Membrane Fluidity

Membrane fluidity was assessed using pyrenedecanoic acid (PDA; Membrane Fluidity Kit; Marker Gene Technologies, Eugene, OR). Cells (∼3 × 10⁶) were used for each independent experiment in triplicate. The labeling solution contained 20 µM of fluorescent lipid reagent and a final concentration of 0.08% Pluronic F127 in media. The cells were incubated with the labeling solution at room temperature for 20 min in dark. The cells were then washed three times with media and plated in a 96-well plate at a final volume of 200 mL. Fluorescence intensities were measured at 420 nm (monomer) and 460 nm (excimer) emissions for excitation at 360 nm using the Synergy2 fluorescence plate reader.