FRAP analysis of secretory granule lipids and proteins in the sea urchin egg

Cortical granules of the sea urchin are secreted at fertilization in response to sperm fusion. Approximately 15,000 of these vesicles are tightly tethered to the cytoplasmic face of the egg plasma membrane prior to insemination such that the vesicle-plasma membrane complex may be isolated and manipulated in vitro. Furthermore, this complex remains fusion competent and can thus be used for in vitro biochemical studies of secretion on a per-vesicle or a population scale. We document approaches to study the dynamics of membrane lipids and proteins in these secretory vesicles. Their large size (1.3-microm diameter), vast number, and ease of manipulation enable several unique approaches to study general secretion mechanisms.