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Recombinant murine g csf

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Recombinant murine G-CSF is a laboratory reagent that functions as a cytokine. It is produced using recombinant DNA technology and is derived from the mouse organism.

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Lab products found in correlation

Recombinant murine gm csf, by Thermo Fisher Scientific (2 mentions) Recombinant murine il 3, by Thermo Fisher Scientific (2 mentions) β estradiol, by Merck Group (2 mentions) Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions) Trypsin, by Harvard Bioscience (1 mentions) Penicilline streptomycin, by Harvard Bioscience (1 mentions) Glucose go assay kit, by Merck Group (1 mentions) Glycolysis cell based assay kit, by Cayman Chemical (1 mentions) Pierce bca protein assay kit, by Merck Group (1 mentions) Lipopolysaccharide lps, by Merck Group (1 mentions) Clodronate liposomes, by Merck Group (1 mentions) C57bl 6j mice, by Daehan Biolink (1 mentions) Amd3100, by R&D Systems (1 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions) Essential 8 medium, by Thermo Fisher Scientific (1 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions) Interleukin 3 (il 3), by Thermo Fisher Scientific (1 mentions) Penicillin, by Atlanta Biologicals (1 mentions) Streptomycin, by Atlanta Biologicals (1 mentions) Dmem f12 medium, by Thermo Fisher Scientific (1 mentions)

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G-CSF (149 citations) Recombinant G-CSF (7 citations) Murine G-CSF (6 citations)

9 protocols using recombinant murine g csf

1

In-vitro Generation of Myeloid-Derived Suppressor Cells

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in-vitro generation of MDSC was performed according to previously established protocols (25 (link), 26 (link)). For in-vitro generation of MDSC non-pregnant WT and myeloid HIF-KO mice were euthanized and femora removed. Bone marrow was collected by rinsing the bones with PBS with a syringe and a 25G needle. Bone marrow cells were then washed, adjusted to 3x105 cells/ml and cultured for 72 h at 37°C in culture medium [Dulbecco's modified eagle medium, DMEM (Thermo Fisher Scientific, Darmstadt, Germany), supplemented with 10% fetal calf serum (FCS, Biochrom, Berlin, Germany) and 1% Penicilline/Streptomycin (Biochrom, Berlin, Germany)] supplemented with 100 ng/ml recombinant murine G-CSF (Peprotech, Hamburg, Germany) and 12.5 ng/ml recombinant murine GM-CSF (Peprotech, Hamburg, Germany). After 72 h of culture non-adherent cells were removed and adherent MDSC detached with trypsin (Biochrom GmbH, Berlin, Germany). >90% of cells were Gr-1+/CD11b+ as determined by flow cytometry, thereby exhibiting surface characteristics of MDSC.
Köstlin-Gille N., Dietz S., Schwarz J., Spring B., Pauluschke-Fröhlich J., Poets C.F, & Gille C. (2019). HIF-1α-Deficiency in Myeloid Cells Leads to a Disturbed Accumulation of Myeloid Derived Suppressor Cells (MDSC) During Pregnancy and to an Increased Abortion Rate in Mice. Frontiers in Immunology, 10, 161.
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2

Glycolysis Profiling of Atg7 KO Cells

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32D CRISPR-Atg7–/– and Atg7+/+ clones were cultured in RPMI 1640 (containing 2mg/mL Glucose) with 1% FCS, 2 mM L-glutamine, 100 U/ml Pen-Strep. In order to induce neutrophil differentiation, 1x106 cells/ml were seeded in 96-well plates with medium supplemented with 100 ng/ml recombinant murine G-CSF (peprotech). At day 3, medium was supplemented with fresh 100 ng/mL G-CSF. Cells and supernatant were harvested at indicated time points and supernatants analyzed for glucose levels and L-lactate levels following the Glucose (GO) Assay Kit (GAGO-20, Sigma) and the Glycolysis Cell-Based Assay Kit (#600450, Cayman Chemicals). Cells were lysed in NP40 buffer and protein was quantified using Pierce BCA Protein Assay Kit (Sigma). Glucose consumption and L-lactate levels were normalized to protein levels for each well individually. Each assay representative of at least two independent experimental repeats.
Riffelmacher T., Clarke A., Richter F.C., Stranks A., Pandey S., Danielli S., Hublitz P., Yu Z., Johnson E., Schwerd T., McCullagh J., Uhlig H., Jacobsen S.E, & Simon A.K. (2017). Autophagy-Dependent Generation of Free Fatty Acids Is Critical for Normal Neutrophil Differentiation. Immunity, 47(3), 466-480.e5.
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3

