We recently established a unique therapeutic modality for musculoskeletal sarcomas, combining acridine orange (AO) with photodynamic surgery (PDS), photodynamic therapy (PDT) and radiodynamic therapy (RDT); excellent results were obtained in the inhibition of local tumor recurrence after intralesional excision. However, AO is not yet approved for clinical use and intravenous injection. Therefore, methylene blue (MB), which has a very similar chemical structure to AO and is already in clinical use for other diseases, was investigated. In vitro studies using mouse osteosarcoma (LM8) cells revealed that MB-PDT had a strong cytocidal effect and that MB was not radiosensitive, showing no effect in RDT. In vivo studies showed that MB did not specifically accumulate in mouse osteosarcoma tissue and that it did not inhibit tumor growth. MB is not a better photosensitizer than AO in PDS, PDT and RDT for osteosarcoma.