[Objective] The aim of this study was to express Mycobacterium tuberculosis Ag85A protein and to evaluate its immunogenicity in mice. [Methods] The cold expressed system and chaperone competent cells BL21 were combined to express Mycobacterium tuberculosis Ag85A protein, then the protein was purified with affinity chromatography and identified by Western Blot analysis. [Results] The immunogenicity of the purified Ag85A protein was evaluated in C57BL/6 mice. Results show that high level of specific IgG was elicited in the serum, and the splenocytes and lymph node cells of immunized mice could produce more Th1 cytokines, such as IFN-γ and TNF-α, after stimulated with specific antigen. [Conclusions] Ag85A protein can induce strong specific humoral and Th1 type cellular immune responses, providing an important biological material for further research and application.