Preparation of Fatty Acid-BSA Solutions

The palmitic acid stock solution was prepared using fatty acid-free BSA (A8806, Sigma) at a molar ratio of 3.3:1 (palmitic:BSA) [45 (link)]. To prepare the stock solution, 500 mM palmitic acid (P0500, Merck) was dissolved in absolute ethanol at 70 °C. Then, 10 μL of palmitic acid solution was added to 1 mL prewarmed 10% BSA (prepared in cell culture medium). The mixture was incubated at 55 °C until it became clear and was then filtered before use. For the vehicle control, 10 μL ethanol was mixed with 1 mL 10% BSA. Oleic acid was prepared at a molar ratio of 6:1 (Oleic acid:BSA) using the product (29557, Cayman, MI, USA). The vehicle stock solution was prepared by dissolving 0.8 mM BSA in 150 mM sodium chloride (S3014, Sigma).