Human GM-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 GM-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.
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: GM-CSF
GM-CSF (Granulocyte-Macrophage Colony Stimulating Factor) induces the development of monocytes, neutrophils, eosinophils, and myeloid and dermal dendritic cells. It also acts as a neutrophil and dendritic cell chemoattractant. GM-CSF promotes Th1 and Th17 cell mediated autoimmune inflammation as well as the inflammatory activation of dendritic cells, microglia, alveolar macrophages, and eosinophils. In addition, it cooperates with G-CSF in promoting tumor cell proliferation and invasion. GM-CSF signals through a receptor complex composed of GM-CSF R alpha and the Common beta Chain (beta c) with Syndecan-2 as a potential co-receptor. The beta c subunit is shared by the receptor complexes for IL-3 and IL-5.
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 GM-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.
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Citations: Showing 1 - 10
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Ventilatory support and inflammatory peptides in hospitalised patients with COVID-19: A prospective cohort trial
Authors: Gysan, MR;Milacek, C;Bal, C;Zech, A;Brugger, J;Milos, RI;Antoniewicz, L;Idzko, M;Gompelmann, D;
PloS one
Species: Human
Sample Types: Serum
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Mast cells disrupt the function of the esophageal epithelial barrier
Authors: Kleuskens, MTA;Bek, MK;Al Halabi, Y;Blokhuis, BRJ;Diks, MAP;Haasnoot, ML;Garssen, J;Bredenoord, AJ;C A M van Esch, B;Redegeld, FA;
Mucosal immunology
Species: Human
Sample Types: Cell Culture Supernates
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A Reconstructed Human Melanoma-in-Skin Model to Study Immune Modulatory and Angiogenic Mechanisms Facilitating Initial Melanoma Growth and Invasion
Authors: Michielon, E;López González, M;Stolk, DA;Stolwijk, JGC;Roffel, S;Waaijman, T;Lougheed, SM;de Gruijl, TD;Gibbs, S;
Cancers
Species: Human
Sample Types: Cell Culture Supernates
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Microbial peptides activate tumour-infiltrating lymphocytes in glioblastoma
Authors: Naghavian, R;Faigle, W;Oldrati, P;Wang, J;Toussaint, NC;Qiu, Y;Medici, G;Wacker, M;Freudenmann, LK;Bont�, PE;Weller, M;Regli, L;Amigorena, S;Rammensee, HG;Walz, JS;Brugger, SD;Mohme, M;Zhao, Y;Sospedra, M;Neidert, MC;Martin, R;
Nature
Species: Human
Sample Types: Cell Culture Supernates
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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
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Mast cell tolerance in the skin microenvironment to commensal bacteria is controlled by fibroblasts
Authors: A Di Nardo, YL Chang, S Alimohamma, K Masuda-Kur, Z Wang, K Sriram, PA Insel
Cell Reports, 2023-04-28;42(5):112453.
Species: Human
Sample Types:
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During early stages of cancer, neutrophils initiate anti-tumor immune responses in tumor-draining lymph nodes
Authors: E Pylaeva, G Korschunow, I Spyra, S Bordbari, E Siakaeva, I Ozel, M Domnich, A Squire, A Hasenberg, K Thangavelu, T Hussain, M Goetz, KS Lang, M Gunzer, W Hansen, J Buer, A Bankfalvi, S Lang, J Jablonska
Cell Reports, 2022-08-16;40(7):111171.
Species: Human
Sample Types: Cell Culture Supernates
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Repeated exposure of bronchial epithelial cells to particular matter increases allergen-induced cytokine release and permeability
Authors: H Janbazacya, J van Bergen, S Varasteh, J Garssen, G Folkerts, S Braber
Cytokine, 2022-04-08;154(0):155878.
Species: Human
Sample Types: Cell Culture Supernates
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CRISPR/Cas9-mediated TGFbetaRII disruption enhances anti-tumor efficacy of human chimeric antigen receptor T cells in vitro
Authors: K Alishah, M Birtel, E Masoumi, L Jafarzadeh, HR Mirzaee, J Hadjati, RH Voss, M Diken, S Asad
Journal of Translational Medicine, 2021-11-27;19(1):482.
