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The aim of this study was to evaluate Photodynamic therapy PDT using diode laser at wavelength (650 nm) with methylene blue MB as a photosensitizer in vitro and in vivo . The cytotoxic effect of PDT with MB was investigated in vitro on murine leiomyosarcoma (L20B) and rhabdomyosarcoma (RD ) cell lines. In this study the cell lines seeded separately, they were divided into two main groups : control group (A) left without any treatment while the second group (B) group of test. Histopathological sections in control group showed the presence of tumor growth characterized by formation of acinar like structures, with pleomorphic tumor cells and hyperchromatic nuclei with high numbers of mitotic figures. Histopathological sections in groups of tumor-bearing female mice, treated with 1% MB+ Laser showed absence of tumor mass replaced by fibrous connective tissue, mononuclear cell infiltration and hyperplasia of lymphoid tissue of white pulp of the spleen . To sum it up, diode laser induce the best photodynamic therapeutic effect on cancer cells in vitro and in vivo after cells photosensitization with MB 1% and 30 min of exposure time.