Cancer/testis (CT) antigens are potential targets for cancer immunotherapy, with NY-ESO-1 being among the most immunogenic. In several clinical trials in malignant melanoma (MM) patients, NY-ESO-1 protein/peptides showed clear evidence of inducing specific immunity. However, little is known about NY-ESO-1 expression in primary and metastatic MM and its relationship to disease progression. We analyzed NY-ESO-1 expression immunohistochemically in a series of primary and metastatic MMs and its relation to prognostic parameters and survival. We studied 61 primary and 63 metastatic MM specimens (from 61 and 56 patients, respectively). The prevalence of NY-ESO-1 expression was significantly higher in metastatic versus primary tumors [18/56 (32%) versus 8/61 (13%), P = 0.015]. There was a significant association between initial stage at presentation and NY-ESO-1 expression [stage I (3.45%), stage II (9.52%) and stage III (45.45%), P = 0.0014]. Primary MMs expressing NY-ESO-1 were significantly thicker than NY-ESO-1 negative cases (median thickness 4.7 mm versus 1.53 mm respectively, P = 0.03). No significant difference was seen in overall survival. In conclusion, NY-ESO-1 is more frequently expressed in metastatic than in primary MM and its expression is associated with thicker primary lesions and a higher frequency of metastatic disease, indicative of a worse prognosis. Our study suggests that patients with metastatic MM who express NY-ESO-1 may benefit from NY-ESO-1-based immunotherapy.