Species: Human
Sample Types: Cell Culture Supernates
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IL-6 promotes drug resistance through formation of polyploid giant cancer cells and stromal fibroblast reprogramming
Authors: N Niu, J Yao, RC Bast, AK Sood, J Liu
Oncogenesis, 2021-09-29;10(9):65.
Species: Human
Sample Types: Cell Culture Supernates
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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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Biomarkers for Comorbidities Modulate the Activity of T-Cells in COPD
Authors: K Jamal Jame, WJ Gallert, SD Yanik, S Panek, J Kronsbein, D Jungck, A Koch, J Knobloch
International Journal of Molecular Sciences, 2021-07-02;22(13):.
Species: Human
Sample Types: Cell Culture Supernates
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Bacterial genotoxins induce T�cell senescence
Authors: SL Mathiasen, L Gall-Mas, IS Pateras, SDP Theodorou, MRJ Namini, MB Hansen, OCB Martin, CK Vadivel, K Ntostoglou, D Butter, M Givskov, C Geisler, AN Akbar, VG Gorgoulis, T Frisan, N Ødum, T Krejsgaard
Cell Reports, 2021-06-08;35(10):109220.
Species: Human
Sample Types: Cell Culture Supernates
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Severe T cell hyporeactivity in ventilated COVID-19 patients correlates with prolonged virus persistence and poor outcomes
Authors: K Renner, T Schwittay, S Chaabane, J Gottschlin, C Müller, C Tiefenböck, JN Salewski, F Winter, S Buchtler, S Balam, MV Malferthei, M Lubnow, D Lunz, B Graf, F Hitzenbich, F Hanses, H Poeck, M Kreutz, E Orsó, R Burkhardt, T Niedermair, C Brochhause, A Gessner, B Salzberger, M Mack
Nature Communications, 2021-05-21;12(1):3006.
Species: Human
Sample Types: Cell Culture Supernates
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Gain-of-function variants in SYK cause immune dysregulation and systemic inflammation in humans and mice
Authors: L Wang, D Aschenbren, Z Zeng, X Cao, D Mayr, M Mehta, M Capitani, N Warner, J Pan, L Wang, Q Li, T Zuo, S Cohen-Keda, J Lu, RC Ardy, DJ Mulder, D Dissanayak, K Peng, Z Huang, X Li, Y Wang, X Wang, S Li, S Bullers, AN Gammage, K Warnatz, AI Schiefer, G Krivan, V Goda, WHA Kahr, M Lemaire, Genomics E, CY Lu, I Siddiqui, MG Surette, D Kotlarz, KR Engelhardt, HR Griffin, R Rottapel, H Decaluwe, RM Laxer, M Proietti, S Hambleton, S Elcombe, CH Guo, B Grimbacher, I Dotan, SC Ng, SA Freeman, SB Snapper, C Klein, K Boztug, Y Huang, D Li, HH Uhlig, AM Muise
Nature Genetics, 2021-03-29;53(4):500-510.
Species: Human
Sample Types: Cell Culture Supernates
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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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A new cytokine-based dynamic stratification during induction is highly predictive of survivals in acute myeloid leukemia
Authors: P Peterlin, J Gaschet, T Guillaume, A Garnier, M Eveillard, A Le Bourgeo, M Cherel, C Debord, Y Le Bris, O Theisen, C Godon, B Mahé, V Dubruille, S Wuilleme, C Touzeau, T Gastinne, N Blin, A Lok, B Tessoulin, S Le Gouill, P Moreau, MC Béné, P Chevallier
Cancer Medicine, 2020-12-25;0(0):.
Species: Human
Sample Types: Serum
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Altered plasma levels of &betaC and &gammaC chain cytokines and post-treatment modulation in tuberculous lymphadenitis
Authors: GR Kathamuthu, K Moideen, R Sridhar, D Baskaran, S Babu
Cytokine, 2020-12-17;138(0):155405.
Species: Human
Sample Types: Plasma
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T helper cells in synovial fluid of patients with rheumatoid arthritis primarily have a Th1 and a CXCR3+Th2 phenotype
Authors: J Aldridge, AH Ekwall, L Mark, B Bergström, K Andersson, I Gjertsson, AC Lundell, A Rudin
Arthritis Res Ther, 2020-10-16;22(1):245.
