The present study detected p75 neurotrophin receptor (p75NTR) expression in tongue squamous cell carcinoma (TSCC) cell lines, in order to define the biological properties of p75NTR+ cells and to confirm the use of p75NTR+ as a surface marker for TSCC stem cells. p75NTR+ cells were separated from Tca-8113 and CAL-27 TSCC cells by fluorescence-activated cell sorting. Colony formation, MTT and scratch assays, and a tumorigenicity analysis were performed to measure self-renewal and proliferation, multidirectional differentiation, and tumorigenicity of p75NTR+ cells. p75NTR+ cells comprised 3.1 and 1.9% of Tca-8113 and CAL-27 cells (mean of three experiments), respectively, and were more able to form colonies compared with non-sorted cells (P<0.01). In addition, the proportion of p75NTR+ cells generated from monoclonal p75NTR+ cells decreased to 14.5 (Tca-8113) and 5.8% (CAL-27) of cells within 2 weeks, thus suggesting that p75NTR+ cells are able to generate p75NTR+ and p75NTR− cells. Furthermore, p75NTR+ cells exhibited increased proliferation, as evidenced by MTT assay (P<0.01) and had greater metastatic ability according to the scratch assay (P<0.01), compared with non-sorted cells. p75NTR+ cells also exhibited a greater tumorigenic capacity compared with non-sorted cells. In conclusion, p75NTR+ cells isolated from TSCC cell lines possess the characteristics of cancer stem cells; therefore, p75NTR may be considered a useful surface marker for the identification of TSCC stem cells.