2.4. Quantification of Glycerol Release

Glycerol concentration in the medium was measured using a glycerol assay kit (Millipore, MA, USA) in accordance with the manufacturer’s instructions. On day 8, the differentiated adipocytes were incubated with/without 10 μM FA and isoproterenol (ISO-10 μM) for 24 h. ISO was used as a positive control. Then, the culture medium was collected and used for the quantification glycerol release, i.e., 100 μL of medium were transferred to 96-well plates and 200 μL of free glycerol assay reagent was added, and the plates were incubated at room temperature for 15 min. Then, the absorbance was read at 540 nm in a SpectraMax3X reader [10].