2.6. Treatment of cells
After 24 h preincubation the medium was aspirated and replenished with an equal volume of the serum-free medium 199
supplemented with 10 μg/mL insulin, 5 μg/mL transferrin and 3×10−8 M selenium as ITS medium (Sigma), containing 1, 10 and 100 ng/mL leptin for another 48 h.
The control received vehicle only.
Final concentration of DMSO in the medium was below 0.1%.
The medium was renewed every 24 h thereafter.