Introduction The COVID-19 pandemic had a major impact on the mental health of workers, in particular healthcare professionals. The social isolation used as a mitigation measure against infection contributed to a reduction in sleep quality, and an increase in depressive symptoms, anxiety, and burnout. Objectives The aims of this study are to assess the impact of isolation and quarantine on the mental health and sleep quality of healthcare professionals in a Portuguese Oncology Institute. Material and Methods An online form was distributed to all healthcare professionals who were in isolation or quarantine during the year 2020 in the IPO-Porto, consisting of three instruments- sociodemographic survey, Mental Health Inventory (MHI) and Basic Insomnia and Sleep Quality Scale (BaSIQS). Data were collected in two moments, from July 25 to December 25, 2020, and in December 2021. Results A total of 359 healthcare professionals were identified (151 by isolation and 208 by quarantine). In the 2020 analysis, 115 responses were obtained (54 by isolation and 61 by quarantine), resulting in a response rate of 32.0%. In the 2021 analysis, 76 responses were obtained with a response rate of 21.1%. Regarding the healthcare professionals in our study, the dimensions that make up the psychological stress of the MHI, show an improvement after the first year of the pandemic, as well as an improvement in the mean total MHI value from 2020 to 2021. Sleep quality (assessed by the BaSIQS) is in the poor-to-intermediate category in the 2020 assessment. Although the 2021 assessment shows a slightly better mean, there is no statistically significant difference between the two. In this sample of health professionals, mental health and sleep quality correlated for the instruments used. Discussion and conclusions Insomnia and sleep quality are a modifiable factor of mental health, with an impact on the quality of life and productivity of healthcare professionals. Social isolation as a disruptor of sleep, daily routine and healthy interaction in work teams may be a factor that aggravates psychological distress. The investment in mental health promotion programs, as well as the coordination of multidisciplinary teams composed of Psychology, Medicine and Occupational Nursing are essential in the monitoring and surveillance of these workers’ health. KEY WORDS: Mental Health, Healthcare Professionals, Occupational Psychology, COVID-19, Occupational Medicine, Occupational Health.