Induction of Granulopoiesis in Mice

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C57BL/6 J mice (gender and age-matched, 6–8 weeks) were purchased from Daehan Biolink (Korea). Lyz2-cre (Lyz2tm1(cre)Ifo) and ROSA-eYFP (Gt(ROSA)26Sortm1(EYFP)Cos) were generous gifts from Drs. R.M. Locksley and C.A. Lowell (UCSF). S100A8-cre (Tg(S100A8-cre,-EGFP)1Ilw/J)75 (link) were generously provided by Drs. I.L. Weissman (Stanford) and G.O. Ahn (Postech). For induction of demand-adapted granulopoiesis, 2.5 μg recombinant murine G-CSF (Peprotech), 20 μg lipopolysaccharide (LPS; Sigma), or 1 mg clodronate-liposomes (Clodronate Liposomes) was intravenously injected. Live L. monocytogenes 10403 S was injected into the tail vein and mouse footpads at 103 and 104 cfu per injection, respectively. All animal experiments were performed in accordance with guidelines and regulations for rodent experiments provided by the Institutional Animal Care and Use Committee (IACUC) of KAIST. The protocols for the study were approved by KAIST IACUC (KA2010-21, KA2014-09).
Kim M.H., Yang D., Kim M., Kim S.Y., Kim D, & Kang S.J. (2017). A late-lineage murine neutrophil precursor population exhibits dynamic changes during demand-adapted granulopoiesis. Scientific Reports, 7, 39804.
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4

B cell depletion in inhibitor mice

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Anti-CD20 IgG2a antibody (clone 18B12, kindly provided by Biogen Idec), AMD3100 (R & D system, USA) and recombinant murine G-CSF (PeproTech, Rocky Hill, NJ) were administered two weeks per cycle for 3 cycles to deplete B cells in inhibitor mice. Anti-CD20 was given at 250 μg/mouse three i.v. doses 14 days apart; AMD3100 (plerixafor; Mozobil®), at 200 μg/d/mouse in sterile 200 μl of PBS were injected i.p. consecutively for 10-days; G-CSF was administered by daily i.p. injection at a dose of 250 μg/kg/d for 6 days. To assess B cell depletion, peripheral blood was collected at different time points and lymphocytes were isolated for staining and flow cytometry analysis.
Liu C.L., Lyle M.J., Shin S.C, & Miao C.H. (2016). Strategies to Target Long-Lived Plasma Cells for Treating Hemophilia A Inhibitors. Cellular immunology, 301, 65-73.
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5

Murine, Human Cell Line, and iPSC Differentiation Protocol

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The murine IL-3-dependent 32D hematopoietic cell line, human SH-SY5Y, and human induced pluripotent stem cells (hiPSCs, KYOUDXR0109B) were originally obtained from ATCC (Manassas, VA). 32D cells were maintained in RPMI 1640 medium supplemented with 10% heat-inactivated fetal bovine serum (FBS), 100 U/ml penicillin and 100 μg/ml streptomycin (Atlanta Biologicals, Norcross, GA), and 5 ng/mL recombinant murine IL-3 (PeproTech, Rocky Hill, NJ). For inducing 32D cell differentiation, cells were washed with HBSS to remove IL-3, and 500,000 cells were plated on Day 0 in RPMI 1640 containing 10% FBS, 1% penicillin-streptomycin, and 40 ng/mL recombinant murine G-CSF (PeproTech, Rocky Hill, NJ). At the indicated time points, cells were harvested, cytospun, and then Wright’s-stained (EMD Chemicals) for analysis of nuclear morphology to determine the extent of granulocytic differentiation54 (link). SHSY5Y human neuroblastoma cells were cultured in DMEM/F12 medium with 10% fetal bovine serum (Thermo Fisher Scientific/Gibco). Human iPSCs were cultured in Essential 8 Medium (Thermo Fisher Scientific/Gibco), as per the supplier's protocol.
MacNicol M.C., Cragle C.E., McDaniel F.K., Hardy L.L., Wang Y., Arumugam K., Rahmatallah Y., Glazko G.V., Wilczynska A., Childs G.V., Zhou D, & MacNicol A.M. (2017). Evasion of regulatory phosphorylation by an alternatively spliced isoform of Musashi2. Scientific Reports, 7, 11503.
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6

Murine Hematopoietic Stem Cell Expansion

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Lineage negative bone marrow (Lin−) cells were
isolated by magnetic cell separation as above. Cells were maintained at
3–5 ×105/mL throughout the culture process. Cells were
initially cultured in IMDM +20% FBS (AtlantaBiologics) in recombinant murine
IL-3 and SCF (Peprotech), both at 50 ng/mL. After 3 days, recombinant murine
G-CSF (Peprotech) at 50 ng/mL was added to the culture. On day 6, cells were
washed 4X in PBS and resuspended in IMDM +20% FBS and 50 ng/mL G-CSF. Viable
cell counts were determined by trypan blue exclusion.
Braun T.P., Estabrook J., Schonrock Z., Curtiss B.M., Darmusey L., Macaraeg J., Enright T., Coblentz C., Callahan R., Yashar W., Taherinasab A., Mohammed H., Coleman D.J., Druker B.J., Demir E., Lusardi T.A, & Maxson J.E. (2022). Asxl1 Deletion Disrupts MYC and RNA Polymerase II Function in Granulocyte Progenitors. Leukemia, 37(2), 478-487.
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7

