MOESM2 of The Study of Enhanced High-Intensity Focused Ultrasound Therapy by Sonodynamic N2O Microbubbles

Additional file 2: Figure S2. Determination of optimal LIFU irradiation time. (The data were shown as mean ± SD, n = 5 per group, *p < 0.05). To Select the optimal time for ultrasound, MDA-MB-231 cells were seeded in a 96-well plate at a density of 5000 cells per well for 24 h. And then use the strength of 2 W/cm2 ultrasonic processing cells, respectively for the 50 s, 100 s, 150 s and 200 s. After the indicated treatments, CCK8 solution (10 μL) was added in each well. Then, the MDA-MB-231 cells were cultured at 37 °C, 5% CO2 cell incubation box to save. At 3 h, 6 h, 9 h and 12 h, cell viabilities were determined by CCK-8 assay. The optical density of each well was measured at 450 nm with a Bio-Tek microplate reader. Cell viability of the control group was defined as 100%. The test results show (Figure S1) that the cells viability in 50 s and 100 s groups were more than 95% at 3 h, 6 h, 9 h and 12 h after treatment, and the difference in cell viability between the two groups is not statistically significant (p > 0.05). The cell viability decreased in 150 s and 200 s groups (p < 0.01), and the survival rate was 74% in 150 s group and 55% in 200 s group. Combined with the oxygen free radicals generated condition, 100 s was selected as the optimal time of ultrasound.