogenates
  16. Human mesenchymal stem/stromal cells cultured as spheroids are self-activated to produce prostaglandin E2 that directs stimulated macrophages into an anti-inflammatory phenotype.
    Authors: Ylostalo J, Bartosh T, Coble K, Prockop D
    Stem Cells, 2012;30(10):2283-96.
    Species: Mouse
    Sample Types: Cell Culture Supernates
  17. Fasting induces an anti-inflammatory effect on the neuroimmune system which a high-fat diet prevents.
    Authors: Lavin DN, Joesting JJ, Chiu GS, Moon ML, Meng J, Dilger RN, Freund GG
    Obesity (Silver Spring), 2011;19(8):1586-94.
    Species: Mouse
    Sample Types: Serum
  18. TLR4 is a negative regulator in noninfectious lung inflammation.
    Authors: Zhao H, Leu SW, Shi L, Dedaj R, Zhao G, Garg HG, Shen L, Lien E, Fitzgerald KA, Shiedlin A, Shen H, Quinn DA, Hales CA
    J. Immunol., 2010;184(9):5308-14.
    Species: Mouse
    Sample Types: BALF
  19. Activation of NOD2 in vivo induces IL-1beta production in the eye via caspase-1 but results in ocular inflammation independently of IL-1 signaling.
    Authors: Rosenzweig HL, Martin TM, Planck SR, Galster K, Jann MM, Davey MP, Kobayashi K, Flavell RA, Rosenbaum JT
    J. Leukoc. Biol., 2008;84(2):529-36.
    Species: Mouse
    Sample Types: Tissue Homogenates
  20. A role for IL-1 receptor antagonist or other cytokines in the acute therapeutic effects of IVIg?
    Authors: Crow AR, Song S, Semple JW, Freedman J, Lazarus AH
    Blood, 2007;109(1):155-8.
    Species: Mouse
    Sample Types: Serum
  21. Interleukin 1 receptor antagonist mediates the antiinflammatory and antifibrotic effect of mesenchymal stem cells during lung injury.
    Authors: Ortiz LA, Dutreil M, Fattman C, Pandey AC, Torres G, Go K, Phinney DG
    Proc. Natl. Acad. Sci. U.S.A., 2007;104(26):11002-7.
    Species: Mouse
    Sample Types: Cell Culture Supernates
  22. Intrapulmonary delivery of bone marrow-derived mesenchymal stem cells improves survival and attenuates endotoxin-induced acute lung injury in mice.
    Authors: Gupta N, Su X, Popov B, Lee JW, Serikov V, Matthay MA
    J. Immunol., 2007;179(3):1855-63.
    Species: Mouse
    Sample Types: BALF
  23. Absence of IL-1 receptor antagonist impaired wound healing along with aberrant NF-kappaB activation and a reciprocal suppression of TGF-beta signal pathway.
    Authors: Ishida Y, Kondo T, Kimura A, Matsushima K, Mukaida N
    J. Immunol., 2006;176(9):5598-606.
    Species: Mouse
    Sample Types: Tissue Homogenates
  24. Interleukin-1 beta contributes to the upregulation of kappa opioid receptor mrna in dorsal root ganglia in response to peripheral inflammation.
    Authors: Puehler W, Rittner HL, Mousa SA, Brack A, Krause H, Stein C, Schafer M
    Neuroscience, 2006;141(2):989-98.
    Species: Rat
    Sample Types: Cell Culture Supernates
  25. A deficit in astroglial organization causes the impaired reactive sprouting in human apolipoprotein E4 targeted replacement mice.
    Authors: Blain JF, Sullivan PM, Poirier J
    Neurobiol. Dis., 2005;21(3):505-14.
    Species: Mouse
    Sample Types: Tissue Homogenates
  26. Regulation of Pseudomonas aeruginosa corneal infection in IL-1 beta converting enzyme (ICE, caspase-1) deficient mice.
    Authors: Thakur A, Barrett RP, McClellan S, Hazlett LD
    Curr. Eye Res., 2004;29(4):225-33.
    Species: Mouse
    Sample Types: Tissue Homogenates
  27. IL-1 receptor-associated kinase 1 mediates protection against Staphylococcus aureus infection.
    Authors: Verdrengh M, Thomas JA, Hultgren OH
    Microbes Infect., 2004;6(14):1268-72.
    Species: Mouse
    Sample Types: Serum

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