Rat spermatozoa from the epididymis and ductus deferens were observed by surface replica, rapid-freeze and deep-etch, and conventional freeze-fracture methods. By the surface replica method, parallel periodical ridges were observed in the acrosomal region of the spermatozoa from the distal part of the cauda epididymis (zone 6) and from the ductus deferens. The periodicity of the ridges forming a domain was about 35 nm. A quantitative analysis of the spermatozoa along the reproductive tract indicated that 39.4% and 73.5% of the population in zone 6 of the epididymis and in the ductus deferens, respectively, had the domain. None of the spermatozoa from zone 1 through zone 5 had the domain. The results of the rapid-freeze and deep-etch procedure showed that the ridges observed by the surface replica method consisted of linear arrangements of elliptical particles on the ES face of the plasma membrane. The particles were about 30 nm in length and 15 nm in width. On the corresponding PF face of the plasma membrane, linear arrangements of the intramembrane particles (IMPs) of about 8 nm in diameter were observed by both the deep-etch and freeze-fracture methods. The IMPs tended to run in paired parallel lines. A close relationship was observed between the lines of the elliptical particles on the ES and of the IMPs on the PF faces. The elliptical particle may be a protruded part of the IMP(s) or other protein(s) bound to the IMP(s).