The cellularity of the human prepubertal testicular interstitium has not been well studied at the ultrastructural level. In this study, testicular biopsies were obtained from 35 boys aged three to nine years and examined by electron microscopy to clarify and quantitate the cell types present during the prepubertal period. The prepubertal testicular interstitium is found to consist of immature Leydig cells (9%), primitive fibroblastic cells (63%) (intertubular in location), and attenuated peritubular fibroblasts (28%). The primitive fibroblastic cells and peritubular fibroblasts appear closely related, being distinguished mainly by shape and location. The immature Leydig cell type contrasts with the fibroblastic cell types by exhibiting an irregular nucleus with relatively little heterochromatin. The most impressive cytoplasmic feature is the moderate to extensive development of smooth endoplasmic reticulum in the form of anastamosing tubules. In contrast, the rough endoplasmic reticulum is not well developed. Other cytoplasmic characteristics are the highly developed Golgi elements and occasional lipid droplets and lysosomes. Glycogen is also often present and is generally found in those cells that do not contain a well-developed smooth endoplasmic reticulum. The ultrastructure of the immature Leydig cell is compared with that of the mature fetal and adult Leydig cells. Although generally found in small clusters between tubules, these cells are often attenuated and closely associated with the seminiferous tubules. Occasional intermediate cell morphologies suggest a relationship between the primitive fibroblasts and immature Leydig cells. The presence of small cells exhibiting a steroid-producing morphology, classified as immature Leydig cells, in the prepubertal testicular interstitium is an interesting finding and is in accordance with earlier studies on nonhuman mammals. It is unknown whether these cells are remnants of the fetal Leydig cell population or have differentiated neonatally from the primitive fibroblastic cells. It is suggested that the immature Leydig cells are the progenitors of the adult Leydig cell population.