Structural specializations in the plasma membrane of opossum spermatozoa obtained from different levels of the epididymis have been analyzed in thin sections and freeze-fracture replicas. The maturation process was accompanied by a redistribution of intramembranous particles in the flagellar midpiece region. Caput epididymal spermatozoa are immotile, and freeze-fracture replicas of the midpiece plasma membrane reveal a random arrangement of intramembranous particles. As spermatozoa transit the corpus epididymis, the intramembranous particles in the midpiece plasma membrane are redistributed from a random arrangement to an organized packing pattern. This redistribution apparently involves the formation of chains of intramembranous particles which gradually increase in length, orient parallel to the flagellar long axis, and ultimately form numerous parallel rows, each three to five particles wide. In cauda epididymal spermatozoa the intramembranous particles within the rows are packed in an organized manner, and few free intramembranous particles are noted between rows. Analysis of thin sections revealed that the reorganization of intramembranous particles is accompanied by the deposition of a mat of amorphous material at the cytoplasmic face of the membrane. No striking changes in intramembranous particle distribution during epididymal maturation were found in other flagellar segments or in the plasma membrane overlying the sperm head.