The study protocol was approved by the Institutional Review Board (IRB) of Duksung Women's University (IRB No. 2017-002-001, 2018-007-006). ECFCs were isolated from the adherent mononuclear cell fraction of human peripheral blood using CD31-coated magnetic beads (Invitrogen, Waltham, MA, USA) as described previously.14 (link) Isolated ECFCs were expanded on 1% gelatin-coated plates (BD Biosciences, Franklin Lakes, NJ, USA) using EC growth medium MV 2 (EGM-MV 2 without hydrocortisone; Promocell, Heidelberg, Germany) supplemented with 10% fetal bovine serum (FBS; Atlas Biologicals, Fort Collins, CO, USA) and 1% glutamine-penicillin-streptomycin (GPS; Gibco, Waltham, MA, USA). ECFCs obtained between passages 7 and 10 were used for all experiments.
Glutamine penicillin streptomycin gps
Manufactured by Thermo Fisher Scientific
Sourced in United States
Glutamine-penicillin-streptomycin (GPS) is a commonly used cell culture supplement. It contains L-glutamine, penicillin, and streptomycin. GPS is used to support the growth and health of cells in various cell culture applications.
Automatically generated - may contain errors
Lab products found in correlation
Cd31 coated magnetic beads, by Thermo Fisher Scientific (2 mentions)
Gelatin coated plates, by BD (2 mentions)
Egm mv 2 without hydrocortisone, by PromoCell (1 mentions)
L ornithine, by Merck Group (1 mentions)
I2643, by Merck Group (1 mentions)
H4001, by Merck Group (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (1 mentions)
Msc growth medium, by Lonza (1 mentions)
Fetal bovine serum (fbs), by Atlas Biologicals (1 mentions)
Endothelial growth medium 2 egm 2, by Lonza (1 mentions)
3 protocols using glutamine penicillin streptomycin gps
1
Isolation and Expansion of Endothelial Colony-Forming Cells
2
Isolation and Culture of Primary Human Hepatocytes
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A published collagenase perfusion technique was employed for PHH isolation from liver wedges [13 (link)]. Briefly liver was digested and centrifuged to isolate PHH. These cells were resuspended and then plated on 24-well plates previously coated with rat tail collagen type 1 at a density of 3x105 cells/well in Dulbecco’s Modified Eagle’s Medium (DMEM) (Catalogue number [CN]:41965–039; Gibco laboratories, Gaithersburg, MD, USA) supplemented with 10% fetal calf serum (FCS) (CN:10270106; Gibco) and 5% glutamine/penicillin/ streptomycin (GPS) (CN:10378016; Gibco). After 2 hours the cells were washed with Phosphate-buffered saline (PBS) (CN:10010023; Gibco) and the media changed to our standard medium for PHH culture, constituted of Arginine-/Glutamine-free Williams E (CN:12551032; Gibco) with 1% GPS, hydrocortisone (2μg/ml) (H4001; Sigma-Aldrich, St. Louis, MO., USA), insulin (0.124 U/ml) (I2643; Sigma-Aldrich) and L-ornithine (400μM) (O6503; Sigma-Aldrich), subsequently the cells were kept at 37°C in 95% air/ 5% CO2.
3
Isolation and Expansion of ECFCs and MSCs
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The study protocol was approved by the institutional review board of Duksung Women’s University (IRB No. 2017-002-001). Human peripheral blood was provided from the national biobank. ECFCs were isolated from the adherent mononuclear cell (MNC) fraction using CD31-coated magnetic beads (Invitrogen, MA, USA) as described in the previous report (Melero-Martin et al., 2008 (link)). The isolated ECFCs were expanded on 1% gelatin-coated plates (BD Biosciences, NJ, USA) using endothelial growth medium-2 (EGM-2; Lonza, MD, USA) without hydrocortisone supplemented with 10% fetal bovine serum (FBS; Atlas Biologicals, CO, USA) and 1% glutamine-penicillin-streptomycin (GPS; Gibco, MA, USA). In all experiments, ECFCs from passages 7 to 10 were used.
MSCs were obtained from the MNC fraction of human adult bone marrow (Lonza). MSCs were cultured in MSC growth medium (Lonza) containing 10% FBS and 1% GPS until 80% confluence was achieved. MSCs from passage numbers 5 to 8 were used.
MSCs were obtained from the MNC fraction of human adult bone marrow (Lonza). MSCs were cultured in MSC growth medium (Lonza) containing 10% FBS and 1% GPS until 80% confluence was achieved. MSCs from passage numbers 5 to 8 were used.
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