Isolation, purification and culture of male germline stem cells from neo natal bovine testis

In order to establish the method of cell proliferation and differentiation of the newborn calf male germline stem cells(mGSCs) in vitro,calf testis were used in the present study.The method of collagenase and typin was sequentially performed to separate single-cell suspensions,and then used differential adhesion method and discontinued density gradient Percoll sorting to isolate the mGSCs and sertoli cells.For the mGSCs obtained were cultured in vitro.The result showed that the differential adhesion method would seperate the mGSCs and sertoli cells effectively;DMEM/F12 medium containing 2% fetal bovine serum could be used to cultrue mGSCs in vitro.In addition,the medium which added 100 ng.mL-1 GDNF could significantly improve the proliferation rate of mGSCs,and the mGSCs still had pluripotency at the some degree.