Rat Lipocalin-2/NGAL 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 Lipocalin-2/NGAL. 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: Lipocalin-2/NGAL
The Lipocalin family comprises a diverse group of mostly secreted soluble proteins that bind hydrophobic ligands and act as transporters, carrying small molecules to specific cells. Lipocalins are related by possessing an 8-stranded beta-barrel structure. Lipocalin-1, also named tear lipocalin (TL), von Ebners gland protein (VEG) and tear pre-albumin, binds a large number of hydrophobic molecules and exhibits cysteine proteinase inhibitor and endonuclear activities. Lipocalin-2, also known as neutrophil gelatinase-associated lipocalin (NGAL), is a component of granules in neutrophils from tissues that are normally exposed to microorganisms and is upregulated during inflammation. Lipocalin-2 can form homodimers and can heterodimerize with the neutrophil gelatinase MMP-9.
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 Lipocalin-2/NGAL 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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MicroRNA?17?5p alleviates sepsis?related acute kidney injury in mice by modulating inflammation and apoptosis
Authors: Sun, J;Niu, L;Wang, Y;Zhao, G;Tang, L;Jiang, J;Pan, S;Ge, X;
Molecular medicine reports
Species: Mouse
Sample Types: Urine
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Effects of a lysophosphatidic acid receptor 1 antagonist on hypertensive renal injury in Dahl-Iwai salt-sensitive rats
Authors: T Naruse, H Otake, T Takahashi
Oncogene, 2022-05-13;149(4):179-188.
Species: Rat
Sample Types: Urine
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Study of the Role of the Tyrosine Kinase Receptor MerTK in the Development of Kidney Ischemia-Reperfusion Injury in RCS Rats
Authors: T Pelé, S Giraud, S Joffrion, V Ameteau, A Delwail, JM Goujon, L Macchi, T Hauet, F Dkhissi, O Benzakour
International Journal of Molecular Sciences, 2021-11-09;22(22):.
Species: Rat
Sample Types: Plasma
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miR?214 ameliorates sepsis?induced acute kidney injury via PTEN/AKT/mTOR?regulated autophagy
Authors: Z Sang, S Dong, P Zhang, Y Wei
Molecular Medicine Reports, 2021-07-30;24(4):.
Species: Mouse
Sample Types: Urine
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Rats with congenital hydronephrosis show increased susceptibility to renal ischemia-reperfusion injury
Authors: R Vilskersts, K Vilks, M Videja, H Cirule, O Zharkova-M, E Sevostjano, M Dambrova, E Liepinsh
Physiol Rep, 2020-11-01;8(22):e14638.
Species: Rat
Sample Types: Urine
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Fecal dysbiosis associated with colonic hypersensitivity and behavioral alterations in chronically Blastocystis-infected rats
Authors: M Defaye, C Nourrisson, E Baudu, A Lashermes, M Meynier, M Meleine, I Wawrzyniak, V Bonnin, J Barbier, B Chassaing, C Godfraind, A Gelot, N Barnich, D Ardid, M Bonnet, F Delbac, FA Carvalho, P Poirier
Sci Rep, 2020-06-04;10(1):9146.
Species: Rat
Sample Types: Tissue Homogenates
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Glomerular Collagen Deposition and Lipocalin-2 Expression Are Early Signs of Renal Injury in Prediabetic Obese Rats
Authors: EN Bukosza, T Kaucsár, M Godó, E Lajtár, P Tod, G Koncsos, ZV Varga, T Baranyai, NM Tu, H Schachner, C S?ti, P Ferdinandy, Z Giricz, G Szénási, P Hamar
Int J Mol Sci, 2019-08-30;20(17):.
Species: Rat
Sample Types: Plasma
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Hypoxia Induces Astrocyte-Derived Lipocalin-2 in Ischemic Stroke
Authors: F Ranjbar Ta, N Gasterich, M Scheld, R Weiskirche, C Beyer, T Clarner, A Zendedel
Int J Mol Sci, 2019-03-13;20(6):.
Species: Rat
Sample Types: Serum
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Renal injury after uninephrectomy in male and female intrauterine growth-restricted aged rats
Authors: AD Newsome, GK Davis, ON Adah, NB Ojeda, BT Alexander
PLoS ONE, 2019-03-07;14(3):e0213404.
Species: Rat
Sample Types: Urine
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Salt-Responsive Metabolite, ?-Hydroxybutyrate, Attenuates Hypertension
Authors: S Chakrabort, S Galla, X Cheng, JY Yeo, B Mell, V Singh, B Yeoh, P Saha, AV Mathew, M Vijay-Kuma, B Joe
Cell Rep, 2018-10-16;25(3):677-689.e4.
Species: Rat
Sample Types: Serum
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TGF-?-mediated NADPH oxidase 4-dependent oxidative stress promotes colistin-induced acute kidney injury
Authors: BY Jeong, SR Park, S Cho, SL Yu, HY Lee, CG Park, J Kang, DY Jung, MH Park, WM Hwang, SR Yun, JY Jung, SH Yoon
J. Antimicrob. Chemother., 2018-04-01;0(0):.
Species: Rat
Sample Types: Serum
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Neutrophil gelatinase-associated lipocalin level is a prognostic factor for survival in rat and human chronic liver diseases
Authors: K Yoshikawa, M Iwasa, A Eguchi, S Kojima, N Yoshizawa, M Tempaku, R Sugimoto, N Yamamoto, K Sugimoto, Y Kobayashi, H Hasegawa, Y Takei
Hepatol Commun, 2017-10-16;1(9):946-956.
Species: Rat
Sample Types: Plasma
Applications: ELISA Capture -
Insulin treatment prevents wounding associated changes in tissue and circulating neutrophil MMP-9 and NGAL in diabetic rats
Authors: M Abdollahi, TS Ng, A Rezaeizade, S Aamidor, SM Twigg, D Min, SV McLennan
PLoS ONE, 2017-02-09;12(2):e0170951.
Species: Rat
Sample Types: Tissue Secretion
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Cardioprotection and lifespan extension by the natural polyamine spermidine
Nat. Med., 2016-11-14;0(0):.
Species: Rat
Sample Types: Urine
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Chronic recurrent dehydration associated with periodic water intake exacerbates hypertension and promotes renal damage in male spontaneously hypertensive rats
Sci Rep, 2016-09-22;6(0):33855.
Species: Rat
Sample Types: Urine
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Oxidative/Nitrative Stress and Inflammation Drive Progression of Doxorubicin-Induced Renal Fibrosis in Rats as Revealed by Comparing a Normal and a Fibrosis-Resistant Rat Strain.
Authors: Szalay C, Erdelyi K, Kokeny G, Lajtar E, Godo M, Revesz C, Kaucsar T, Kiss N, Sarkozy M, Csont T, Krenacs T, Szenasi G, Pacher P, Hamar P
PLoS ONE, 2015-06-18;10(6):e0127090.
Species: Rat
Sample Types: Urine
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Modulation of early inflammatory response by different balanced and non-balanced colloids and crystalloids in a rodent model of endotoxemia.
Authors: Voigtsberger S, Urner M, Hasler M, Roth Z'Graggen B, Booy C, Spahn D, Beck-Schimmer B
PLoS ONE, 2014-04-07;9(4):e93863.
Species: Rat
Sample Types: Urine
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Intrarenal resistance index for the assessment of acute renal injury in a rat liver transplantation model.
Authors: Kong H, Chen F, He Y, Wu L, Wang L, Zhu S, Zheng S
BMC Nephrol, 2013-03-02;14(0):55.
Species: Rat
Sample Types: Serum
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