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Mouse gm csf

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Mouse GM-CSF is a recombinant mouse granulocyte-macrophage colony-stimulating factor (GM-CSF) protein. GM-CSF is a cytokine that regulates the production, differentiation, and function of granulocytes and macrophages.

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Lab products found in correlation

Mouse il 4, by R&D Systems (4 mentions) Fetal bovine serum (fbs), by Corning (2 mentions) Interleukin 6 (il 6), by R&D Systems (2 mentions) Cellstripper, by Corning (2 mentions) M csf, by R&D Systems (2 mentions) Recombinant human il 1β, by R&D Systems (1 mentions) 2 mercaptoethanol, by Thermo Fisher Scientific (1 mentions) L glutamine, by Thermo Fisher Scientific (1 mentions) Rpmi 1640, by Corning (1 mentions) Silac rpmi 1640, by Thermo Fisher Scientific (1 mentions) Penicillin streptomycin p s, by Thermo Fisher Scientific (1 mentions) L lysine hcl, by Merck Group (1 mentions) 40 μm cell strainer, by Greiner (1 mentions) Mcc950, by Bio-Techne (1 mentions) Imject alum, by Thermo Fisher Scientific (1 mentions) Cxcl1 elisa kit, by R&D Systems (1 mentions) Uric acid, by Merck Group (1 mentions) Cxcl2, by R&D Systems (1 mentions) Anti f4 80, by Bio-Rad (1 mentions) Anti cd206 af2535, by R&D Systems (1 mentions)

Close spelling variants of this product

GM-CSF (599 citations) Recombinant mouse GM-CSF (33 citations) Recombinant mouse granulocyte-macrophage colony-stimulating factor (GM-CSF) (7 citations) Mouse GM-CSF Quantikine ELISA kit (6 citations) Recombinant mouse GM-CSF and IL-4 (3 citations) Mouse GM-CSF Duo Set ELISA kit (2 citations) Mouse GM-CSF (415-ML) (2 citations) Recombinant mouse GM-CSF (415-ML) (2 citations)

13 protocols using mouse gm csf

1

Bone Marrow-Derived Macrophage Activation

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Bone marrow-derived macrophages were generated from whole bone marrow cells isolated from the femurs and tibias of control and Fasf/f, LysMcre mice followed by in vitro differentiation in RPMI 1640 media, supplemented with 10% (v/v) FBS, 20 ng/ml of mouse GM-CSF (R&D) and 0.1 % (v/v) of 2-mercaptoethanol (GIBCO/invitrogen), as described (18 (link)). At day 7-9, the in vitro differentiated macrophages were collected by gentle pipetting, washed with PBS and were allowed to rest for 1 hour prior to activation.
Recombinant human IL-1β was purchased from R&D. Recombinant N-terminal domain of gp96 was prepared and purified with extensive endotoxin removal procedures, as previously described (16 (link), 19 (link)). The endotoxin level in the recombinant gp96 used was below the level of detection by the limulus amebocyte lysate assay (16 (link)). Macrophages were seeded at 1×105/200 μl/well in 96 well cell culture plates. Cell activation was performed in RPMI 1640 medium supplemented with 10% FBS and 1μg/ml of polymyxin B. Macrophages were incubated with IL-1β or gp96 for 4 or 20 hours and the supernatants were harvested and examined for IL-10, IL-6 and CXCL5 by ELISA. The concentration of each cytokine was adjusted for cell number employing the MTT assay (O.D. 490 nm) at the time of terminating the activation.
Huang Q.Q., Birkett R., Koessler R.E., Cuda C.M., Kenneth Haines G., Jin J.P., Perlman H, & Pope R.M. (2014). Fas Signaling in Macrophages Promotes Chronicity in K/BxN Serum Transfer Induced Arthritis. Arthritis & rheumatology (Hoboken, N.J.), 66(1), 68-77.
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2

