Infection with the ubiquitous parasite Toxoplasma gondii is a threat for immunocompromised patients and pregnant women and effective immune-prophylaxis is still lacking. Here we tested a mixture of recombinant T. gondii antigens expressed in different developmental stages, i.e., SAG1, MAG1 and GRA7 (SMG), and a lysate derived from T. gondii tachyzoites (TLA) for prophylactic vaccination against cyst formation. Both vaccine formulations were applied systemically followed by an oral TLA-booster in BALB/c mice. Systemic priming with SMG and oral TLA-booster did not show significant induction of protective immune responses. In contrast, systemic priming and oral booster with TLA induced higher levels of Toxoplasma-specific IgG, IgG1 and IgG2a in sera as well as high levels of Toxoplasma-specific IgG1 in small intestines. Furthermore, high levels of Toxoplasma-specific Th1-, Th17- and Th2-associated cytokines were only detected in restimulated splenocytes of TLA-vaccinated mice. Importantly, in mice orally infected with T. gondii oocysts, only TLA-vaccination and booster reduced brain cysts. Furthermore, sera from these mice reduced tachyzoites invasion of Vero cells in vitro, indicating that antibodies may play a critical role for protection against Toxoplasma infection. Additionally, supernatants from splenocyte cultures of TLA-vaccinated mice containing high levels of IFN-γ lead to substantial production of nitric oxide (NO) after incubation with macrophages in vitro. Since NO is involved in the control of parasite growth, the high levels of IFN-γ induced by vaccination with TLA may contribute to the protection against T. gondii. In conclusion, our data indicate that prime-boost approach with TLA, but not with the mixture of recombinant antigens SMG, induces effective humoral and cellular Toxoplasma-specific responses and leads to significant reduction of cerebral cysts, thereby presenting a viable strategy for further vaccine development against T. gondii infection.