Phosphatidylethanolamine-binding protein (PBP) has been described previously in the male reproductive tract, where it has been implicated in the biogenesis and maintenance of antigen segregation of membranes. In the present study we have used a specific antiserum to PBP to determine its expression and localisation in the adult and prepubertal rat testis and epididymis by Western blotting and immunohistochemistry. In the adult rat testis, PBP was localised to step 17-19 elongating spermatids, residual bodies, and interstitial Leydig cells. In the adult epididymis, PBP was localised to epithelial cells of the caput, corpus, and cauda regions and to the cytoplasmic droplets of spermatozoa in the lumen of the initial segment, caput, and corpus epididymidis. In prepubertal animals, PBP was expressed in both testes and epididymides from day 1 and day 3 postpartum, respectively (day 3 being the earliest epididymal tissue taken). In prepubertal testes, PBP was localised to Leydig cells from day 1 postpartum and was not detected in any other cell type until the differentiation of elongate spermatids, when it was detected in step 17-19 elongating spermatids. These data suggest that PBP may be involved in the organisation of sperm membranes during spermiogenesis. The presence of PBP in Leydig cells, however, suggests diverse roles for this protein as a lipid carrier or binding protein.