Background: Large volumes of sitting time have been associated with multiple health risks. To reduce sitting time of office workers working for a Dutch insurance company, the Dynamic Work intervention was developed. The primary objective of this paper is to describe the study protocol of the Dynamic Work study, which aims to evaluate if this multicomponent intervention is (cost-)effective in reducing total sitting time on the short-term (≈3 months) and longer-term (≈12 months) compared to usual practice. Methods/design: This two-arm cluster randomized controlled trial will recruit 250 desk-based office workers working at different locations of an insurance company in the Netherlands. After baseline measurements, departments will be matched in pairs and each pair will be randomly assigned to the control or intervention condition. The multicomponent intervention contains organizational (i.e. face to face session with the head of the department), work environmental (i.e. the introduction of sit-stand desks and cycling workstations), and individual elements (i.e. counselling and activity/sitting tracker with a self-help program booklet). The counselling involves two group intervention sessions and four on-site department consultations with an occupational physiotherapist. Sitting time (primary outcome), upright time and step counts will be assessed objectively using the activPAL activity monitor at baseline, short-term (approximately 3 months) and longer-term (12 months). Other outcomes will include: self-reported lifestyle behaviours, anthropometrics, work-related outcomes (i.e. absenteeism, presenteeism, work performance, work-related stress), health-related outcomes (i.e. vitality, musculoskeletal symptoms, need for recovery, quality of life), and costs from both company and societal perspective. The study will include economic and process evaluations. Discussion: This study will assess the longer-term (cost-) effectiveness of a multicomponent workplace intervention aimed at reducing sitting time in comparison with usual practice. Furthermore, the process evaluation will provide insights in factors associated with successful implementation of this intervention. Trial registration: ClinicalTrials.gov NCT03115645; Registered 13 April 2017. Retrospectively registered.