Bone Marrow Dendritic Cell Culture

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BMDC-RPMI: 10% fetal bovine serum (FBS, HyClone), 1% L-glutamine (Gibco), 1% Penicillin/Streptomycin (P/S, Gibco), 55μM 2-ME (Sigma), and 10μg/mL mouse GM-CSF (R&D Systems) in RPMI-1640 (Corning). Complete (C)-DMEM: 10% bovine calf serum (BCS, HyClone,) and 1% P/S in standard DMEM (Corning). L-ARG-free SILAC RPMI: 10% dialyzed FBS (Corning), 1% P/S, 219μM L-lysine-HCl (Sigma), and 55μM 2-ME in SILAC RPMI-1640 (Thermo Scientific).
Crowther R.R., Schmidt S.M., Lange S.M., McKell M.C., Robillard M.C., Zhao J., Haffey W.D., Wyder M.A., Greis K.D., Setchell K.D, & Qualls J.E. (2022). L-arginine transfer from antigen presenting cells sustains CD4+ T cell viability and proliferation. Journal of immunology (Baltimore, Md. : 1950), 208(4), 793-798.
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3

Isolation and Activation of Neonatal Microglia

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Neonatal microglia were isolated from C57BL/6 P1-4 mouse pups (Charles River, UK) as previously described (Carrillo-Jimenez et al., 2018 (link)). In brief, brains were dissected by removing meninges, cerebellum, and olfactory bulbs before tissue was finely minced with a scalpel then digested with papain solution for 20 min at 37 °C. Tissue was dissociated and the resulting cell suspension was passed through a 40 μm cell strainer (Greiner Bio-One). Cells were centrifuged at 300 g for 5 min then resuspended in culture medium (DMEM, 10% FBS, 1% PenStrep) and plated in T75 collagen-coated flasks (Greiner Bio-One). A full medium change was caried out on day 2 while a half medium was changed on day 5 in the presence of 5 ng/ml of mouse GM-CSF (R&D Systems). Microglia were shaken off between day 9 and 11 and plated onto poly-d-lysine coated 96-well plates (μclear, Greiner Bio-One) at a density of 20,000 cells/well. The following day cells were primed with LPS (100 ng/ml) for 3.5 h prior to treatment with C101248, MCC950 (Tocris), or vehicle (0.1% DMSO) for 30 min. NLRP3 was then activated by a complete medium change with an isotonic K+-free buffer (148 mM NaCl, 10 mM HEPES, 10 mM glucose, 2 mM CaCl2, 1 mM MgCl2 at pH 7.4) in the presence of test compounds or DMSO for 1 h after which supernatants were collected and stored at −20 °C until the measurement of IL-1β.
Ossola B., Rifat A., Rowland A., Hunter H., Drinkall S., Bender C., Hamlischer M., Teall M., Burley R., Barker D.F., Cadwalladr D., Dickson L., Lawrence J.M., Harvey J.R., Lizio M., Xu X., Kavanagh E., Cheung T., Sheardown S., Lawrence C.B., Harte M., Brough D., Madry C., Matthews K., Doyle K., Page K., Powell J., Brice N.L., Bürli R.W., Carlton M.B, & Dawson L.A. (2023). Characterisation of C101248: A novel selective THIK-1 channel inhibitor for the modulation of microglial NLRP3-inflammasome. Neuropharmacology, 224, 109330.
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4

Comprehensive ELISA Assays for Inflammatory Mediators

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Mouse M-CSF, mouse GM-CSF, human caspase-1, and mouse IL-1β, IL-6, CCL2, CXCL2, and CXCL1 ELISA kits were purchased from R&D Systems (Minneapolis, MN, USA). Uric acid and human IgG1 (hIgG) were obtained from Sigma-Aldrich (St. Louis, MO, USA). Anti-Ly6G Ab was purchased from BD Pharmingen (San Jose, CA, USA), anti-F4/80 was purchased from Serotec (Oxford, UK), anti-arginase 1 (#93668) was purchased from Cell Signaling Technology (Danvers, MA, USA), and anti-CD206 (AF2535) was purchased from R&D (Minneapolis, MN, USA). Silica (MIN-U-SIL-5; average particle diameter 1.7 μm) was obtained from US Silica Co. (Katy, TX, USA). Imject Alum was obtained from Thermo Scientific (Waltham, MA, USA).
Pan Y.G., Huang M.T., Sekar P., Huang D.Y., Lin W.W, & Hsieh S.L. (2021). Decoy Receptor 3 Inhibits Monosodium Urate-Induced NLRP3 Inflammasome Activation via Reduction of Reactive Oxygen Species Production and Lysosomal Rupture. Frontiers in Immunology, 12, 638676.
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5

