2.8. Oxygen Consumption (Energy Consumption) Assay

To determine the energy expenditure in Tg^(−/−) and Tg^(+/+) with salt-treated WAT and BLA, oxygen (O₂) consumption (RFU/h) in the respiratory system of mitochondria in differentiated and live adipocytes was measured by the Mito Xpress pH-Xtra data visualization tool system (Agilent, Santa Clara, CA, USA) [19].The working solution was prepared by diluting the medium with Xpress agent at a 1:9 ratio. After the differentiation, the cells were incubated with the working solution and two drops of HS mineral oil per well to block the O₂ in flow from the medium. Absorbance was measured at 380 and 650 nm using a fluorescence plate reader (Molecular Devices), and the O₂ consumption rate was recorded every 17 sec for a total of 100 min. We collected the data at two points during the experiments, such as at the inflection points of the curve (almost at 30 min from the beginning), and at the end points.