Objective To establish an efficient and stable method for protein extraction of Streptococcus mutans. Methods The collected bacteria were treated by freeze-thaw and ultrasonic(method 1), ultrasonic(method 2), boiling (method 3), boiling and ultrasonic(method 4), respectively. The index such as state of bacteria broken, concentration of extracted protein and SDS-PAGE of protein were employed to evaluate the effects of above four methods. Results Beside the method 3, the other three methods could break the bacteria effectively, of which ultrasonic was the key factor. The pattern of SDS-PAGE which treated by method 1, method 2 and method 4 was almost same, but method 1 resulted in the best abundance. There was significantly difference among the four protein concentration extracted by four methods(P