Induction and Treatment of Monoarticular Arthritis

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Monoarticular arthritis was induced as before [5 (link), 8 (link), 40 (link)] by intra-articular (i.a.) injection of 100 μg mBSA in 10 μl saline into the right knee on day 0, the left knee being injected with saline, followed by a s.c. injection, in the scruff of the neck on days 0–2, of either mouse GM-CSF (500 ng, R&D Systems), mouse TNF (500 ng, R&D Systems), mouse CCL17 (600 ng, R&D Systems), or saline. Mice were sacrificed (day 7), and knee joints collected for histology [8 (link), 9 ].
For monoclonal antibody (mAb) administration, mice were given 150 μg anti-IL-23p19 (G23-8, eBiosceience™) or isotype (IgG1) control. For prophylactic administration of mAb, mice were i.p. injected on days − 2 and 0; for therapeutic administration of mAb, mice were i.p. injected after pain was evident (i.e. days 1 and 4 for mBSA/TNF and mBSA/GM-CSF models, respectively).
Lee K.M., Zhang Z., Achuthan A., Fleetwood A.J., Smith J.E., Hamilton J.A, & Cook A.D. (2020). IL-23 in arthritic and inflammatory pain development in mice. Arthritis Research & Therapy, 22, 123.
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6

Osteoclast Differentiation from Mouse BMMs

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Mouse BMMs were prepared using osteoclast precursors as described previously63 (link). BMMs were cultured in 24-well plates (7 × 104 cells/well) in the presence of M-CSF (50 ng/ml) in αMEM (Sigma-Aldrich) containing 10% fetal bovine serum (FBS) (JRH Biosciences, Lenexa, KS). The BMMs were further cultured in the presence or absence of GST-RANKL (100 ng/ml), mouse GM-CSF (10 ng/ml; R&D Systems), mouse IL-4 (10 ng/ml; R&D Systems), or mouse IFN-γ (20 ng/ml; Peprotech, Rock Hill, NJ) with M-CSF (50 ng/ml) for 24 h. Total RNA was collected 24 h after stimulation.
Koide M., Yamashita T., Murakami K., Uehara S., Nakamura K., Nakamura M., Matsushita M., Ara T., Yasuda H., Penninger J.M., Takahashi N., Udagawa N, & Kobayashi Y. (2020). Sclerostin expression in trabecular bone is downregulated by osteoclasts. Scientific Reports, 10, 13751.
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7

Generation of Murine Bone Marrow-Derived Dendritic Cells

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Bone marrow leukocytes were flushed from mouse tibia and femur and cultured in RPMI-1640 (Lonza, Allendale, NJ, USA) containing 10% fetal bovine serum (Invitrogen, Carlsbad, CA, USA), 1% antibiotic–antimycotic (Invitrogen) and supplemented with 20 ng/mL mouse GM-CSF (R&D Systems, Minneapolis, MN, USA) and 10 ng/mL mouse IL-4 (R&D Systems). Cells were cultured in a humidified chamber at 37°C and 5% atmospheric CO2. On day 3, half the culture medium was gently removed and replenished with fresh medium and cytokines. BMDC were harvested on day 6 with Cell Dissociation Buffer (Invitrogen). The immature BMDC were then treated with lipopolysaccharide (Sigma-Aldrich) or maturation cytokine cocktail comprising 10 ng/ml IL-1β (R&D Systems), 10 ng/ml TNF-α (R&D Systems), 15 ng/ml IL-6 (R&D Systems), and 1 µg/ml PGE2 (Sigma-Aldrich).
Liang D., Tian L., You R., Halpert M.M., Konduri V., Baig Y.C., Paust S., Kim D., Kim S., Jia F., Huang S., Zhang X., Kheradmand F., Corry D.B., Gilbert B.E., Levitt J.M, & Decker W.K. (2018). AIMp1 Potentiates TH1 Polarization and Is Critical for Effective Antitumor and Antiviral Immunity. Frontiers in Immunology, 8, 1801.
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8

