Increased expression of aldehyde dehydrogenase 1 (ALDH1) has recently been reported in several cancers. However, whether member A1 of aldehyde dehydrogenase 1 (ALDH1A1) is involved in the formation of nasopharyngeal carcinoma (NPC) remains unknown. To investigate the expression of ALDH1A1 in NPC and its association with the tumorigenesis of NPC, we examined the expression of ALDH1A1 in NPC specimens using immunohistochemistry (IHC), quantitative RT-PCR (qRT-PCR) and Western blot. Moreover, we sorted ALDH1A1(high) cells from NPC cell line CNE-2 by flow cytometry and examined the expression of primitive embryonic stem cell markers OCT4, SOX2 and Nanog. Finally, we investigated the capacities of growth, proliferation, colony- formation and tumorigenesis of ALDH1A1(high) cells in vitro and in vivo. We found ALDH1A1 was significantly increased in human NPC samples via IHC, qRT-PCR and Western blot (p < 0.05). ALDH1A1(high) cells sorted from NPC cell line CNE-2 by flow cytometry had higher expression of primitive embryonic stem cell markers OCT4, SOX2 and Nanog, and showed enhanced capacities of growth, proliferation, colony formation and tumorigenesis in vitro and in vivo when compared with ALDH1A1(low) cells (p < 0.05). Our findings indicated that increased expression of ALDH1A1 in NPC was associated with enhanced invasiveness.