Measurement of the translational mobility of concanavalin A in glycerol-saline solutions and on the cell surface by fluorescence recovery after photobleaching

Abstract

The fluorescence recovery kinetics of succinyl-fluorescein Concanavalin A (S-F-ConA) in glycerol-physiological saline solutions of high viscosity and when bound to the surface of mouse fibroblasts were measured following brief photobleaching using a laser excited fluorescence microscope. In the high viscosity solutions, the recovery kinetics, interpreted on the basis of a simple diffusion model, yielded a diffusion coefficient in close agreement with the values predicted by the Stokes-Einstein equation. Recovery kinetics for S-F-ConA bound to the surface of mouse 3T3 and SV3T3 cells cultured in vitro yielded diffusion coefficients in the range of 5-10-10(-11) cm2/s, values considerably lower than those reported previously for membrane proteins. These measurements indicated that a considerable fraction of the S-F-ConA molecules bound to the cell surface are immobilized. These results are discussed in relation to current concepts of lateral motion of protein components within natural membranes.