Summary
Product Features
Kit Content
Other Reagents Required
Data Examples
Product Datasheets
Preparation and Storage
Background
Human EN-RAGE DuoSet ELISA Summary
Assay Type
Solid Phase Sandwich ELISA
Format
96-well strip plate
Assay Range
7.8 - 500 pg/mL
Sufficient Materials
For five 96-well plates*
Specificity
Please see the product datasheet
* 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 human Extracellular Newly Identified RAGE-binding Protein (EN-RAGE). The suggested diluent is suitable for the analysis of most cell culture supernate, serum, and plasma 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)
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
Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: EN-RAGE/S100A12
EN-RAGE, also named S100 calcium-binding protein A12, calcium-binding protein in amniotic fluid (CAAFl), calgranulin-related protein (CGRP), and calgranulin C, is a member of the S100/calgranulin superfamily.
Long Name:
Extracellular Newly Identified RAGE-binding Protein
Entrez Gene IDs:
6283 (Human)
Alternate Names:
CAAF1; CAAF1Neutrophil S100 protein; CAAFI; CAGC; CAGCS100 calcium binding protein A12 (calgranulin C); CAGCS100; Calcium-binding protein in amniotic fluid 1; Calgranulin C; calgranulin-C; CGRPEN-RAGE; ENRAGE; EN-RAGE; Extracellular newly identified RAGE-binding protein; MRP6; p6calgranulin C; protein S100-A12; S100 calcium binding protein A12; S100 calcium-binding protein A12 (calgranulin C); S100 calcium-binding protein A12; S100A12
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 Human EN-RAGE 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.
7
Citations: Showing 1 - 7
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Assessment of EN-RAGE, sRAGE, and its isoforms: cRAGE, esRAGE in obese patients treated by moderate caloric restriction combined with physical activity conducted in hospital condition
Authors: Kanikowska, D;Kanikowska, A;Strojny, Z;Kawka, E;Zawada, A;Rutkowski, R;Litwinowicz, M;Sato, M;Grzymis?awski, M;Br?borowicz, A;Witowski, J;Korybalska, K;
Cytokine
Species: Human
Sample Types: Serum
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Differential Expression of Circulating Damage-Associated Molecular Patterns in Patients with Coronary Artery Ectasia
Authors: Tsoporis, JN;Triantafyllis, AS;Kalogeropoulos, AS;Izhar, S;Rigopoulos, AG;Rallidis, LS;Sakadakis, E;Toumpoulis, IK;Salpeas, V;Leong-Poi, H;Parker, TG;Rizos, I;
Biomolecules
Species: Human
Sample Types: Plasma
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Antibody microarray analysis of amniotic fluid proteomes in women with cervical insufficiency and short cervix, and their association with pregnancy latency length
Authors: S Hong, KH Park, YE Lee, JE Lee, YM Kim, E Joo, I Cho
PLoS ONE, 2022-02-07;17(2):e0263586.
Species: Human
Sample Types: Amniotic Fluid
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Higher levels of the soluble receptor for advanced glycation end products and lower levels of the extracellular newly identified receptor for advanced glycation end products were associated with lipid-lowering drugs in patients with type 1 diabetes: a comparative cross-sectional study
Authors: EO Melin, J Dereke, M Hillman
Lipids Health Dis, 2020-10-14;19(1):223.
Species: Human
Sample Types: Plasma
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Distinct synovial tissue macrophage subsets regulate inflammation and remission in rheumatoid arthritis
Authors: S Alivernini, L MacDonald, A Elmesmari, S Finlay, B Tolusso, MR Gigante, L Petricca, C Di Mario, L Bui, S Perniola, M Attar, M Gessi, AL Fedele, S Chilaka, D Somma, SN Sansom, A Filer, C McSharry, NL Millar, K Kirschner, A Nerviani, MJ Lewis, C Pitzalis, AR Clark, G Ferracciol, I Udalova, CD Buckley, E Gremese, IB McInnes, TD Otto, M Kurowska-S
Nat. Med., 2020-06-29;0(0):.
Species: Human
Sample Types: Cell Culture Supernates
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MMP-12 and S100s in saliva reflect different aspects of periodontal inflammation
Authors: SB Holmström, R Lira-Junio, S Zwicker, M Majster, A Gustafsson, S Åkerman, B Klinge, M Svensson, EA Boström
Cytokine, 2018-07-06;0(0):.
Species: Human
Sample Types: Saliva
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Increased Plasma Levels of Danger-Associated Molecular Patterns Are Associated With Immune Suppression and Postoperative Infections in Patients Undergoing Cytoreductive Surgery and Hyperthermic Intraperitoneal Chemotherapy
Authors: GP Leijte, H Custers, J Gerretsen, A Heijne, J Roth, T Vogl, GJ Scheffer, P Pickkers, M Kox
Front Immunol, 2018-04-05;9(0):663.
Species: Human
Sample Types: Plasma
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Reviews for Human EN-RAGE DuoSet ELISA
Average Rating: 4.5 (Based on 2 Reviews)
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Human EN-RAGE DuoSet ELISA
By Jonatan Dereke
on 08/06/2019
Sample Tested: EDTA Plasma
We ran human EDTA-plasma at a 1:300 dilution for this analysis with low intra-assay variation.
Human EN-RAGE DuoSet ELISA
By Anonymous
on 10/12/2017
Sample Tested: Plasma
