•The average SARS-CoV-2 seroprevalence in healthcare workers of a a tertiary hospital, University Hospital Essen, Germany was 1.6 % in the period between March to April 2020.•The local hygiene standards seem to be effective to prevent patient-to-staff virus transmission at the designated COVID-19 units wards.•Awareness for COVID-19 infections is crucial even in non−COVID-19 wards. The novel coronavirus SARS-CoV-2 is associated with a severe respiratory manifestation, COVID-19, and presents a challenge for healthcare systems worldwide. Healthcare workers are a vulnerable cohort for SARS-CoV-2 infection due to frequent and close contact to patients with COVID-19. Serum samples from 316 healthcare workers of the University Hospital Essen, Germany were tested for SARS-CoV-2-IgG antibodies. A questionnaire was used to collect demographic and clinical data. Healthcare workers were grouped depending on the frequency of contact to COVID-19 patients in high-risk-group (n = 244) with daily contact to known or suspected SARS-CoV-2 positive patients, intermediated-risk-group (n = 37) with daily contact to patients without known or suspected SARS-CoV-2 infection at admission and low-risk-group (n = 35) without patient contact. In 5 of 316 (1.6 %) healthcare workers SARS-CoV-2-IgG antibodies could be detected. The seroprevalence was higher in the intermediate-risk-group vs. high-risk-group (2/37 (5.4 %) vs. 3/244 (1.2 %), p = 0.13). Four of the five subject were tested negative for SARS-CoV-2 via PCR. One (20 %) subject was not tested via PCR since he was asymptomatic. The overall seroprevalence of SARS-CoV-2 in healthcare workers of a tertiary hospital in Germany is low (1.6 %). The data indicate that the local hygiene standard might be effective.