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At 24 hours after plating, the medium was changed with new medium completed with 500 µM of copper sulfate (CuSO4), copper chloride (CuCl2), sodium sulfate (Na2SO4) or sodium chloride (NaCl).
Based on CFSE peak fluorescence intensity halving, the calculated number of cell divisions was only 4.5 for melanocytes cultured in M254 medium, while cells cultured in Mel-mix medium completed 8 dividing cycles in the 120-hour experimental period (Figure 4).
All experiments were performed three times on each of the three cell lines cultivated in usual conditions (three days in Leibovitz's L-15 Medium completed with FCS).
HUVECs were cultured in complete EBM (Endothelial Basal Medium, Lonza, Walkersville MD) medium completed with growth factors and antibiotics (EGM SingleQuots, Lonza) in cells culture dishes (Corning, USA) coated with Collagen S type I, 0.3 μg/mL (Roche, Mannheim, Germany).
Education level was grouped into three categories based on the highest education level attained: low (illiterate individuals and those who did not complete primary education), medium (completed primary education), and high (completed secondary or higher-level education).
Cells (3.10 cells) were pelleted by centrifugation and suspended in DMEM/F-12 medium completed with 10% fetal bovine serum and 10% DMSO and frozen at -80°C until the preparation of cell extracts.
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After 12 days of culturing in the feather meal medium, complete decomposition of feathers was observed only in flask B4.
In some experiments, medium (complete or defined) or PBS was treated separately from the cells, designated 'separated treatment'.
The particles were stable in water as well as in cell culture medium (complete DMEM).
After 4 hr incubation at 37°C to allow cell adhesion, the medium was replaced either by starvation medium (complete medium without insulin) or complete medium (for autophagy studies), and cells were further incubated at 37°C overnight.
Then medium was changed to RPMI medium complete medium containing 60 μg/ml hygromycin B and cells were cultured for 2 weeks for selection of stable integration events.
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