Mouse CCL21/6Ckine 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 CCL21/6Ckine. 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: CCL21/6Ckine
CCL21, also known as 6Ckine, TCA-4, SLC, Exodus-2, and A21, is a homeostatic chemokine that signals through CCR7 or CXCR3. CCL21 is constitutively presented on initial lymphatic vessels, high endothelial venules (HEV), and lymph node dendritic cells (DC) where it promotes the docking of DC to lymphatic vessels and the retention of T cells by lymph node DC, resulting in T cell priming for activation. Its upregulation during chronic inflammation or tissue damage promotes fibrosis, inflammatory cytokine production, and neuropathic pain. The soluble chemokine is elevated in rheumatoid arthritis synovial fluid and in the serum of coronary artery disease patients.
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 CCL21/6Ckine 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.
21
Citations: Showing 1 - 10
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Aging modulates homeostatic leukocyte trafficking to the peritoneal cavity in a sex-specific manner
Authors: Hopkin, SJ;Pezhman, L;Begum, J;Kavanagh, D;McGettrick, HM;Iqbal, AJ;Chimen, M;
Journal of leukocyte biology
Species: Mouse
Sample Types: Peritoneal Lavage Fluid
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Neuroimmune proteins can differentiate between tauopathies
Authors: JD Cherry, ZH Baucom, KG Eppich, D Kirsch, ER Dixon, Y Tripodis, KF Bieniek, K Farrell, K Whitney, M Uretsky, JF Crary, D Dickson, AC McKee
Journal of Neuroinflammation, 2022-11-19;19(1):278.
Species: Human
Sample Types: CSF
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Heparan sulfate is essential for thymus growth
Authors: HP Hsu, YT Chen, YY Chen, CY Lin, PY Chen, SY Liao, CCY Lim, Y Yamaguchi, CL Hsu, IL Dzhagalov
The Journal of Biological Chemistry, 2021-02-15;0(0):100419.
Species: Mouse
Sample Types: Tissue Homogenates
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Dual-mechanism based CTLs infiltration enhancement initiated by Nano-sapper potentiates immunotherapy against immune-excluded tumors
Authors: Y Huang, Y Chen, S Zhou, L Chen, J Wang, Y Pei, M Xu, J Feng, T Jiang, K Liang, S Liu, Q Song, G Jiang, X Gu, Q Zhang, X Gao, J Chen
Nat Commun, 2020-01-30;11(1):622.
Species: Mouse
Sample Types: Tissue Homogenates
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Cytotoxic chemotherapy reduces T cell trafficking to the spleen by downregulating the expression of C-C motif chemokine ligand 21 and C-C motif chemokine ligand 19
Authors: L Liu, L Zhao, Y Yang, J Gao, C Hu, B Guo, B Zhu
Oncol Lett, 2018-08-09;16(4):5013-5019.
Species: Mouse
Sample Types: Tissue Homogenates
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Regulation of T cell afferent lymphatic migration by targeting LT?R-mediated non-classical NF?B signaling
Authors: W Piao, Y Xiong, K Famulski, CC Brinkman, L Li, N Toney, C Wagner, V Saxena, T Simon, JS Bromberg
Nat Commun, 2018-08-01;9(1):3020.
Species: Mouse
Sample Types: Cell Culture Supernates
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Identification of periplakin as a major regulator of lung injury and repair in mice
Authors: V Besnard, R Dagher, T Madjer, A Joannes, M Jaillet, M Kolb, P Bonniaud, LA Murray, MA Sleeman, B Crestani
JCI Insight, 2018-03-08;3(5):.
Species: Mouse
Sample Types: BALF
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Expression of the B cell differentiation factor BAFF and chemokine CXCL13 in a murine model of Respiratory Syncytial Virus infection
Authors: W Alturaiki, AJ McFarlane, K Rose, R Corkhill, PS McNamara, J Schwarze, BF Flanagan
Cytokine, 2018-02-01;0(0):.
Species: Mouse
Sample Types: Tissue Homogenates
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Infection Programs Sustained Lymphoid Stromal Cell Responses and Shapes Lymph Node Remodeling upon Secondary Challenge
Authors: JL Gregory, A Walter, YO Alexandre, JL Hor, R Liu, JZ Ma, S Devi, N Tokuda, Y Owada, LK Mackay, GK Smyth, WR Heath, SN Mueller
Cell Rep, 2017-01-10;18(2):406-418.
Species: Mouse
Sample Types: Tissue Homogenates
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Age-Dependent Cell Trafficking Defects in Draining Lymph Nodes Impair Adaptive Immunity and Control of West Nile Virus Infection.
Authors: Richner J, Gmyrek G, Govero J, Tu Y, van der Windt G, Metcalf T, Haddad E, Textor J, Miller M, Diamond M
PLoS Pathog, 2015-07-23;11(7):e1005027.
