Effects of ultrasonic ice seeding and different concentrations of trehalose on the cryopreservation of L-02 hepatocytes were studied by replacing bio toxic dimethyl sulfoxide with trehalose in a self-built ultrasonic ice seeding device. The results show that the survival rate of hepatocytes (87.6±2.0)% is higher when cryopreserving L-02 hepatocytes with 0.3 mol/L trehalose by ultrasonic ice seeding , which is not significantly different from the traditional ice seeding group (84.3±1.0)%, and a significantly different from the non-ice seeding group (76.9±3.1)%.