The localization of proacrosin was determined by using colloidal gold labeling and electron microscopy of boar germ cells during spermiogenesis to post-ejaculation. Proacrosin was first localized in round spermatids during the Golgi phase of spermiogenesis; it was associated with the electron-dense granule, or acrosomal granule that was conspicuous within the acrosome. It remained within the acrosomal granule during the cap and acrosome phases of spermiogenesis. At these stages, there was no apparent association of the proacrosin molecule with the acrosomal membranes. During the maturation phase of spermiogenesis, proacrosin was seen to become dispersed into all regions of the acrosome except the equatorial segment. When sperm from different segments of the epididymis and ejaculated sperm were examined, localization was observed throughout the acrosome except for the equatorial segment. Here proacrosin appeared to be localized on both the inner and outer acrosomal membranes as well as with the acrosomal matrix, although further studies are required to verify the membrane localization. No labeling was seen on the plasma membrane. These data suggest that the synthesis and movement of proacrosin to sites in the acrosome are controlled by an as yet unknown process. The absence of proacrosin on the plasma membrane of mature ejaculated sperm makes it unlikely that this enzyme plays a role in sperm-zona adhesion prior to capacitation.