*4.5. DCF Assay for Determining ROS Generation*

We adapted the method of Wang and Joseph [67]. Briefly, A549 cells in 96-well plates were incubated for 30 min with 10 or 50 µM membrane-permeant and non-fluorescent 2<sup>0</sup> ,70 -dichlorofluorescein diacetate (DCFDA) (Sigma-Aldrich, Saint Louis, MO, USA), which was deacetylated by non-specific intracellular esterases into the membrane-impermeant and non-fluorescent DCFH2. Cells were then washed with Hank's balanced salt solution (HBSS) and incubated with H2O<sup>2</sup> + LPS with/without BUD, HPβCD, or BUD:HPβCD complex in HBSS. Oxidative stress was evaluated through the measure of the fluorescence of DCF resulting from the oxidation of DCFH<sup>2</sup> by intracellular oxidants (λex = 490 nm; λem = 523 nm).
