OBJECTIVES: Patients with cancer are at higher risk for severe COVID-19 infection. COVID-19 surveillance of workers in oncological centres is crucial to assess infection burden and prevent transmission. We estimate the SARS-CoV-2 seroprevalence among healthcare workers (HCWs) of a comprehensive cancer centre in Catalonia, Spain, and analyse its association with sociodemographic characteristics, exposure factors and behaviours. DESIGN: Cross-sectional study (21 May 2020-26 June 2020). SETTING: A comprehensive cancer centre (Institut Català d'Oncologia) in Catalonia, Spain. PARTICIPANTS: All HCWs (N=1969) were invited to complete an online self-administered epidemiological survey and provide a blood sample for SARS-CoV-2 antibodies detection. PRIMARY OUTCOME MEASURE: Prevalence (%) and 95% CIs of seropositivity together with adjusted prevalence ratios (aPR) and 95% CI were estimated. RESULTS: A total of 1266 HCWs filled the survey (participation rate: 64.0%) and 1238 underwent serological testing (97.8%). The median age was 43.7 years (p25-p75: 34.8-51.0 years), 76.0% were female, 52.0% were nursing or medical staff and 79.0% worked on-site during the pandemic period. SARS-CoV-2 seroprevalence was 8.9% (95% CI 7.44% to 10.63%), with no differences by age and sex. No significant differences in terms of seroprevalence were observed between onsite workers and teleworkers. Seropositivity was associated with living with a person with COVID-19 (aPR 3.86, 95% CI 2.49 to 5.98). Among on-site workers, seropositive participants were twofold more likely to be nursing or medical staff. Nursing and medical staff working in a COVID-19 area showed a higher seroprevalence than other staff (aPR 2.45, 95% CI 1.08 to 5.52). CONCLUSIONS: At the end of the first wave of the pandemic in Spain, SARS-CoV-2 seroprevalence among Institut Català d'Oncologia HCW was lower than the reported in other Spanish hospitals. The main risk factors were sharing household with infected people and contact with COVID-19 patients and colleagues. Strengthening preventive measures and health education among HCW is fundamental.