To evaluate the anti-tumor effect of PFP@Polymer NPs activated by dual-frequency HIFU irradiation in vitro, 4T1 cells were incubated with PFP@Polymer NPs and irradiated with single-frequency HIFU (1.1 or 5 MHz) and dual-frequency HIFU (1.1 and 5 MHz), respectively. This evidence concerns the gene PRF1 and neoplasm.