Melanoma is an aggressive disease that arises from mutations in the cells that produce the pigment melanin, melanocytes. Melanoma is characterized by a high mortality rate, due to avoidance of applied therapies and metastasis to other organs. The peculiar features of boron neutron capture therapy (BNCT), particularly its cell-level selectivity, make BNCT a promising modality for melanoma treatment. However, appropriate cellular models should be used to study new therapies or improve the efficacy of existing therapies. Spheroids, which have been used for years for in vitro studies of the efficacy of anti-cancer therapies, have many characteristics shared with tumors through which they can increase the accuracy of the cellular response compared to 2D culture in vitro studies and reduce the use of animals for research in the future. To the best of our knowledge, when we started researching the use of spheroids in BNCT in vitro, there was no publication showing such use. Our study aimed to evaluate the efficacy of a 3D cellular model (spheroids) for testing BNCT on melanoma cells. We assessed boronophenylalanine (