In Italy, from January 2021, the Ministry of Health indicated a vaccination plan against COVID for frail patients and physicians based on a three-dose scheme. However, conflicting results have been reported on which biomarkers permit immunization assessment. We used several laboratory approaches (i.e., antibodies serum levels, flow cytometry analysis, and cytokines release by stimulated cells) to investigate the immune response in a cohort of 53 family pediatricians (FPs) at different times after the vaccine. We observed that the BNT162b2-mRNA vaccine induced a significant increase of specific antibodies after the third (booster) dose; however, the antibody titer was not predictive of the risk of developing the infection in the six months following the booster dose. The antigen stimulation of PBMC cells from subjects vaccinated with the third booster jab induced the increase of the activated T cells (i.e., CD4+CD154+); the frequency of CD4+CD154+TNF-α+cells, as well as the TNF-α secretion, was not modified, while we observed a trend of increase of IFN-γ secretion. Interestingly, the level of CD8+IFN-γ+(independently from antibody titer) was significantly increased after the third dose and predicts the risk of developing the infection in the six months following the booster jab. Such results may impact also other virus vaccinations.