Generation and Stimulation of Mouse BMDCs

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Mouse bone marrow was flushed from mouse femurs with cold RPMI-1640 culture medium (Corning, Manassas, VA, USA), and passed through a 70 μm cell strainer. The cells were centrifuged at 300 × g for 7 minutes at 4°C and counted with an automated veterinary cell counter (Heska, Loveland, CO, USA). 500,000 cells per well were seeded in flat bottom 96 well plates. Non-adherent cells were removed after 2 hours, and the wells were washed with PBS (Gibco, New York, NY, USA) and adherent cells were differentiated into BMDC with RPMI-1640 containing 10% FBS (Corning), 20 ng/mL mouse GM-CSF, and 20 ng/mL mouse IL-4 (R & D Systems). The differentiation medium was refreshed every other day. After 6 days the cells were treated with RPMI or mouse or human recombinant IL-38 (precursor or 3–152) for one hour before the cells were stimulated with 10 ng/mL lipopolysaccharide (LPS; O55:B5, Sigma-Aldrich, St. Louis, MO, USA). After 24 hours, the supernatants were collected and stored at −20°C until ELISA was performed.
de Graaf D.M., Maas R.J., Smeekens S.P., Eisenmesser E., Redzic J.S., Helsen M.M., Powers N.E., Li S., Kalabokis V., Gresnigt M.S., Joosten L.A., Dinarello C.A, & van de Veerdonk F.L. (2020). Human recombinant interleukin-38 suppresses inflammation in mouse models of local and systemic disease. Cytokine, 137, 155334.
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9

Isolation of Mouse Lung APC and Spleen CD4 T Cells

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Mouse lung or spleen single-cell suspensions were prepared by mincing whole organs through a 40-μm cell strainer (BD Falcon, San Jose, CA) followed by red blood cell (RBC) lysis (ACK lysis buffer, Sigma–Aldrich) for 3 min. For isolation of lung APCs, RBC-free whole lung cells were labeled with anti-CD11c-conjugated magnetic beads (Miltenyi Biotec, San Diego, CA) and then isolated by autoMACS (Miltenyi Biotec). For isolation of spleen CD4 T cells, RBC-free whole splenocytes were labeled with anti-CD4 conjugated magnetic beads (Miltenyi Biotec) and then isolated by autoMACS. Mouse BMDCs were prepared as previously described with some modification (Lutz et al., 1999 (link)) Femurs and tibias of 4- to 8-week-old female were isolated and freed from the surrounding tissue. Intact bones were kept in 70% ethanol for 3 min followed by a PBS wash. Both ends of the bones were cut with scissors, and the marrow was flushed out with RPMI-1640 medium through a syringe with 26.5 needle. RBCs were then removed by ACK lysis buffer and cell debris or tissue clusters were filtered out. Cells from bone marrow were cultured in a 6-well plate with 20 ng/ml mouse GM-CSF and 10 ng/ml mouse IL-4 (R&D Systems, Minneapolis, MN) for 5 to 6 days.
You R., Lu W., Shan M., Berlin J.M., Samuel E.L., Marcano D.C., Sun Z., Sikkema W.K., Yuan X., Song L., Hendrix A.Y., Tour J.M., Corry D.B, & Kheradmand F. (2015). Nanoparticulate carbon black in cigarette smoke induces DNA cleavage and Th17-mediated emphysema. eLife, 4, e09623.
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10

Generating Activated Mouse Bone Marrow-Derived Macrophages

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BMDMs were prepared from bone marrow extracted from the tibia and femur of WT C57BL/6J mice and cultured in non–tissue culture–treated Petri dishes in high glucose DMEM medium containing 10% FBS, 100 U/mL penicillin, 100 μg/mL streptomycin, and supplemented with 10 ng/mL mouse GM-CSF and M-CSF (R&D Systems). After 7 days of culture, BMDMs were activated with 100 U/mL EB lipopolysaccharide (LPS; Invivogen) for 24 hours (M1 polarization). After washing the monolayer, cells were detached after 10 minutes incubation in Cellstripper (Corning) using cell lifters. BMDMs in suspension were loaded with 100 ng/mL SIINFEKL peptide at 37°C for 1 hour and washed before being counted and injected in 1X PBS into mice as indicated below.
Cunha P.P., Bargiela D., Minogue E., Krause L.C., Barbieri L., Brombach C., Gojkovic M., Marklund E., Pietsch S., Foskolou I., Branco C.M., Veliça P, & Johnson R.S. (2022). Infiltration of Tumors Is Regulated by T cell–Intrinsic Nitric Oxide Synthesis. Cancer Immunology Research, 11(3), 351-363.
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