The squamous cell carcinoma cell line NOKT-1 was successfully established from the right tongue of a 74-yearold Japanese man. Pathological diagnosis of the original tumor was moderately differentiated squamous cell carcinoma. NOKT-1 cells were transplanted subcutaneously into nude mice and xenograft was formed. In addition, the NOKT-1-XG cell line was established from the transplanted tumor of NOKT-1 cells. NOKT-1 cells and NOKT-1-XG cells were epithelial neoplastic and pleomorphic cells, which were similar. Immunocytochemistry revealed that NOKT-1 and NOKT-1-XG cells were CK17 and human mitochondria positive. To authenticate the NOKT-1 cell line and NOKT-1-XG cell line, we examined cross-contamination with other cell lines using short tandem repeat analysis, the results of which showed that NOKT-1 and NOKT-1-XG are new cell lines. Four of the 16 loci, corresponding to 25%, were different between these two cell lines, which indicates that the NOKT-1 genome was altered by transplantation. Moreover, in AM, NOKT-1 did not have a Y chromosome, whereas NOKT-1-XG had. Despite the genetic differences, a collagen gel droplet-embedded culture drug susceptibility test demonstrated that NOKT-1 cells derived from the original tumor and the NOKT-1-XG cell line had the same sensitivity. This cell line could be very useful for the development of immunotherapy and chemotherapy regimens and research on cancer etiology.