Giant vesicles, laurdan, and two-photon fluorescence microscopy: Evidence of lipid lateral separation in bilayers

Abstract

This chapter describes giant vesicles, Laurdan, and two-photon fluorescence microscopy. The combination of Laurdan, giant unilamellar vesicles (GUVs), and two-photon fluorescence microscopy has been extremely useful in producing a microscopic picture of lipid-phase coexistence in the GUV bilayer model system. Laurdan is a unique probe, giving simultaneous information about morphology and phase state of lipid domains from fluorescence images. The critical issues in membrane biophysics today are centered on the molecular dynamics of the bilayer structure. The interplay between lipids that results in the formation of domains on a bilayer surface and the interactions among these domains and relevant membrane-associated biomolecules is of particular interest. The advantages of using a microscope as the optical arrangement are clear. The light collection efficiency of a well-designed microscope is greatly enhanced over other optical arrangements. In addition, the flexibility of fluorescence microscopes creates for the spectroscopist a malleable optical compartment that can be designed and readily redesigned as needed.