Objectives Risk of reinfection with SARS-CoV-2 among health-care workers (HCWs) is unknown. We assessed the incidence rate of SARS-CoV-2 reinfection in the real-life setting of a longitudinal observational cohort of HCWs from the Hôpital Européen Georges Pompidou, Assistance Publique-Hôpitaux de Paris, France, during the first and second waves of COVID-19 epidemic. Methods From March to December 2020, HCWs were subjected to molecular and serology testing of SARS-CoV-2. Reinfection was defined as a positive test result during the first wave, either by serology or PCR, followed by a positive PCR during the second wave. Evolution of COVID-19 status of HWCs was assessed by a Sankey diagram. Results A total of 7765 tests (4579 PCR and 3186 serology) were carried out and 4168 HCWs had at least one test result during the follow-up period with a positivity rate of 15.9%. No case of reinfection during the second wave could be observed among 102 positive HCWs of the first wave, nor among 175 HCWs found positive by PCR during the second wave who were negative during the first wave. Conclusions SARS-CoV-2 reinfection was not observed among HCWs, suggesting a protective immunity against reinfection that lasts at least 8 months post infection.