Quantitative G-CSF Sandwich ELISA

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Rat antieG-CSF mAb (clone 67604) and biotinylated rabbit antieG-CSF polyclonal Ab (R&D Systems, Minneapolis, MN) were used in a sandwich enzyme-linked immunosorbent assay. Recombinant murine G-CSF (Peprotech, Rocky Hill, NJ) was used as a standard.
Goldberg G.L., Cornish A.L., Murphy J., Pang E.S., Lim L.L., Campbell I.K., Scalzo-Inguanti K., Chen X., McMenamin P.G., Maraskovsky E., McKenzie B.S, & Wicks I.P. (2016). G-CSF and Neutrophils Are Nonredundant Mediators of Murine Experimental Autoimmune Uveoretinitis. The American journal of pathology, 186(1).
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8

Hoxb8-Induced Neutrophil Differentiation and Interferon Response

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Eight week-old male B6.A2G-Mx1 and B6.A2G-Mx1-Tyk2−/− were used to generate Hoxb8 neutrophil cultures as described (49 (link)). Briefly, myeloid progenitor cells were derived from bone marrow, retrovirally transduced with an estrogen-regulated Hoxb8 construct (MSCV-ERHBD-Hoxb8 (50 (link)) and selected for 4 weeks in the presence of stem cell factor (SCF) to generate neutrophil progenitor lines. Polyclonal progenitor cell lines were cultured in OptiMEM + GlutaMAX medium (Life Technologies) supplemented with 10 % FCS, 30 μM β-mercaptoethanol (Life Technologies), 1 μM β-estradiol (Sigma-Aldrich) and 1 % supernatant from SCF-producing CHO cells. Differentiation was induced by β-estradiol removal in the presence of 1% SCF supernatant and 20 ng/ml murine recombinant G-CSF (Peprotech). After 3.5-4 days of differentiation cells were used for experiments. For IFN treatment, cells were treated for 4 h with the indicated concentrations of either IFN-αB/D or recombinant mouse IFN-λ2 and then processed for RNA isolation and subsequent RT-qPCR analysis.
Schnepf D., Crotta S., Thamamongood T., Stanifer M., Polcik L., Ohnemus A., Vier J., Jakob C., Llorian M., Gad H.H., Hartmann R., Strobl B., Kirschnek S., Boulant S., Schwemmle M., Wack A, & Staeheli P. (2021). Selective Janus Kinase Inhibition Preserves Interferon-Λ-mediated Antiviral Responses. Science immunology, 6(59), eabd5318.
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9

Neutrophil Progenitor Differentiation Protocol

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Hoxb8 neutrophil progenitors were derived from bone marrow of C57BL/6 wt or vav-Bcl-2-tg mice24 (link) expressing Bcl-2 throughout the hematopoietic lineage. Polyclonal neutrophil progenitor cell lines were established by retroviral transduction of Hoxb8 and selection in the presence of SCF.20 (link) Progenitor cells were cultured in Opti-MEM medium (Life Technologies, Carlsbad, CA, USA) supplemented with 10% FCS, 30 mM β-mercaptoethanol, 1% supernatant from SCF-producing CHO cells (kindly provided by Hans Häcker) and 1 μM β-estradiol (Sigma-Aldrich, Munich, Germany). Differentiation was induced by estrogen removal and culture in medium containing 1% SCF supernatant. Murine GM-SCF was either used as 1% cell culture supernatant of GM-CSF-producing B16 cells corresponding to a final concentration of approximately 10 ng/ml according to Wang et al.20 (link) (kindly provided by Hans Häcker), or as commercially available recombinant murine GM-CSF at a concentration of 10 ng/ml (Peprotech, Hamburg, Germany). Both GM-CSF preparations led to comparable results. Human recombinant GM-CSF (Peprotech), murine recombinant G-CSF (Peprotech), LPS (E. coli O55:B5, Sigma-Aldrich cat no. L2637, Munich, Germany) or the inhibitors LY94002 (20 μM, Sigma-Aldrich) Jak inhibitor 1 (JI1, 1 μM), Stattic (0.5 μM) and U0126 (10 μM, all from Calbiochem, San Diego, CA, USA) were used as indicated.
Vier J., Groth M., Sochalska M, & Kirschnek S. (2016). The anti-apoptotic Bcl-2 family protein A1/Bfl-1 regulates neutrophil survival and homeostasis and is controlled via PI3K and JAK/STAT signaling. Cell Death & Disease, 7(2), e2103-.
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