COVID-19 is a global threat with an increasing number of infections. Research on IgG seroprevalence among health care workers (HCWs) is needed to re-evaluate health policies. This study was performed in three pandemic hospitals in Istanbul and Kocaeli. Different clusters of HCWs were screened for SARS-CoV-2 infection. Seropositivity rate among participants was evaluated by chemiluminescent microparticle immunoassay. We recruited 813 non-infected and 119 PCR-confirmed infected HCWs. Of the previously undiagnosed HCWs, 22 (2.7%) were seropositive. Seropositivity rates were highest for cleaning staff (6%), physicians (4%), nurses (2.2%) and radiology technicians (1%). Non-pandemic clinic (6.4%) and ICU (4.3%) had the highest prevalence. HCWs in “high risk” group had similar seropositivity rate with “no risk” group (2.9 vs 3.5 p = 0.7). These findings might lead to the re-evaluation of infection control and transmission dynamics in hospitals.