Background: Knowledge about the COVID-19 outbreak is still sparse, especially in a cross-national setting. COVID-19 is caused by a SARS-CoV-2 infection. The aim of the study is to contribute to the surveillance of the pandemic by bringing new knowledge about SARS-CoV-2 seropositivity among healthcare workers. It seeks to evaluate whether certain job functions are associated with a higher risk of being infected and to clarify if such association is mediated by the number of individuals that employees meet during a workday. In addition, we investigate regional and national differences in seroprevalence. Methods: This research involved a bi-national prospective observational cohort study including 3272 adults employed at Falck in Sweden and Denmark. Participants were tested for SARS-CoV-2 antibodies every second week for a period of 8 weeks from 22 June 2020 until 10 August 2020. Descriptive statistics as well as multivariable logistic regression analyses were applied. Results: Of the 3272 Falck employees participating in this study, 159 (4.9%) tested positive for SARS-CoV-2 antibodies. The seroprevalence was lower among Danish Falck employees than among those from Sweden (2.8% in Denmark and 8.3% in Sweden). We also found that the number of customer or patient contacts during a workday was the most prominent predictor for seropositivity and that ambulance staff was the most vulnerable staff group. Conclusion: Our study presents geographical variations in seroprevalence within the Falck organization and shows evidence that social interaction is one of the biggest risk factors for becoming infected with SARS-CoV-2.