Owing to the limitations of traditional Bacillus Calmette-Guerin vaccines and the low efficacy of DNA vaccines expressing single antigens of Mycobacterium tuberculosis, there is a pressing requirement for adjuvants capable of strengthing the immunogenicity and effectiveness of these vaccines against tuberculosis (TB). T-bet (TBX21) is a transcription factor, which controls the optimal development of type-1 immune responses responsible for the potent protection of vaccines against TB. However, little is known about the efficiency of the TB vaccine combined with T-bet. In this study, we report an approach to intensify the immunogenicity of Ag85B DNA-based vaccines using T-bet as an adjuvant. Balb/c mice were immunized by 3 intramuscular inoculations with pcDNA3.1-FLAG-T-bet in combination with pcDNA3.1-FLAG-Ag85B, and the immune responses were compared with those induced by vaccination with Ag85B DNA alone. We found that pcDNA3.1-T-bet-Ag85B not only induced evidently higher IgG2a antibody responses, but also increased the production of interferon-γ (IFN-γ) and interleukin (IL)-2 with the concomitant repression of IL-4 and IL-10 compared with pcDNA3.1-Ag85B alone or the empty vector. Thus, plasmid DNA coding for T-bet enhanced Ag85B-specific immune responses and shifted them to a predominant Th1-type immune response. In conclusion, T-bet is an efficacious Th1-inducing adjuvant in the context of the Ag85B DNA-based vaccination, and could also prove to be a promising candidate for DNA vaccine development against TB.