Objective:To validate fatty acid amide hydrolase (FAAH) expression in mouse spermatogonial stem cells (SSCs). Methods: We established a stable mouse SSCs line in vitro and verified FAAH expression in various tissues of mice and the cell line by Western blotting assay. Immunofluorescence was performed to detect FAAH location in the testis and SSCs. Results: Western blotting assay showed that FAAH was highly expressed in testicular tissues. Double-immunofluorescent staining showed that FAAH located on the membrane of mouse SSCs and co-localized with SSCs separation marker, GDNF family receptor alpha-1 (GFRα1). Conclusion: FAAH is a new candidate separation surface marker of SSCs, which provides a basis for the research of FAAH function in spermatogenesis.