FIGURE 4.

Fusion of a single Rh-DOPE-labeled Syb1–116 vesicle to an acceptor SNARE complex-containing bilayer. A, fluorescence micrographs of a docked vesicle at different time points during fusion showing the dissipation of the fluorescent lipid tracer into the planar membrane. The peak fluorescence intensity trace of this particular vesicle is shown in Fig. 3A. Here, time point zero is set to the onset of fusion. B, diffusion of Rh-DOPE from the fusion site. Total fluorescence intensities of the central pixel (black) and of rings with radii of 2 pixels (0.75 μm) (red) and 4 pixels (1.5 μm) (blue) from the vesicle center are plotted for the same fusion event as in A as a function of time. The solid lines are the corresponding theoretically expected intensities from a simulation calculated for lipids that diffuse into the planar bilayer from a 90-nm disk source with a hypothetical diffusion coefficient of 1 μm²/s and taking into account a 1/e half-width point spread function of 0.25 μm.