Rat CXCL2/CINC-3 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 rat CINC-3. 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: CXCL2/GRO beta/MIP-2/CINC-3
CXCL2/GRO beta, also called MIP-2 in mouse and CINC-3 in rat, is a member of the CXC chemokine family. Human CXCL2/GRO beta is 107 amino acids (aa) in length with a predicted molecular weight of 11 kDa. The mouse and rat orthologs share 70% and 71% aa sequence identity with the human protein, respectively. N-terminal aa 1-4 of CXCL2/GRO beta can be post-translationally cleaved which confers enhanced hematopoietic bioactivity. CXCL2/GRO beta is produced by a variety of cell types including monocytes and macrophages at sites of inflammation and is chemotactic for granulocytes, including neutrophils.
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 Rat CXCL2/CINC-3 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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CXCR2-Blocking Has Context-Sensitive Effects on Rat Glioblastoma Cell Line Outgrowth (S635) in an Organotypic Rat Brain Slice Culture Depending on Microglia-Depletion (PLX5622) and Dexamethasone Treatment
Authors: Falter, J;Lohmeier, A;Eberl, P;Stoerr, EM;Koskimäki, J;Falter, L;Rossmann, J;Mederer, T;Schmidt, NO;Proescholdt, M;
International journal of molecular sciences
Species: Rat
Sample Types: Tissue Culture Supernates
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Pyridostigmine bromide elicits progressive and chronic impairments in the cholinergic anti-inflammatory pathway in the prefrontal cortex and hippocampus of male rats
Authors: HE Burzynski, VA Macht, JL Woodruff, JN Crawford, JM Erichsen, GG Piroli, CA Grillo, JR Fadel, LP Reagan
Neurobiology of stress, 2022-04-15;18(0):100446.
Species: Rat
Sample Types: Plasma
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Exposure to Nickel Oxide Nanoparticles Induces Acute and Chronic Inflammatory Responses in Rat Lungs and Perturbs the Lung Microbiome
Authors: MJ Jeong, S Jeon, HS Yu, WS Cho, S Lee, D Kang, Y Kim, YJ Kim, SY Kim
International Journal of Environmental Research and Public Health, 2022-01-04;19(1):.
Species: Rat
Sample Types: BALF
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Obesity and Type 2 Diabetes mellitus induce lipopolysaccharide tolerance in rat neutrophils
Authors: WMT Kuwabara, CNF Yokota, R Curi, TC Alba-Loure
Sci Rep, 2018-12-03;8(1):17534.
Species: Rat
Sample Types: BALF
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The role of Src & ERK1/2 kinases in inspiratory resistive breathing induced acute lung injury and inflammation
Authors: D Toumpanaki, V Vassilakop, I Sigala, P Zacharatos, I Vraila, V Karavana, S Theocharis, T Vassilakop
Respir. Res., 2017-12-13;18(1):209.
Species: Rat
Sample Types: Tissue Homogenates
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Supernatant of stored platelets causes lung inflammation and coagulopathy in a novel in vivo transfusion model.
Authors: Vlaar AP, Hofstra JJ, Kulik W
Blood, 2010-05-17;116(8):1360-8.
Species: Rat
Sample Types: Tissue Homogenates
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The lectin-like domain of tumor necrosis factor improves lung function after rat lung transplantation--potential role for a reduction in reactive oxygen species generation.
Authors: Hamacher J, Stammberger U, Roux J, Kumar S, Yang G, Xiong C, Schmid RA, Fakin RM, Chakraborty T, Hossain HM, Pittet JF, Wendel A, Black SM, Lucas R
Crit. Care Med., 2010-03-01;38(3):871-8.
Species: Rat
Sample Types: BALF
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Novel role for the liver X nuclear receptor in the suppression of lung inflammatory responses.
Authors: Birrell MA, Catley MC, Hardaker E, Wong S, Willson TM, McCluskie K, Leonard T, Farrow SN, Collins JL, Haj-Yahia S, Belvisi MG
J. Biol. Chem., 2007-08-31;282(44):31882-90.
Species: Rat
Sample Types: Tissue Homogenates
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Utility of exhaled nitric oxide as a noninvasive biomarker of lung inflammation in a disease model.
Authors: Birrell MA, McCluskie K, Hardaker E, Knowles R, Belvisi MG
Eur. Respir. J., 2006-09-27;28(6):1236-44.
Species: Rat
Sample Types: BALF
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Dichotomal role of TNF in experimental pulmonary edema reabsorption.
Authors: Braun C, Hamacher J, Morel DR, Wendel A, Lucas R
J. Immunol., 2005-09-01;175(5):3402-8.
Species: Rat
Sample Types: BALF
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