Unlabelled fatty acids were purchased from Cayman Chemical
Company in solution in ethanol. The cells were cultured in the standard
culture medium (see above) but with the FBS concentration set at 2, 5 or
10% (v/v). The cells were seeded at a density of 25,000 cells cm−2 into
the required number of 75 cm2 cell culture flasks in a final volume of
15 mL of the culture medium. Once the cells had adhered to the flasks
(after approximately 4 h), 125 μM vitamin E and 10 μM of each
unlabelled fatty acid, complexed with fatty acid free bovine serum albumin,
were added to the appropriate flasks as described previously
(Scholefield and Schuller, 2014). The cells were then incubated at
25 °C for either 24 or 72 h.