At present, very little is known about the types and heterogeneity of T cell responses and immunodominant epitopes of melanoma-associated antigens at coexisting sites of primary melanoma and metastatic lesions. To address this issue, we compared the T cell receptor (TCR) gene usage, complementary-determining region 3 diversity, and melanoma-associated antigens expression patterns of primary and metastatic melanoma specimens from three patients with partially homologous HLA class-1 types. Results obtained showed an overall predominance of a very limited number of TCRV regions with AV13 and BV14 being most frequently overexpressed. Sequencing of the dominating TCR transcripts confirmed the restricted usage of certain TCR specificities and, in two of the three patients, identified several identical TCR clonotypes at more than one metastatic site. Nevertheless, we failed to detect TCR transcripts that were common to all tumor deposits in a given patient and, within the majority of coexisting metastases, tumor-infiltrating lymphocytes preferentially used individual site-specifically expanded TCR beta-chain VJ segment combinations. This occurrence of individual responses simultaneously executed at and influenced in their specificity by the different sites of tumor growth, has important implications for the type of strategies chosen in the development of efficacious vaccines for patients with metastatic melanoma.