A recently developed procedure which permits the isolation of discrete populations of seminiferous epithelial cells from the prepuberal and adult mouse testis is reviewed in this presentation. The procedure involves an initial incubation of the testis with collagena. se in an enriched Krebs-Ringer medium (EKRB). This step dissociates the interstitial tissue but leaves the dispersed seminiferous tubules(or cords) relatively intact. The purified seminiferous epithelium is then dissociated with trypsin to produce a monodisperse cell suspension. Furthermore, suspensions enriched in specific cell types can be obtained by selecting animals of the appropriate developmental age. These respective cell suspensions are subjected to sedimentation velocity at unit gravity using a 2-4% bovine serum albumin gradient in EKRB. This technique enables the isolation of highly enriched populations of prepuberal Sertoli cells and primitive type A spermatogonia on day 6, type A spermatogonia and type B spermato-gonia on day 8, preleptotene, leptotene/zygotene and early-mid pachytene spermatocytes on day 18 and finally late pachytene spermatocytes, round spermatids and residual bodies from the adult seminiferous epithelium. The advantages of this sequential enzymatic procedure include:(a) the elimination of both blood and interstitial tissue,(b) a marked reduction in the incidence of symplasts(multinucleated cells) and (c) an apparent preservation of cellular integrity as assessed by the exclusion of trypan blue, ultrastructural morphology, oxygen consumption and a quantitative and qualitative recovery of ribonucleic acid and histone species. Limitations inherent to the dissociation technique and isolated cell populations are considered in detail, including some counteractive measures.

The mammalian testis contains two discrete morphological compartments, interstitial tissue and the seminiferous tubules. Interstitial tissue forms the minor compartment and is composed primarily of vascular, lymphatic and connective tissue elements, macrophages, fibroblasts and the androgen secreting interstitial cells of Leydig(15). The seminiferous epithelium forms the major compartment and contains the differentiating germ cells intercalated between the supporting Sertoli cells. Spermatogenesis is a continuum of germ cell differentiation which occurs in three principal phases: the mitotic renewal and proliferation of spermatogonia, meiosis and spermiogenesis(Fig. 1)(29, 43, 44). The proliferation of spermatogonia oc-