Human G-CSF DuoSet ELISA
Human G-CSF 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 human G-CSF. 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.
Normal Goat Serum: (Catalog # DY005)
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)
Normal Goat Serum: (Catalog # DY005)
Scientific Data
Product Datasheets
Preparation and Storage
Background: G-CSF
Granulocyte-colony stimulating factor (G-CSF) is a 24-25 kDa monomeric glycoprotein that regulates the proliferation, differentiation, and activation of hematopoietic cells in the neutrophilic granulocyte lineage. Mature human G-CSF is a 178 amino acid (aa) O-glycosylated protein that contains two intrachain disulfide bridges. In humans, alternate splicing generates a second minor isoform with a 3 aa deletion. Mouse and human G-CSF share 76% aa sequence identity, and the two proteins show species cross-reactivity. G-CSF is produced by activated monocytes and macrophages, fibroblasts, endothelial cells, astrocytes, neurons, and bone marrow stroma cells. In addition, various tumor cells express G-CSF constitutively.
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 with NGS, 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 G-CSF 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.
19
Citations: Showing 1 - 10
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c-Met Mediated Cytokine Network Promotes Brain Metastasis of Breast Cancer by Remodeling Neutrophil Activities
Authors: Liu, Y;Smith, MR;Wang, Y;D'Agostino, R;Ruiz, J;Lycan, T;Kucera, GL;Miller, LD;Li, W;Chan, MD;Farris, M;Su, J;Song, Q;Zhao, D;Chandrasekaran, A;Xing, F;
Cancers
Species: Human
Sample Types: Cell Culture Supernates
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DRP1-Mediated Mitochondrial Fission Regulates Lung Epithelial Response to Allergen
Authors: SR Bruno, A Kumar, ZF Mark, R Chandrasek, E Nakada, N Chamberlai, B Mihavics, J Walzer, J Cahoon, AE Dixon, B Cunniff, V Anathy
International Journal of Molecular Sciences, 2021-10-15;22(20):.
Species: Human
Sample Types: Cell Culture Supernates
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Neutrophil function and bactericidal activity against Staphylococcus aureus after cardiac surgery with cardiopulmonary bypass
Authors: M Lesouhaiti, M Gregoire, A Gacouin, V Coirier, A Frerou, C Piau, V Cattoir, E Dumontet, M Revest, P Tattevin, A Roisne, JP Verhoye, E Flecher, Y Le Tulzo, K Tarte, JM Tadié
Journal of leukocyte biology, 2021-08-23;0(0):.
Species: Human
Sample Types: Plasma
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Characterization of low-density granulocytes in COVID-19
Authors: LE Cabrera, PT Pekkarinen, M Alander, KHA Nowlan, NA Nguyen, S Jokiranta, S Kuivanen, A Patjas, S Mero, SH Pakkanen, S Heinonen, A Kantele, O Vapalahti, E Kekäläinen, T Strandin
PloS Pathogens, 2021-07-06;17(7):e1009721.
Species: Human
Sample Types: Serum
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The dynamics of human bone marrow adipose tissue in response to feeding and fasting
Authors: PK Fazeli, MA Bredella, OG Pachon-Peñ, W Zhao, X Zhang, AT Faje, M Resulaj, SP Polineni, TM Holmes, H Lee, EK O'Donnell, OA MacDougald, MC Horowitz, CJ Rosen, A Klibanski
JCI Insight, 2021-06-22;0(0):.
Species: Human
Sample Types: Serum
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A digital single-molecule nanopillar SERS platform for predicting and monitoring immune toxicities in immunotherapy
Authors: J Li, A Wuethrich, AAI Sina, HH Cheng, Y Wang, A Behren, PN Mainwaring, M Trau
Nature Communications, 2021-02-17;12(1):1087.
Species: Human
Sample Types: Serum
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Gene expression network analyses during infection with virulent and avirulent Trypanosoma cruzi�strains unveil a role for fibroblasts in neutrophil recruitment and activation
Authors: AER Oliveira, MCA Pereira, AT Belew, LRP Ferreira, LMN Pereira, EGA Neves, MDCP Nunes, BA Burleigh, WO Dutra, NM El-Sayed, RT Gazzinelli, SMR Teixeira
PLoS Pathog., 2020-08-18;16(8):e1008781.
