Objective To study the antitumor effects of exosomes derived from heat-shocked E.G7-OVA tumor cells in vivo. Methods Exosomes derived from E.G7-OVA tumor cells were isolated and purified by serial centrifugation and sucrose gradients ultracentrifugation. Exosomes from heat-shocked or non-heat-shocked E.G7-OVA tumor cells were named as Exo/HS and Exo correspondingly. Exosomes were viewed by electron microscopy. Protein components of exosomes were detected by Western blot. Exo, Exo/ HS or PBS were injected into mice before injection of E.G7-OVA tumor cells, and antitumor effects were ob-served in each group. Mouse model bearing E.G7-OVA tumor cells were established to examine immunother-apy effects of Exo or Exo/HS. Cytotoxity of spleen CTL were measured by LDH. Results Exosomes con-tained bi-layer membrane and their diameters are between 40 nm and 100 nm under electron microscopy. The Western blot results showed that HSC70, HSP70, HSP60, HSP90, MHC Ⅰ and OVA were present in both Exo and Exo/HS. However, Exo/HS contained more HSP70 and MHC Ⅰ than Exo. Protective antitu-mor immunity suggested that tumor-free survival (90 days) rate in Exo/HS vaccinated mice was significantly higher than those in Exo or PBS vaccinated mice (50%, 20%, 0%, P<0.01). Therapeutic antitumor effects showed that immunization by Exo/HS resulted in dramatically enhanced antitumor effects when com-pared to the Exo- or PBS-treated groups (P<0.01). CTL results showed that immunization with Exo/HS in-duced higher level of OVA-specific CTL responses as compared with those from Exo or PBS (P<0.01). Conclusion Exosomes derived heat-shocked E.G7-OVA tumor cells may be used as potent cancer vaccine.