Species: Human
Sample Types: Plasma
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LPS Induces GM-CSF Production by Breast Cancer MDA-MB-231 Cells via Long-Chain Acyl-CoA Synthetase 1
Authors: F Al-Rashed, R Thomas, A Al-Roub, F Al-Mulla, R Ahmad
Molecules, 2020-10-14;25(20):.
Species: Human
Sample Types: Cell Culture Supernates
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The human glomerular endothelial cells are potent pro-inflammatory contributors in an in vitro model of lupus nephritis
Authors: P Dimou, RD Wright, KL Budge, A Midgley, SC Satchell, M Peak, MW Beresford
Sci Rep, 2019-06-06;9(1):8348.
Species: Human
Sample Types: Cell Culture Supernates
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Historical BCG vaccination combined with drug treatment enhances inhibition of mycobacterial growth ex vivo in human peripheral blood cells
Authors: SA Prabowo, A Zelmer, L Stockdale, U Ojha, SG Smith, K Seifert, HA Fletcher
Sci Rep, 2019-03-19;9(1):4842.
Species: Human
Sample Types: Cell Culture Supernates
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Interferon-gamma inhibits seminal plasma induction of colony stimulating factor 2 in mouse and human reproductive tract epithelial cells
Authors: DJ Sharkey, DJ Glynn, JE Schjenken, KP Tremellen, SA Robertson
Biol. Reprod., 2018-09-01;0(0):.
Species: Human
Sample Types: Cell Culture Supernates
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Inhibition of IRE1 RNase activity modulates the tumor cell secretome and enhances response to chemotherapy
Authors: SE Logue, EP McGrath, P Cleary, S Greene, K Mnich, A Almanza, E Chevet, RM Dwyer, A Oommen, P Legembre, F Godey, EC Madden, B Leuzzi, J Obacz, Q Zeng, JB Patterson, R Jäger, AM Gorman, A Samali
Nat Commun, 2018-08-15;9(1):3267.
Species: Human
Sample Types: Cell Culture Supernates
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Cross-Tissue Transcriptomic Analysis of Human Secondary Lymphoid Organ-Residing ILC3s Reveals a Quiescent State in the Absence of Inflammation
Authors: YE Bar-Ephrai, F Cornelisse, N Papazian, T Konijn, RM Hoogenboez, MA Sanders, BA Westerman, M Gönültas, J Kwekkeboom, JMM Den Haan, RM Reijmers, RE Mebius, T Cupedo
Cell Rep, 2017-10-17;21(3):823-833.
Species: Human
Sample Types: Cell Culture Supernates
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A microfluidics assay to study invasion of human placental trophoblast cells
Authors: Y Abbas, CM Oefner, WJ Polacheck, L Gardner, L Farrell, A Sharkey, R Kamm, A Moffett, ML Oyen
J R Soc Interface, 2017-05-01;14(130):.
Species: Human
Sample Types: Cell Culture Supernates
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Caspase-8 Acts in a Non-enzymatic Role as a Scaffold for Assembly of a Pro-inflammatory FADDosome Complex upon TRAIL Stimulation
Authors: CM Henry, SJ Martin
Mol. Cell, 2017-02-16;65(4):715-729.e5.
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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CREB regulates TNF-?-induced GM-CSF secretion via p38 MAPK in human lung fibroblasts
Authors: Y Koga, T Hisada, T Ishizuka, M Utsugi, A Ono, M Yatomi, Y Kamide, H Aoki-Saito, H Tsurumaki, K Dobashi, M Yamada
Allergol Int, 2016-04-22;0(0):.
Species: Human
Sample Types: Cell Culture Supernates
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Chronic Endotoxemia in Subjects with Type-1 Diabetes Is Seen Much before the Onset of Microvascular Complications.
Authors: Aravindhan V, Mohan V, Arunkumar N, Sandhya S, Babu S
PLoS ONE, 2015-09-14;10(9):e0137618.
Species: Human
Sample Types: Serum
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Roles for the mitogen-activated protein kinase (MAPK) phosphatase, DUSP1, in feedback control of inflammatory gene expression and repression by dexamethasone.
Authors: Shah S, King E, Chandrasekhar A, Newton R
J Biol Chem, 2014-04-01;289(19):13667-79.
Species: Human
Sample Types: Cell Culture Supernates
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Activin A as a mediator of NK-dendritic cell functional interactions.
