Spermatogonial stem cells are at the beginning of the spermatogenic process. In non-primate mammals they are single cells (As spermatogonia). Their daughter cells either migrate away from each other and become two new stem cells or stay together, connected by an intercellular bridge. The latter designates these cells to differentiate and ultimately become spermatozoa. The purification of spermatogonial stem cells is hampered by the lack of specific markers. Present protocols only allow for a purity of 10% at best. Spermatogonial stem cells are difficult to culture in the absence of serum and a feeder layer. Better results have been obtained with co-cultures of Sertoli cells with mouse and bovine spermatogonia.Spermatogonial stem cells can be transplanted to recipient testes the endogenous spermatogenesis of which has been removed which makes it possible to perform functional assays of stem cell capacity of germ cell suspensions. The ratio between self renewal and differentiation of spermatogonial stem cells can be regulated as after cell loss self-renewal is increased. A major factor that plays a role in regulating stem cell fate is glial cell line derived neurotrophic factor (GDNF) produced by Sertoli cells which inhibits stem cell differentiation. Furthermore, spermatogonial stem cells are mainly localized to those tubule areas that border on the interstitial tissue. Apparently, also factor(s) from the interstitium, possibly testosterone, inhibit stem cell differentiation. Research on spermatogonial stem cells and the regulation of their differentiation may become facilitated by spermatogonial (stem) cell lines thathave recently been developed.