HuMSCs were purchased from Beijing Beina Chuanglian Biotechnology Institute. Cells were cultured in 25-cm2 culture flasks containing HyClone™ Dulbecco's modified Eagle's medium (HyClone; GE Healthcare Life Sciences), supplemented with 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.), 100 U/ml penicillin and 100 µg/ml streptomycin (Gibco; Thermo Fisher Scientific, Inc.). Cells were grown at 37°C under 5% CO2 atmosphere. The culture medium was replaced every 3 days and the HuMSCs were digested with trypsin (Gibco; Thermo Fisher Scientific, Inc.) once they reached 70–80% confluency. The cells in the fourth passage were used for further differentiation. Before hepatic differentiation, the multipotency of the cultured HuMSCs was confirmed by differentiation experiments. The cells were treated with osteogenic medium containing L-glutamine, decamethasone, ascorbate and β-glycerophosphate (Sigma-Aldrich; Merck KGaA) and chondrogenic medium containing h-Insulin, L-glutamin, dexamethasone, indomethacin and 3-isobuty-I-methyl-xanthine (Sigma-Aldrich; Merck KGaA) according to previous studies (15 (link),16 (link)).
Hyclone dulbecco s modified eagle s medium
Manufactured by GE Healthcare
Sourced in United States
Hyclone Dulbecco's modified Eagle's medium is a cell culture medium formulation that provides the necessary nutrients and components to support the growth and maintenance of a variety of cell types in in vitro experiments.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Ascorbate, by Merck Group (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
Trypsin, by Thermo Fisher Scientific (1 mentions)
Dexamethasone (dex), by Merck Group (1 mentions)
Indomethacin, by Merck Group (1 mentions)
β glycerophosphate, by Merck Group (1 mentions)
Antibiotic antimycotic, by Thermo Fisher Scientific (1 mentions)
Hek293t cell, by American Type Culture Collection (1 mentions)
Cryostat, by Leica (1 mentions)
Black filter paper, by Merck Group (1 mentions)
Gibco rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)
Hyclone fetal bovine serum fbs, by GE Healthcare (1 mentions)
Lipopolysaccharides (lps), by Merck Group (1 mentions)
Hyclone fetal bovine serum, by GE Healthcare (1 mentions)
Hyclone rpmi 1640 medium, by GE Healthcare (1 mentions)
Mda mb 231, by GE Healthcare (1 mentions)
Mcf 7, by GE Healthcare (1 mentions)
Mcf10a, by GE Healthcare (1 mentions)
8 protocols using hyclone dulbecco s modified eagle s medium
1
Differentiation of Human Mesenchymal Stem Cells
2
Bacterial Strains and Cell Culture
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Bacterial strains and plasmids used in this study are listed in Table S1 . Escherichia coli strains were grown in Luria-Bertani (LB) medium at 37 °C with appropriate antibiotics. HEK293T cells were purchased from the American Type Culture Collection (ATCC) and cultured at 37 °C with 5% CO2 in Hyclone Dulbecco's modified Eagle's medium (GE Healthcare) supplemented with 1% antibiotic/antimycotic (Gibco-BRL) and 10% fetal bovine serum (Gibco-BRL).
3
Preparation and Fixation of Retinal Samples
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The eyes were dissected in refrigerated HyClone Dulbecco’s Modified Eagles Medium (GE Healthcare Life Sciences, Logan, UT). The eyecups were then immersion-fixed in 4% (w/v) paraformaldehyde (PFA) in 0.1 M phosphate buffered saline (PBS), pH 7.4 for 45 minutes to 1 hour and cryoprotected overnight in 30% sucrose. These eyecups were sectioned at 16–20 μm with a Leica cryostat (Leica Microsystems, Buffalo Grove, IL) and mounted onto slides, which were stored at -20°C. For whole-mounted retinas, the sclera was removed and the retina was flattened photoreceptor side down on black filter paper (EMD Millipore Corporation, Bedford, MA). The retina was subsequently immersion-fixed in 4% PFA in 0.1 M PBS for 1 hour. The whole-mounted retina was stored in 0.1 M PBS until processing for immunohistochemistry.
4
Cell culture conditions for CRC lines
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Four CRC cell lines (SW480, HCT116, HT29, and SW620) and an intestinal epithelial cell line (NCM460) were donated by Dr Jinjun Rao (College of Pharmacy, Southern Medical University, Guangzhou, China). The cancer cell lines were incubated in Gibco RPMI-1640 medium (Thermo Fisher Scientific, Waltham, MA, USA) while NCM460 cells were cultured in HyClone Dulbecco’s Modified Eagle’s Medium (GE Healthcare Life Sciences, Logan, UT, USA); both types of media were supplemented with 10% HyClone fetal bovine serum (FBS; GE Healthcare Life Sciences). Cells were cultured at 37°C in a humidified environment containing 5% CO2.
