Abstract T follicular helper (Tfh) cells and cytokine IFN-γ are involved in Ig class switching in B cells to produce IgG2a(c). Tfh cells mainly produce IL-21 as their signature cytokine, and several studies have also shown that some Tfh cells are capable of expressing Th1 signature cytokine IFN-γ without expressing the Th1 master transcription factor T-bet. In Toxoplasma gondii peptide AS15/Complete Freund’s Adjuvant (CFA)-induced immune response, we found antigen AS15-specific IgG2a(c) were abolished in CD4-Cre-mediated T-bet-deficient mice. By using T-bet-fate-mapping mouse strains, we reported that transient expression of T-bet epigenetically imprints the Ifng locus for cytokine production in a subset of Tfh cells; In germinal centers, multi-color tissue imaging revealed that the ex-T-bet Tfh cells express IFN-γ in situ; IFN-γ-expressing Tfh cells are absent in T-bet-deficient mice. More interestingly, we noticed that IFN-γ-expressing Tfh cells were fully present in mice with T-bet deletion at late stages of T cell differentiation, and antigen AS15-specific IgG2a(c) were even further up-regulated in such “T-bet late knockout” mice. Taken together, our findings demonstrate that IFN-γ expressed by T cells plays a dominant role in antibody IgG2a(c) production; T-bet expression is fine-tuned as spatiotemporal manner for balancing Th1 and Tfh cell differentiation in vivo and regulating the strength of antibody production.