The fraction of CD31+ cells (106 cells/1 mL of medium) obtained after magnetic sorting was cultured on gelatin-coated plastic tablets for T-25 cell cultures in the medium consisted of 50% DMEM (Sigma-Aldrich, St. Louis, MO), 10% fetal bovine serum (FBS, Sigma-Aldrich, USA), 2 mM L-glutamine (Sigma-Aldrich, St. Louis, MO), 100 U/mL penicillin and 100 μg/mL streptomycin (Sigma-Aldrich, St. Louis, MO). The next day, we have replaced medium on medium with M199 (Sigma-Aldrich, St. Louis, MO) with 15% FBS (Sigma-Aldrich, St. Louis, MO), 1 mM L-glutamine (Sigma-Aldrich, St. Louis, MO), 100 μg/mL heparin (Sigma-Aldrich, St. Louis, MO) and endothelial growth factor 50 μg/mL (Sigma-Aldrich, St. Louis, MO). Cells were cultured for 5 days in standard gas (3.5% CO2) and temperature conditions (37°C). The medium was changed every 1–2 days. Evaluation of the effectiveness of the cultivation carried out to increase the mass of CD31+ cells was performed using flow cytometry.
Endothelial growth factor
Manufactured by Merck Group
Sourced in United States
Endothelial growth factor is a protein that stimulates the growth and proliferation of endothelial cells, which are the cells that line the interior of blood vessels. This protein plays a crucial role in the formation of new blood vessels, a process known as angiogenesis. It is commonly used in cell culture and research applications to study and promote the growth of endothelial cells.
Automatically generated - may contain errors
Lab products found in correlation
Heparin, by Merck Group (4 mentions)
L glutamine, by Merck Group (2 mentions)
Basic fibroblast growth factor (bfgf), by Merck Group (2 mentions)
Fetal bovine serum (fbs), by Merck Group (2 mentions)
Penicillin, by Thermo Fisher Scientific (2 mentions)
Streptomycin, by Thermo Fisher Scientific (2 mentions)
Dulbecco modified eagle medium (dmem), by Merck Group (1 mentions)
Penicillin, by Merck Group (1 mentions)
Streptomycin, by Merck Group (1 mentions)
Dispase solution, by Thermo Fisher Scientific (1 mentions)
Gentamycin, by Merck Group (1 mentions)
48 well plate, by Corning (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Thermo Fisher Scientific (1 mentions)
Medium 199, by Thermo Fisher Scientific (1 mentions)
Insulin, by Merck Group (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Hydrocortisone, by Merck Group (1 mentions)
Endothelium cell medium, by ScienCell (1 mentions)
Huvecs, by ScienCell (1 mentions)
Foetal bovine serum (fbs), by Lonza (1 mentions)
7 protocols using endothelial growth factor
1
Expansion of Endothelial Progenitor Cells
2
Mammosphere Formation Assay
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Single cells were plated at a density of 1,000 cells/mL in 96-well ultralow attachment plates (Corning, New York, USA). Cells were grown in serum-free DMEM/F12 supplemented with B27 (1:50, Invitrogen), 20 ng/mL endothelial growth factor, 20 ng/mL basic fibroblast growth factor and 4 μg/mL heparin (Sigma, St. Louis, MO, USA). Cells were cultured for 7 d, and mammospheres with >50 μm diameter were counted.
3
Culturing HUVECs with hTERT for longevity
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The HUEhT-1 (HUVECs modified with pIRES-hTERT-hygr) cell line was purchased from the RIKEN Cell Bank (Ibaraki, Japan). Cells were cultured in a humidified incubator with 5% CO2 at 37 °C using MCDB107 (Peptide Institute, Osaka, Japan) base medium supplemented with 10 ng/mL endothelial growth factor, 10 ng/mL basic fibroblast growth factor (Sigma-Aldrich), and 10 v/v% fetal bovine serum (FBS).
4
Isolation of Human Umbilical Vein Endothelial Cells
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Individual donor endothelial cells (HUVECs) were isolates from human umbilical cords according to the methods of Jaffe [55 (link)] and Scheglovitova [56 (link)]. Umbilical cords were obtained after normal parturition from five healthy donors following informed written consent.
