Background: The objective of this study was to explore the effects of 30 kHz ultrasound on the efficacy of paclitaxel in subcutaneous breast tumors in Balb/c mice in vivo. Materials and Methods: 40 Balb/c female mice were divided into 5 groups: model control group, paclitaxel intraperitoneal (ip) group (20 mg/kg, ip, at 3 day intervals for a total of 3 doses (q3d×3)), paclitaxel intratumoral (it) group (20 mg/kg, it, q3d×3), paclitaxel intraperitoneal (20 mg/kg, ip, q3d×3) combined with low-frequency ultrasound (LFU) group, and paclitaxel intratumoral (20 mg/kg, it, q3d×3) combined with LFU group. Ultrasound parameters were 30 kHz, 200 MW intensity, 15 min, q3d×3. The antitumoral effect was determined by examining the tumor weight in subcutaneously inoculated EMT6 breast carcinoma models in Balb/c mice. All subcutaneous tumors were examined using high performance liquid chromatography (HPLC) upon completion of the experiments. Drug concentrations in the subcutaneous tumors were also analyzed using HPLC. Finally, paraffin sections of the subcutaneous tumors were made, and after hematoxylin and eosin staining, the tumor morphology was examined under a light microscope. Results: LFU combined with paclitaxel significantly restrained tumor growth in transplanted subcutaneous EMT6 tumors in Balb/c mice, and this effect correlated with increased local concentrations of paclitaxel in the tumors. Body weight measurement did not reveal significant adverse effects on the Balb/c mice during the study. Conclusion: LFU combined with paclitaxel has a significant synergistic effect in the treatment of breast cancer.