Objective： To isolate and identify Sertoli cells from mouse testis. Methods： Testis were isolated from male mouse aged 18-20 days old and were cultured by enzymatic digestion. The Sertoli cell gene with a zinc finger domain （SERZ） was obtained by RT-PCR and was inserted into pcDNA3.0 to construct recombinant plasimid pcDNA3.0-SERZ, pcDNA3.0- SERZ was then cleaved by restriction endonuclease Kpn I and Xba I to obtain the linear templets for preparation of the probes. The expression of SERZ mRNA in the cultured cells was analyzed by in situ hybridization with the prepared probes. The expression of androgen binding protein （ABP） mRNA in the cultured cells was detected by RT-PCR. Results： Under light microscope, most Sertoli cells were polygonal and were completely extended,mimicking a membrane. The nuclei were triangular or irregular, weakly stained and with obvious nucleoli. The neighbouring cells were interlaced with one another and the cell purity was （85.1± 2.5） %. SERZ mRNA was highly expressed in the cytoplasm of the cultured cells. RT-PCR showed that ABP mRNA was expressed in the cultured cells. Conclusion： We have successfully isolated and identified Sertoli cells from mouse testis, with the cell purity being （85.1±2. 5）%.