The human nasopharyngeal carcinoma (NPC) cell line 5–8F and Sune1, human colon cancer cell line SW480 and lung cancer cell line H460 were obtained from the Cancer Research Institute of Southern Medical University (Guangzhou, China). In addition, the human NPC cell line C666-1 (EBV+) was obtained from Professor George Sai Wah Tsao (The University of Hong Kong, Hong Kong, China). The cell lines used in the current study were cultured in RPMI-1640 medium (Biological Industries, Kibbutz Beit-Haemek, Israel) supplemented with 10% fetal bovine serum (Biological Industries, Kibbutz Beit-Haemek, Israel) and incubated at 37°C with 5% CO2 for 48 h. The cells were collected, washed to remove serum, and then suspended in serum-free Dulbecco's modified Eagle's medium (DMEM)/F12 (Biological Industries) supplemented with 20 ng/ml epidermal growth factor, 10 ng/ml basic fibroblast growth factor and 2% B27 (1:50 dilution). These cell factors were purchased from PeproTech, Inc. (Rocky Hill, NJ, USA). The cells were cultured in ultra-low attachment 6-well or 96-well plates (Corning, Inc., Corning, NY, USA) at a density of up to 2,000 cells/ml and maintained at 37°C under 5% CO2 in a humidified incubator for 7 days.
Basic fibroblast growth factor (bfgf)
Manufactured by Sartorius
Sourced in Israel
Basic fibroblast growth factor is a protein that stimulates the proliferation and differentiation of various cell types, including fibroblasts, endothelial cells, and smooth muscle cells. It plays a crucial role in tissue repair and regeneration processes.
Automatically generated - may contain errors
Lab products found in correlation
Epidermal growth factor (egf), by Sartorius (2 mentions)
Rpmi 1640 medium, by Sartorius (1 mentions)
Fetal bovine serum (fbs), by Sartorius (1 mentions)
Ultra low attachment 6 well or 96 well plates, by Corning (1 mentions)
Pen strep, by Thermo Fisher Scientific (1 mentions)
Pen strep, by Merck Group (1 mentions)
L dmem, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Dexamethasone (dex), by Merck Group (1 mentions)
Insulin, by Thermo Fisher Scientific (1 mentions)
Indomethacin, by Merck Group (1 mentions)
Dexamethasone (dex), by Fujifilm (1 mentions)
Isobutylmethylxanthine ibmx, by Merck Group (1 mentions)
β glycerophosphate, by Merck Group (1 mentions)
Oil red o, by Merck Group (1 mentions)
Pen strep, by STEMCELL (1 mentions)
Ascorbate 2 phosphate, by Fujifilm (1 mentions)
Penicillin, by Sartorius (1 mentions)
Streptomycin, by Sartorius (1 mentions)
L glutamine, by Sartorius (1 mentions)
4 protocols using basic fibroblast growth factor (bfgf)
1
Culturing Cancer Cell Lines
2
Multilineage Differentiation of MSCs
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Adipogenic and osteogenic differentiation were performed according to the protocol mentioned previously [14 (link)]. To induce adipogenic differentiation, cells (3,000 cells/cm2) were cultured in L-DMEM (Gibco) supplemented with 10% fetal bovine serum (FBS) (Gibco), 0.5 mM isobutyl-methylxanthine (IBMX) (Sigma, St. Louis, MO, USA), 1 μM dexamethasone (Sigma), 10-μg/mL insulin (Gibco), 200 μM indomethacin (Sigma), and 1% Pen/Strep (Gibco) for 3 weeks. The presence of intracellular lipid droplets was confirmed by staining with 0.5 % Oil red O (Sigma). For osteogenic differentiation, cells (3,000 cells/cm2) were cultured in L-DMEM supplemented with 0.1 μM dexamethasone, 0.05 μM ascorbate-2-phosphate (Wako Chemicals, Richmond, VA, USA), 10 mM β-glycerophosphate (Sigma), 1% Pen/Strep and 10% FBS. After 4 weeks, osteogenic differentiation was assessed via staining with 2% alizarin red (pH 4.1–4.3; Fluka, Buchs, Switzerland).
For neurogenic differentiation, cells on collagen (type-I) coated coverslips were cultivated until 70% confluency. Cells were further cultured in differentiation medium (L-DMEM supplemented with 0.5 mM IBMX), epidermal growth factor (Biological Industries, Kibbutz Beit Haemek, Israel), basic fibroblast growth factor (Biological Industries), neural stem cell proliferation supplements (StemCell Technology, British Columbia, Canada), and 1% Pen/Strep.
For neurogenic differentiation, cells on collagen (type-I) coated coverslips were cultivated until 70% confluency. Cells were further cultured in differentiation medium (L-DMEM supplemented with 0.5 mM IBMX), epidermal growth factor (Biological Industries, Kibbutz Beit Haemek, Israel), basic fibroblast growth factor (Biological Industries), neural stem cell proliferation supplements (StemCell Technology, British Columbia, Canada), and 1% Pen/Strep.
3
Isolation and Purification of Microvascular Endothelial Cells
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MVEC were isolated from human adult foreskins using a previously described technique [17 (link),23 ]. In brief, foreskins obtained from adult circumcisions were placed in PBS supplemented with penicillin 200 units/mL, streptomycin 200 μg/mL and amphotericin B 0.5 μg/mL (all from Biological Industries, Kibbutz Beit Haemek, Israel). Foreskins were cut into 3 mm squares and placed in PBS containing 0.3% trypsin and 1% EDTA at 37°C for 30 minutes. Segments were then washed several times with PBS, placed in a Petri dish in M199 containing 10% foetal bovine serum (FBS), and individually compressed with the side of a scalpel blade to express microvascular fragments. The microvascular segments were passed through a 150 μm stainless steel mesh and collected by centrifugation at 300xg for 15 minutes. MVEC were seeded on a gelatin-coated cell culture flask and cultured in medium MCDB131 with 20% FBS; L-glutamine 2 mmol/L, penicillin 200 units/mL, streptomycin 200 μg/ml, EGF 20 ng/mL and bFGF 5 ng/mL (all from Biological Industries). When cells reached confluence, they were purified with Dynabeads CD31 (Dynabeads, Invitrogen Dynal ASA, Oslo, Norway) following the manufacturer’s instructions. Flow cytometry and positive staining for CD31, platelet endothelial cell adhesion molecule-1 (PECAM-1) confirmed the purity of the cell population. MVEC were used in passage 3
4
Culturing Human Mesenchymal Stem Cells and iPSCs
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Human bone marrow mesenchymal stem cells (hMSC) passages 2–6 (Lonza, Basel, Switzerland) were cultured using αMEM, supplemented with 10% FCS, 1% Pen-Strep®, 0.4% Fungizone®, and basic fibroblast growth factor (bFGF, 5 ng ml−1) (Biological Industries, Beit Haemek, Israel). The media was replaced at least every 2 days. hiPSC (kindly provided by Prof. Lior Gepstein, Technion) were cultured on matrigel (Corning, Glendale, AZ, USA) in mTeSR1 Basal Medium and supplemented with mTeSR1 5X supplement (STEMCELLTM Technologies, Vancouver, Canada). The medium was replaced every day. All Cells were cultured at 37 °C in a humidified incubator with 5% CO2.
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