Preparation of individual electrically and video-recorded eggs for integrated temporal and electron microscopic analyses

A method for correlative studies of early fertilization events that integrates techniques of intracellular electrophysiological recording, video-imaging, and electron microscopy is described. A key feature of the method is its ability to identify the fertilizing sperm and to record the moment of egg excitation. Since the site of gamete interaction is recognizable throughout all stages of preparation, difficulties associated with locating the site of fertilization and determining specimen orientation for microtomy and electron microscopic examination are eliminated. Virtually all samples yield useful information. An example of interacting gametes fixed 4 sec after initiation of the fertilization potential and serial sectioned is described. The method is applicable to systems other than fertilizing eggs when functional, temporal, and spatial relationships of individual cells need to be correlated with changes in ultrastructure.