Species: Mouse
Sample Types: Tissue Homogenates
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De novo-induced self-antigen-specific Foxp3+ regulatory T cells impair the accumulation of inflammatory dendritic cells in draining lymph nodes.
Authors: Alissafi T, Hatzioannou A, Ioannou M, Sparwasser T, Grun J, Grutzkau A, Verginis P
J Immunol, 2015-05-06;194(12):5812-24.
Species: Mouse
Sample Types: Tissue Homogenates
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CCRL1/ACKR4 is expressed in key thymic microenvironments but is dispensable for T lymphopoiesis at steady state in adult mice.
Authors: Lucas B, White A, Ulvmar M, Nibbs R, Sitnik K, Agace W, Jenkinson W, Anderson G, Rot A
Eur J Immunol, 2015-01-16;45(2):574-83.
Species: Mouse
Sample Types: Tissue Homogenates
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Plasmodium suppresses expansion of T cell responses to heterologous infections.
Authors: White C, Villarino N, Sloan S, Ganusov V, Schmidt N
J Immunol, 2014-12-10;194(2):697-708.
Species: Mouse
Sample Types: Tissue Homogenates
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The B lymphocyte differentiation factor (BAFF) is expressed in the airways of children with CF and in lungs of mice infected with Pseudomonas aeruginosa.
Authors: Neill, Daniel R, Saint, Gemma L, Bricio-Moreno, Laura, Fothergill, Joanne L, Southern, Kevin W, Winstanley, Craig, Christmas, Stephen, Slupsky, Joseph R, McNamara, Paul S, Kadioglu, Aras, Flanagan, Brian F
PLoS ONE, 2014-05-21;9(5):e95892.
Species: Mouse
Sample Types: Tissue Homogenates
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A TNF-alpha-CCL20-CCR6 axis regulates Nod1-induced B cell responses.
Authors: Paradis M, Mindt B, Duerr C, Rojas O, Ng D, Boulianne B, McCarthy D, Yu M, Summers deLuca L, Ward L, Waldron J, Philpott D, Gommerman J, Fritz J
J Immunol, 2014-02-17;192(6):2787-99.
Species: Mouse
Sample Types: Serum
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Hypomorphic mutation in the RAG2 gene affects dendritic cell distribution and migration.
Authors: Maina, Virginia, Marrella, Veronica, Mantero, Stefano, Cassani, Barbara, Fontana, Elena, Anselmo, Achille, Del Prete, Annalisa, Sozzani, Silvano, Vezzoni, Paolo, Poliani, Pietro L, Villa, Anna
J Leukoc Biol, 2013-09-19;94(6):1221-30.
Species: Mouse
Sample Types: Tissue Homogenates
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Membrane-associated CD93 regulates leukocyte migration and C1q-hemolytic activity during murine peritonitis.
Authors: Greenlee-Wacker MC, Briseno C, Galvan M, Moriel G, Velazquez P, Bohlson SS
J. Immunol., 2011-08-17;187(6):3353-61.
Species: Mouse
Sample Types: Peritoneal Lavage Fluid
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Induction of targeted cell migration by cutaneous administration of a DNA vector encoding a biologically active chemokine CCL21.
Authors: Jalili A, Pashenkov M, Kriehuber E, Wagner C, Nakano H, Stingl G, Wagner SN
J. Invest. Dermatol., 2010-02-25;130(6):1611-23.
Species: Mouse
Sample Types: Cell Culture Supernates
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Role of lymphotoxin-alpha in cigarette smoke-induced inflammation and lymphoid neogenesis.
Authors: Demoor T, Bracke KR, Maes T, Vandooren B, Elewaut D, Pilette C, Joos GF, Brusselle GG
Eur. Respir. J., 2009-01-22;34(2):405-16.
Species: Mouse
Sample Types: BALF
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Cyclooxygenase inhibition during allergic sensitization increases STAT6-independent primary and memory Th2 responses.
Authors: Zhou W, Newcomb DC, Moore ML, Goleniewska K, O'Neal JF, Peebles RS
J. Immunol., 2008-10-15;181(8):5360-7.
Species: Mouse
Sample Types: Tissue Homogenates
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Therapeutic targeting of CC ligand 21 or CC chemokine receptor 7 abrogates pulmonary fibrosis induced by the adoptive transfer of human pulmonary fibroblasts to immunodeficient mice.
Authors: Pierce EM, Carpenter K, Jakubzick C, Kunkel SL, Flaherty KR, Martinez FJ, Hogaboam CM
Am. J. Pathol., 2007-04-01;170(4):1152-64.
Species: Mouse
Sample Types: Tissue Homogenates
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Excellent post-sale service. Many thanks to Drs. Nagarajan and Riedeman, for the excellent technical support.
