Total internal reflection fluorescence microscopy has been applied to image
the final stage of constitutive exocytosis, which is the fusion of single
post-Golgi carriers with the plasma membrane. The use of a membrane protein
tagged with green fluorescent protein allowed the kinetics of fusion to be
followed with a time resolution of 30 frames/s. Quantitative analysis allo
wed carriers undergoing fusion to be easily distinguished from carriers mov
ing perpendicularly to the plasma membrane. The flattening of the carriers
into the plasma membrane is seen as a simultaneous rise in the total, peak,
and width of the fluorescence intensity. The duration of this flattening p
rocess depends on the size of the carriers, distinguishing small spherical
from large tubular carriers. The spread of the membrane protein into the pl
asma membrane upon fusion is diffusive. Mapping many fusion sites of a sing
le cell reveals that there are no preferred sites for constitutive exocytos
is in this system.