Laurdan fluorescence experiments

The experiments were performed in a QuantaMaster 40 spectrofluorometer (Photon Technology International, Lawrenceville, NJ) using Laurdan. Laurdan is a solvatochromic dye that exhibits an increase in charge separation when excited in polar solvents, which results in a larger dipole moment⁵⁸. Laurdan shows different maximum emission intensities with liquid-ordered (440 nm) and liquid-disordered phases (490 nm). The emission spectrum changes in response to variations in the membrane environment, particularly in the glycerol backbone region in the phospholipid membrane. In order to quantify the spectral changes the generalized polarization function (GP) is used, that is obtained from measurements of wavelength displacements.

GP measurements are performed using excitation light at 360 nm and recording emission intensities both at 440 and 490 nm. Multilamellar vesicles (MLVs) were prepared as described above with 1 mol% Laurdan, and measurements were carried out at room temperature and constant stirring. Theoretically, GP values can vary from −1.0 (disordered) to +1.0 (ordered phases) but experimental values usually occur in the −0.3 to +0.6 range⁵⁹.