Spermatogonial stem cells (SSCs) reside in certain niches inside seminiferous tubules. These niches are believed to secrete chemotactic aspects for SSCs, since SSCs migrate to them upon transplantation. Nevertheless, the identity of these chemotactic molecules stays unknown. Here, we established a testis feeder cell moreover culture method and utilised it to determine SSC Dilution Calculator chemotactic aspects. When seeded on testis cells from infertile mice, SSCs migrated beneath the Sertoli cells and formed colonies having a cobblestone physical appearance that were pretty just like these created by hematopoietic stem cells. Cultured cells maintained SSC action and fertility for at the very least 5 months. Cobblestone colony formation depended on GDNF and CXCL12, and dominant-negative GDNF receptor transfection or CXCL12 receptor deficiency lowered SSC colonization. Additionally, GDNF upregulated CXCL12 receptor expression, and CXCL12 transfection in Sertoli cells improved homing efficiency. General, our findings identify GDNF and CXCL12 as SSC chemotactic elements in customer reviews vitro and in vivo.