Primary cells isolated from pig bone marrow were differentiated into M1 macrophages using porcine granulocyte-macrophage colony-stimulating factor (GM-CSF) as previously described (Gao et al., 2018 (link)). In short, the bone marrow cells were thawed, seeded at 50,000 cells per well, and cultured in 0.1% GM-CSF (Biorad) in RPMI +/+ (Thermo Fisher Scientific). At day 3 of the culture, cells were supplemented with 50 μL of 0.1% GM-CSF. At day 6, the differentiated macrophages were stimulated for 24 h with isolated OMVs (0.5 µg/mL protein) or purified LPS (10 ng/mL, kindly provided by J. Pérez Ortega, Utrecht University). Thereafter, cell markers and cytokines were measured using fluorescence-activated cell sorting (FACS, Becton, Dickinson and Company) and enzyme-linked immunosorbent assay (ELISA), respectively. For FACS, antibodies against the following surface markers and their dilutions were: Swc3α-PE (1:4000, Invitrogen, Carlsbad, California, USA), recombinant CTLA-4-MuIg-APC (1:1000, Ancell, Stillwater, Minnesota, USA) which binds to porcine CD80/86, CD163-FITC (1:1000), and human CD14-PB (1:100, both from Biorad). For cytokine detection, corresponding ELISA kits were used (R&D Systems, Minneapolis, Minnesota, USA).
Gm csf
Manufactured by Bio-Rad
Sourced in United States
GM-CSF is a recombinant cytokine that stimulates the growth and differentiation of granulocyte and macrophage progenitor cells. It is a hematopoietic growth factor that can be used in cell culture and research applications.
Automatically generated - may contain errors
Lab products found in correlation
Ifn γ, by Bio-Rad (10 mentions)
Tnf α, by Bio-Rad (9 mentions)
G csf, by Bio-Rad (6 mentions)
Eotaxin, by Bio-Rad (5 mentions)
Mcp 1, by Bio-Rad (5 mentions)
Interleukin 6 (il 6), by Bio-Rad (4 mentions)
Mip 1α, by Bio-Rad (4 mentions)
Rantes, by Bio-Rad (4 mentions)
Interleukin 2 (il 2), by Bio-Rad (4 mentions)
Il 10, by Bio-Rad (3 mentions)
Ip 10, by Bio-Rad (3 mentions)
Interleukin 4 (il 4), by Bio-Rad (3 mentions)
Vascular endothelial growth factor (vegf), by Bio-Rad (3 mentions)
Il 1b, by Bio-Rad (2 mentions)
Il 17a, by Bio-Rad (2 mentions)
Il 23, by Thermo Fisher Scientific (2 mentions)
Ifn λ1, by Thermo Fisher Scientific (2 mentions)
Ifn λ2, by Thermo Fisher Scientific (2 mentions)
Ifnλ3, by Thermo Fisher Scientific (2 mentions)
Ifnλ4, by Thermo Fisher Scientific (2 mentions)
13 protocols using gm csf
1
Porcine Macrophage Differentiation and Stimulation
2
Quantifying Cytokines in Lung Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To quantify cytokine concentrations in cell free supernatants of lung tissue in vitro, ELISA (IL-18; MBL, Nagoya, Japan) and multiplex-assay (IL1b, IL6, IL8, IL10, IL12, IL17A, G-CSF, GM-CSF, TNF-α; Bio-Rad, München, Germany) were used after incubation of 0.3–0.4 g pieces of lung tissue at 37 °C and 5% CO2 in 2 mL RPMI medium (RPMI 1640 + 10% FCS) for 24 h. Cytokine assays were done according to kit instructions and were performed on 11 of the 13 lung specimens due to tissue shortage after pathological assessment in the remaining cases.
3
Serum Cytokine Detection Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For serum cytokine detection, blood samples were collected together with CRP and PCT samples at time points 0, 12, 24, 48, 72, and 96 h after study inclusion. The serum samples were kept at −75 °C until they were analyzed at the Virus Diagnostics Laboratory, University of Turku. Serum cytokine levels were determined by 27-plex immunoassay (IL-1β, IL-1rα, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12 (p70), IL-13, IL-15, IL-17, eotaxin, FGF basic, G-CSF, GM-CSF, IFN-γ, IP-10, MCP-1, MIP-1α, platelet-derived growth factor (PDGF-ββ), MIP-1, RANTES, TNF-α, vascular endothelial growth factor (VEGF)) from Bio-Rad Laboratories, Inc. (California, United States)). According to the manufacturer’s instructions, except that the amount of beads, detection antibodies, and streptavidin-phycoerythrin conjugate were used at 50% of their recommended concentration, which was tested previously as appropriate for the analysis system. The results were analyzed with Bio-Plex Manager 6.0 software. For statistical analyses and calculating the geometric mean cytokine levels, samples under the detection limit were given a value that was the detection limit divided by two. This was done in order to enable the inclusion of negative values (0 values) for geometric mean calculations.
