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Background: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a constantly evolving virus, resulting in an increased burden on the existing COVID-19 vaccines. Healthcare workers (HCWs) are the first line of defense against the coronavirus disease 2019 (COVID-19) pandemic and have been prioritized among the risk categories receiving the COVID-19 vaccine. This work aimed to investigate the maintenance of antibody response of the Oxford−AstraZeneca vaccine (ChAdOx1/nCoV-19). Methods: Anti-spike immunoglobulin G (IgG) was measured at baseline point (immediately prior to vaccination) and 12- and 24-week (w) points following vaccination. Adverse reactions to the vaccine were reported. Participants were followed up for the incidence of COVID-19 during the 12 w interval between vaccination doses for 24 w after the second dose. Results: A total of 255 HCWs participated in the study. Prior to vaccination, 54.1% experienced COVID-19, 88.2% were seropositive after the first dose, while seropositivity reached 95.7% after the second dose. Following the first and second doses, the anti-spike IgG serum level was significantly higher in subjects with past COVID-19 than in others (p < 0.001 and =0.001, respectively). Conclusions: The Oxford−AstraZeneca vaccine is generally safe and provides a highly effective long-term humoral immune response against the Delta and Omicron variants of SARS-CoV-2.