In this study, we described the establishment of the HNSCC primary cell line designated 132P1 derived from well-differentiated, localized oral squamous cell carcinoma tissue obtained by the primary tumor biopsy and also establishment and characterization of CD44+/CD90−, CD44−/CD90−, CD44+/CD90+, and CD44−/CD90− cell subpopulations derived from this primary cell line 132P1. Here, CD44 is linked to head and neck squamous cell carcinoma.