4
Interictal Plasma Cytokine Profiling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Interictal samples were collected at day 7 from the last seizure attack. The plasma was harvested within 30 min at 37 °C of venipuncture from EDTA-anticoagulated blood samples and stored at −80 °C for subsequent cytokine analysis. The concentrations of IL-2, IL-4, IL-6, IL-8, IL-10, IFNγ, GM-CSF, TNFα (Bio-Rad, USA), IL-17a (PeproTech, Rocky Hill, NJ, USA), IL-1β (Bender MedSystems, Vienna, Austria), IL1Ra (Cytoscreen, Biosource, Belgium), IFNλ1, IFNλ2, IFNλ3, IFNλ4 (eBioscience, CA, USA), and IL-23 (Invitrogen, Carlsbad, CA, USA) were measured by ELISA according to manufacturers’ instruction [16 (link)–20 (link)]. CSF concentrations of IL-6, IFNγ, IFNλ3 and IL-17a were measured using human cytoline/chemokine MILLIplex kits (Millipore Corp, Billerica, MA, USA) [21 (link)]. ELISAs were performed in duplicate.
5
Cytokine Profile Analysis in Plasma
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Blood samples were collected by venipuncture into EDTA-anticoagulation vials. Plasma was harvested from the collected samples within 30 min and stored at −80 °C for subsequent cytokine analysis. Concentrations of IL-2, IL-4, IL-6, IL-8, IL-10, IFNγ, GM-CSF, TNFα (all antibodies from Bio-Rad, USA), IL-17a (PeproTech, Rocky Hill, NJ, USA), IL-1β (Bender MedSystems, Vienna, Austria), IL1Ra (Cytoscreen, Biosource, Belgium), IFNλ1, IFNλ2, IFNλ3, IFNλ4 (eBioscience, CA, USA), and IL-23 (Invitrogen, Carlsbad, CA, USA) were measured by ELISA according to manufacturers’ instruction16 (link)22 (link)23 (link). ELISAs were performed in duplicate.
6
Cytokine Profiling of LPS-Stimulated Monocytes
7
Multiplex Cytokine Profiling in Serum
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Serum cytokine levels were analyzed using single-plex sets for IL-1b, IL-2, IL-5, IL-4, IL-6, IL-8, IL-10, IL-12p40, GM-CSF, IFN-γ, CXCL10, and TNF-α (Bio-Rad, Hercules, CA, USA) following the manufacturer's instructions. Serum aliquots (50 μL) were used for analysis, with a minimum of 50 beads acquired per analyte. Median fluorescence intensities were measured using a Luminex 200 analyzer. Data collected was analyzed with MasterPlex CT control software and MasterPlex QT analysis software (Hitachi Software, San Bruno, CA, USA). Standard curves for each analyte were generated using standards provided by the manufacturer.
8
Cytokine and Growth Factor Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Interleukin 6 (IL-6), IL-8 (Sanquin, Amsterdam, Netherlands), tumor necrosis factor α (TNF-α, R&D, the Netherlands) concentrations in the culture supernatant were measured by commercial ELISA kits according to the instruction of the manufacturer. GM-CSF, MCP1, and VEGF (all Bio-rad, Veenendaal, the Netherlands) were measured by Magpix (Luminex Corporation). Lactate was measured by a Lactate Fluorometric Assay Kit (Biovision, CA, USA).
9
Comprehensive Cytokine Profiling in BAL Fluid
10
Anticancer Peptide Effects on Gastric Cancer
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The anticancer active peptide used in the present study, anticancer bioactive peptide (ACBP) is a low-molecular-weight active substance that was obtained from the Clinical Medical Research Center, Affiliated Hospital of Inner Mongolia Medical University (Hohhot, China). The poorly differentiated human gastric adenocarcinoma cell line of MKN-45, the highly differentiated human gastric adenocarcinoma cell line NCI-N87 and the immortalized and non-tumorigenic human gastric mucosal epithelial cell line GES-1 were purchased from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China). RPMI-1640 medium and high-glucose DMEM were purchased from Hyclone (Cytiva). Penicillin, streptomycin, fetal bovine serum and 0.25% trypsin were from Gibco (Thermo Fisher Scientific, Inc.).
The Bio-Plex ProT Human Cytokine 27-plex Assay (cat. no. M500KCAFOY; Bio-Rad Laboratories, Inc.) is a multiplex assay that detects cytokines including IL-1β, IL-1RA, IL-2, IL-4, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-13, IL-15, IL-17, Eotaxin, bFGF, GM-CSF, IFN-γ, MCP-1 (MCAF), IP-10, MIP-1α, PDGF-BB, MIP-1β, RANTES, TNF-α and VEGF. OXA was obtained from Jiangsu Hengrui Pharmaceutical Co., Ltd.
The Bio-Plex ProT Human Cytokine 27-plex Assay (cat. no. M500KCAFOY; Bio-Rad Laboratories, Inc.) is a multiplex assay that detects cytokines including IL-1β, IL-1RA, IL-2, IL-4, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-13, IL-15, IL-17, Eotaxin, bFGF, GM-CSF, IFN-γ, MCP-1 (MCAF), IP-10, MIP-1α, PDGF-BB, MIP-1β, RANTES, TNF-α and VEGF. OXA was obtained from Jiangsu Hengrui Pharmaceutical Co., Ltd.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!