The efficacy and effectiveness of influenza vaccines depend primarily on the vaccine recipient and the virus similarity to the endemic virus. Regulatory T cells (Tregs) and cytokines are known to restrict immune responses against viral infections. We conducted this study to explore the role of Tregs, cytokines, and antibody production after influenza vaccination. The whole blood was collected from healthy subjects (n = 36) before and two weeks after influenza vaccine immunization for two or three consecutive years. The cell surface markers, intracellular staining of Foxp3 (+) Tregs, and Th1/Th2 cytokines were determined. The antibody titer was detected using the hemagglutination inhibition test. The CD3 (+) , CD127 (+) , CD4 (+) CD25 (+) and CD4 (+) Foxp3 (+) cells were increased significantly post vaccination. The plasma level of the transforming growth factor (TGF-β), but not interleukin (IL)-2, IL-4, IL-5, IL-10, IFN-γ, TNF-α, was also found to increase significantly after vaccination. We further correlated the cytokine fold-increases with the anti-influenza antibody titer for individual post vaccination. It was found that the IL-10 level after vaccination correlated with the fold-increases of anti-H1N1, anti-H3N2, anti-B/Yamagata, and anti-B/Victoria antibodies. But, a negative relationship occurs between the TGF-β level and fold-increases of anti-H1N1, anti-H3N2, anti-B/Yamagata, and anti-B/Victoria antibodies post vaccination. Treg cells and TGF-β seem to participate in the downregulation of the anti-influenza antibody response post influenza vaccination. Alteration of Treg activity might enhance influenza vaccine antibody responses and efficacy.