ABSTRACTAlthough many studies have been conducted on the increase in spike antibody levels after vaccination, there is insufficient prospective and longitudinal information on the BA.5-adapted bivalent vaccine up to the fifth vaccination. In this study, we conducted a follow-up study of spike antibody levels and infection history in 46 health care workers who received up to 5 vaccinations. Monovalent vaccines were administered for the first to fourth vaccinations, and a bivalent vaccine was administered for the fifth vaccination. 11 serum samples were collected from each participant, and antibody levels were measured in a total of 506 serum samples. During the observation period, 43 of the 46 health care workers had no infection history, and 3 had a history of infection. Spike antibody levels peaked at 1 week after the second booster vaccination and gradually declined until the 27th week after the second vaccination. After 2 weeks following the fifth BA.5-adapted bivalent vaccine, the spike antibody levels significantly increased (median: 23,756 [IQR: 16,450 to 37,326]), compared to those measured before vaccination (median: 9,354 [IQR: 5,904 to 15,784]) (paired Wilcoxon signed-rank test,P= 5.7 × 10−14). These changes in antibody kinetics were observed regardless of age or sex. These results suggest that booster vaccination increased the spike antibody levels. Regular vaccination is effective in maintaining long-term antibody levels.IMPORTANCEA COVID-19 bivalent mRNA vaccine was developed and administered to health care workers. The COVID-19 mRNA vaccine induces a robust antibody response. However, little is known about the antibody response to vaccines in serially collected blood samples from the same individuals. Here, we provide two-year follow-up data on the humoral immune response to COVID-19 mRNA vaccines in health care workers who received up to five vaccinations, including the BA.5-adapted bivalent vaccine. The results suggest that regular vaccination is effective in maintaining long-term antibody levels and have implications for vaccine efficacy and booster dose strategies in health care settings.