Abstract: To provide reference for the clinical application of focused ultrasound in tumor treatment, the minimum thermal dose of irreversible thermal damage in isolated biological tissues under the action of focused ultrasound is studied in this paper. In this study, a high-resolution infrared camera is used to measure the temperature distribution of tissues during focused ultrasound irradiation. Based on the thermal damage image and the temperature change rule, the dose required for thermal damage of biological tissues is analyzed, and the results obtained by different thermal damage prediction methods are compared. The experimental results show that by using the "threshold method", when the tissue temperature exceeds 51 ℃, in vitro tissue rapidly undergoes thermal damage. The thermal damage threshold obtained using T_index is T₉₀=56 ℃. Using the equivalent thermal dose model t_CEM43, the minimum thermal dose that can cause thermal damage is (4.7 ± 1.9) min. Using the cumulative equivalent time of T₉₀ (t_CEM43,T90), the minimum heat dose for thermal damage is (215 ± 96) min. Research has shown that using T₉₀=56 ℃ as the thermal dose threshold has higher confidence, and using t_CEM43,T90 as the quantitative method for thermal dose has stronger robustness. The research in this paper can provide experimental basis for ultrasound hyperthermia and thermal injury evaluation, which is helpful for the clinical application of focused ultrasound.