Briefly, fresh umbilical veins were cannulated and filled with dispase solution (2 mg/mL) (Gibco, New York, NY, USA) and incubated at 37 °C for 30 min. Then, the veins were perfused with PBS. Cells were collected from the perfusate by centrifugation at 1000 rpm for 10 min, resuspended in Medium 199 (Gibco) supplemented with 10% fetal calf serum (HighClone, Logan, UT, USA), 200 μg/mL endothelial growth factor (Sigma), 100 μg/mL heparin (Moskovskii endokrinnyi zavod, Moscow, Russia), and 50 μg/mL gentamycin (KRKA), and seeded into 25–75 sm2 (6-well plates) cultural flasks. Cells were cultured in a humidified atmosphere of 5% CO2 at 37 °C. The confluent primary monolayers were washed and trypsinized (0.05% trypsin + 0.02% EDTA, Gibco). The cells were resuspended into a complete medium, seeded in 48-well plates (140,000 cells/mL) (Costar, Washington, DC, USA), and cultured for four days. Only the first subcultures were used for further experiments.
Briefly, fresh umbilical veins were cannulated and filled with dispase solution (2 mg/mL) (Gibco, New York, NY, USA) and incubated at 37 °C for 30 min. Then, the veins were perfused with PBS. Cells were collected from the perfusate by centrifugation at 1000 rpm for 10 min, resuspended in Medium 199 (Gibco) supplemented with 10% fetal calf serum (HighClone, Logan, UT, USA), 200 μg/mL endothelial growth factor (Sigma), 100 μg/mL heparin (Moskovskii endokrinnyi zavod, Moscow, Russia), and 50 μg/mL gentamycin (KRKA), and seeded into 25–75 sm2 (6-well plates) cultural flasks. Cells were cultured in a humidified atmosphere of 5% CO2 at 37 °C. The confluent primary monolayers were washed and trypsinized (0.05% trypsin + 0.02% EDTA, Gibco). The cells were resuspended into a complete medium, seeded in 48-well plates (140,000 cells/mL) (Costar, Washington, DC, USA), and cultured for four days. Only the first subcultures were used for further experiments.
5
Doxorubicin Cytotoxicity in Cell Lines
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Doxorubicin was purchased from Zurich pharma, and dimethyl sulfoxide (DMSO) (Sigma–Aldrich cat 276855), L-glutamine, insulin, hydrocortisone, and endothelial growth factor were purchased from Sigma–Aldrich, Chemie GmbH, Taufkirchen, Germany.
6
HUVEC Isolation and Culture
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Human umbilical vein endothelial cells (HUVECs) were purchased from Sciencell Research Laboratories, Inc., (cat. no. 8000). The cells were cultured in the endothelium cell medium (Sciencell Research Laboratories, Inc.) containing 5% fetal bovine serum (Sigma-Aldrich; Merck KGaA), 1% endothelial growth factor (Sigma-Aldrich; Merck KGaA), 100 µg/ml streptomycin (Gibco; Thermo Fisher Scientific, Inc.) and 100 U/ml penicillin (Gibco; Thermo Fisher Scientific, Inc.) in an incubator with 95% O2 and 5% CO2 at 37°C.
7
Dural Membrane Cell Culture Protocol
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Samples (∼1 cm2), of dural membrane were dissected aseptically away from the arachnoid and the pia mater (avoid vascular endothelial contamination) and placed in a six-well plate (Thermo Fisher Scientific Ltd), and cultured in medium m199 (Sigma) supplemented with foetal bovine serum (20% v/v, Lonza), l -glutamine (2 mM, Lonza), sodium pyruvate (1.1 mg mL−1, Sigma), heparin (10 U mL−1), penicillin/streptomycin (50 U mL−1, Lonza) and endothelial growth factor (15 µg mL−1, Sigma) at 37°C in 5% (v/v) CO2 in air. After 7 days of outgrowth, the cells were harvested and transferred to 75 cm2 flasks (Fisher) and expanded in supplemented m199 medium.
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