Dispersion-relation fluorescence spectroscopy

Abstract

Because of its ability to study specifically labeled structures, fluorescence microscopy is the most widely used technique for investigating live cell dynamics and function. Fluorescence correlation spectroscopy is an established method for studying molecular transport and diffusion coefficients at a fixed spatial scale. We propose a new approach, dispersion-relation fluorescence spectroscopy (DFS), to study the transport dynamics over a broad range of spatial and temporal scales. The molecules of interest are labeled with a fluorophore whose motion gives rise to spontaneous fluorescence intensity fluctuations that are analyzed to quantify the governing mass transport dynamics. These data are characterized by the effective dispersion relation. We report on experiments demonstrating that DFS can distinguish diffusive from advection motion in a model system, where we obtain quantitatively accurate values of both diffusivities and advection velocities. Because of its spatially resolved information, DFS can distinguish between directed and diffusive transport in living cells. Our data indicate that the fluorescently labeled actin cytoskeleton exhibits active transport motion along a direction parallel to the fibers and diffusive in the perpendicular direction.