Species: Human
Sample Types: Cell Culture Supernates
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Interleukin (IL)-17/IL-36 axis participates to the crosstalk between endothelial cells and keratinocytes during inflammatory skin responses
Authors: L Mercurio, CM Failla, L Capriotti, C Scarponi, F Facchiano, M Morelli, S Rossi, G Pagnanelli, C Albanesi, A Cavani, S Madonna
PLoS ONE, 2020-04-30;15(4):e0222969.
Species: Human
Sample Types: Cell Culture Supernates
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Probiotic Lactobacillus rhamnosus GR-1 is a unique prophylactic agent that suppresses infection-induced myometrial cell responses
Authors: B Kim, O Shynlova, S Lye
Sci Rep, 2019-03-18;9(1):4698.
Species: Human
Sample Types: Cell Culture Supernates
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Regulation of CXCL1 chemokine and CSF3 cytokine levels in myometrial cells by the MAFF transcription factor
Authors: J Saliba, B Coutaud, V Solovieva, F Lu, V Blank
J. Cell. Mol. Med., 2019-01-22;0(0):.
Species: Human
Sample Types: Cell Culture Supernates
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Interleukin-1 primes human mesenchymal stem cells towards an anti-inflammatory and pro-trophic phenotype in vitro
Authors: E Redondo-Ca, C Cunningham, J Miller, L Martuscell, S Aoulad-Ali, NJ Rothwell, CM Kielty, SM Allan, E Pinteaux
Stem Cell Res Ther, 2017-04-17;8(1):79.
Species: Human
Sample Types: Cell Culture Supernates
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IL-17 Promotes Neutrophil-Mediated Immunity by Activating Microvascular Pericytes and Not Endothelium
J Immunol, 2016-08-17;0(0):.
Species: Human
Sample Types: Cell Culture Supernates
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Opposite cytokine synthesis by fibroblasts in contact co-culture with osteosarcoma cells compared with transwell co-cultures.
Authors: David M, Kelly E, Zoellner H
Cytokine, 2013-03-19;62(1):48-51.
Species: Human
Sample Types: Cell Culture Supernates
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IL-17C regulates the innate immune function of epithelial cells in an autocrine manner.
Authors: Ramirez-Carrozzi V, Sambandam A, Luis E
Nat. Immunol., 2011-10-12;12(12):1159-66.
Species: Human
Sample Types: Cell Culture Supernates
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Novel markers of inflammation identified in tumor necrosis factor receptor-associated periodic syndrome (TRAPS) by transcriptomic analysis of effects of TRAPS-associated tumor necrosis factor receptor type I mutations in an endothelial cell line.
Authors: Rebelo SL, Amel-Kashipaz MR, Radford PM, Bainbridge SE, Fiets R, Fang J, McDermott EM, Powell RJ, Todd I, Tighe PJ
Arthritis Rheum., 2009-01-01;60(1):269-80.
Species: Human
Sample Types: Cell Culture Supernates
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Human epithelial cells establish direct antifungal defense through TLR4-mediated signaling.
Authors: Weindl G, Naglik JR, Kaesler S, Biedermann T, Hube B, Korting HC, Schaller M
J. Clin. Invest., 2007-12-01;117(12):3664-3672.
Species: Human
Sample Types: Cell Culture Supernates
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Role of mesothelial cell-derived granulocyte colony-stimulating factor in interleukin-17-induced neutrophil accumulation in the peritoneum.
Authors: Witowski J, Ksiazek K, Warnecke C, Kuzlan M, Korybalska K, Tayama H, Wisniewska-Elnur J, Pawlaczyk K, Trominska J, Breborowicz A, Jorres A
Kidney Int., 2007-01-17;71(6):514-25.
Species: Human
Sample Types: Cell Culture Supernates
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Prostanoid receptor expression by human airway smooth muscle cells and regulation of the secretion of granulocyte colony-stimulating factor.
Authors: Clarke DL, Belvisi MG, Smith SJ, Hardaker E, Yacoub MH, Meja KK, Newton R, Slater DM, Giembycz MA
Am. J. Physiol. Lung Cell Mol. Physiol., 2005-02-01;288(2):L238-50.
Species: Human
Sample Types: Cell Culture Supernates
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Toxoplasma gondii induces granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor secretion by human fibroblasts: implications for neutrophil apoptosis.
Authors: Channon JY, Miselis KA, Minns LA, Dutta C, Kasper LH
Infect. Immun., 2002-11-01;70(11):6048-57.
Species: Human
Sample Types: Cell Culture Supernates
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