We have determined fusion pore closure time by dynamically counting the amount of C4 tail lipid beads present in the space between the lipid bilayers (see figure ). For planar and parallel membranes there are no C4 beads between the bilayers simply because all lipid tails point toward their respective bilayer interiors. On the other hand, for the formed fusion pore (where lipids surrounding the water channel are gradually tilted toward the opposite bilayer describing a clean membrane fusion path), the amount of C4 beads increases as a function of the fusion pore width. For more details on lipid tilting and splaying during membrane fusion see Mirjanian et al. and Caparotta et al. .