5
Bone Marrow-Derived Macrophage Treatment
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Bone marrow-derived macrophages (BMDM) were cultured for 6 days in HyClone™ Dulbecco's Modified Eagles Medium (GE Healthcare Life Sciences, Missassauga, Ontario, Canada) conditioned with medium from L926 cells, as we described (Chen et al., 2015) and treated with lipopolysaccharides (LPS; Sigma-Aldrich, Oakville, Ontario, Canada, 100 ng/mL) with or without Dabrafenib (10 μM) for 4 hours. RNA was isolated for quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis.
6
Culturing Human Breast Cell Lines
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Cell lines and culture. Human breast cancer cell lines (MCF-7, BT474, MDA-MB-231, and MDA-MB-468), normal breast epithelial cell line MCF-10A and 293T cells were purchased from the American Type Culture Collection (Manassas, VA, USA). BT474 and MDA-MB-231 cells were cultured in Hyclone RPMI-1640 medium (GE Healthcare Life Sciences, Logan, UT, USA) while MCF-7, MDA-MB-468, MCF-10A and 293T cells were cultured in Hyclone Dulbecco's modified Eagle's medium (GE Healthcare Life Sciences). All cells were grown in medium containing 10% fetal bovine serum (Hyclone; GE Healthcare Life Sciences) supplemented with 1% penicillin-streptomycin (Gibco; Thermo Fisher Scientific Inc., Waltham, MA, USA) in a humidified atmosphere of 5% CO 2 at 37˚C.
7
Cytotoxicity Assay for OP-B Treatment
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To detect cell viability following exposure to various concentrations (0, 5, 10 and 20 mmol/l) of OP-B (Jrdun Biotechnology Corp., New York, NY, USA), 100 μl SGC-7901 cells (5×104/ml) were seeded in 96-well plates and were cultured for 0, 12, 24, 48 and 72 h at 37°C. Subsequently, 100 μl serum-free Dulbecco's modified Eagle's medium (Hyclone; GE Healthcare Life Sciences) containing 10% Cell Counting kit (CCK)-8 reagents (v/v) (Dojin Laboratories, Kumamoto, Japan) was added to each well, and the cells were cultured for 1 h in a 5% CO2 atmosphere at 37°C. Optical density was then measured at 450 nm using a Model 550 microplate reader (Bio-Rad Laboratories, Inc., Hercules, CA, USA).
8
Gastric Cancer Cell Line Characterization
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Gastric cancer cell lines. The present study used four different gastric cancer cell lines with different degrees of differentiation, including the MGC-803 well-differentiated gastric adenocarcinoma cell line, the SGC-7901 moderately differentiated gastric adenocarcinoma cell line, the BGC-823 poorly differentiated gastric adenocarcinoma cell line, the HGC-27 undifferentiated gastric adenocarcinoma cell line and the GES-1 normal gastric epithelial cell line (Stem Cell Bank, Chinese Academy of Sciences, Shanghai, China). Cell culture was performed using Dulbecco's modified Eagle's medium (Hyclone; GE Healthcare Life Sciences, Logan, UT, USA) containing 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc., Waltham, MA, USA) and the cells were incubated at 37˚C and 5% CO 2 .
Antibodies. Anti-HOXC6 mouse monoclonal antibody (sc-376330; 1:500) and anti-β-actin mouse monoclonal antibody (sc-47778; 1:1,000) were purchased from Santa Cruz Biotechnology, Co., Ltd. (Dallas, TX, USA). Mouse monoclonal anti-Flag antibody (F1804; 1:1,000) was purchased from Sigma-Aldrich (St. Louis, MO, USA). Rabbit polyclonal phosphorylated-extracellular signal-regulated kinase (ERK; #9101; 1:1,000) (Thr202/Tyr204) and rabbit monoclonal MMP9 (#13667; 1:1,000) antibodies were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA).
Antibodies. Anti-HOXC6 mouse monoclonal antibody (sc-376330; 1:500) and anti-β-actin mouse monoclonal antibody (sc-47778; 1:1,000) were purchased from Santa Cruz Biotechnology, Co., Ltd. (Dallas, TX, USA). Mouse monoclonal anti-Flag antibody (F1804; 1:1,000) was purchased from Sigma-Aldrich (St. Louis, MO, USA). Rabbit polyclonal phosphorylated-extracellular signal-regulated kinase (ERK; #9101; 1:1,000) (Thr202/Tyr204) and rabbit monoclonal MMP9 (#13667; 1:1,000) antibodies were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA).
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