Authors: Seeger P, Bosisio D, Parolini S, Badolato R, Gismondi A, Santoni A, Sozzani S
J Immunol, 2014-01-06;192(3):1241-8.
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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Leptin up-regulates TLR2 in human monocytes.
Authors: Jaedicke K, Roythorne A, Padget K, Todryk S, Preshaw P, Taylor J
J Leukoc Biol, 2013-01-22;93(4):561-71.
Species: Human
Sample Types: Cell Culture Supernates
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Interleukin-1alpha controls allergic sensitization to inhaled house dust mite via the epithelial release of GM-CSF and IL-33.
J. Exp. Med., 2012-07-16;209(8):1505-17.
Species: Human
Sample Types: Cell Culture Supernates
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ERK5 protein promotes, whereas MEK1 protein differentially regulates, the Toll-like receptor 2 protein-dependent activation of human endothelial cells and monocytes.
Authors: Wilhelmsen K, Mesa K, Lucero J, Xu F, Hellman J
J Biol Chem, 2012-06-15;287(32):26478-94.
Species: Human
Sample Types: Cell Culture Supernates
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The role of IL-33 and its receptor ST2 in human nasal epithelium with allergic rhinitis.
Authors: Kamekura R, Kojima T, Takano K, Go M, Sawada N, Himi T
Clin. Exp. Allergy, 2012-02-01;42(2):218-28.
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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Functional regulatory T cells accumulate in aged hosts and promote chronic infectious disease reactivation.
Authors: Lages CS, Suffia I, Velilla PA, Huang B, Warshaw G, Hildeman DA, Belkaid Y, Chougnet C
J. Immunol., 2008-08-01;181(3):1835-48.
Species: Mouse
Sample Types: Cell Culture Supernates
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Fungal proteases induce Th2 polarization through limited dendritic cell maturation and reduced production of IL-12.
Authors: Lamhamedi-Cherradi SE, Martin RE, Ito T, Kheradmand F, Corry DB, Liu YJ, Moyle M
J. Immunol., 2008-05-01;180(9):6000-9.
Species: Human
Sample Types: Cell Culture Supernates
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Human CD4(+) T cells recognize an epitope within alpha-fetoprotein sequence and develop into TGF-beta-producing CD4(+) T cells.
Authors: Alisa A, Boswell S, Pathan AA, Ayaru L, Williams R, Behboudi S
J. Immunol., 2008-04-01;180(7):5109-17.
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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Expression and role of EGFR ligands induced in airway cells by PM2.5 and its components.
Authors: Rumelhard M, Ramgolam K, Hamel R, Marano F, Baeza-Squiban A
Eur. Respir. J., 2007-09-05;30(6):1064-73.
Species: Human
Sample Types: Cell Culture Supernates
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B and T lymphocytes are the primary sources of RANKL in the bone resorptive lesion of periodontal disease.
Authors: Kawai T, Matsuyama T, Hosokawa Y, Makihira S, Seki M, Karimbux NY, Goncalves RB, Valverde P, Dibart S, Li YP, Miranda LA, Ernst CW, Izumi Y, Taubman MA
Am. J. Pathol., 2006-09-01;169(3):987-98.
Species: Human
Sample Types: Tissue Homogenates
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Differential survival of leukocyte subsets mediated by synovial, bone marrow, and skin fibroblasts: site-specific versus activation-dependent survival of T cells and neutrophils.
Authors: Filer A, Parsonage G, Smith E, Osborne C, Thomas AM, Curnow SJ, Rainger GE, Raza K, Nash GB, Lord J, Salmon M, Buckley CD
Arthritis Rheum., 2006-07-01;54(7):2096-108.
Species: Human
Sample Types: Cell Culture Supernates
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Activation of resting human primary T cells with chimeric receptors: costimulation from CD28, inducible costimulator, CD134, and CD137 in series with signals from the TCR zeta chain.
Authors: Finney HM, Akbar AN, Lawson AD
J. Immunol., 2004-01-01;172(1):104-13.
Species: Human
Sample Types: Cell Culture Supernates
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Adherence influences monocyte responsiveness to interleukin-10.
Authors: Petit-Bertron AF, Fitting C, Cavaillon JM, Adib-Conquy M
J. Leukoc. Biol., 2003-01-01;73(1):145-54.
Species: Human
Sample Types: Cell Culture Supernates
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The ELISA worked well. Easy to prepare it.
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