Intracellular membrane traffic in cells has been postulated to require small vesicles which shuttle between fixed endomembrane compartments of the cell. Such vesicles are thought to carry contents in a vectorial manner and return membrane material to the point of origin after delivery. Though often postulated, little morphological evidence has been forthcoming to show that these are real. Cryoelectron microscopy will be utilized to attempt to visualize such vesicles in Paramecium. Cryofixation will be used to physically fix cells ultrarapidly so intermediate steps in fission or fusion can be preserved. Subsequent cryosubstitution or deep-etch rotary-shadow steps will allow observation of the cell interior in a nearly native state, making it possible to locate small transport or shuttle vesicles. Monoclonal antibody histochemistry will be used to identify membrane components of the shuttle vesicles. These studies will provide new information on the movements of membranes and membrane components within cells.
