Abstract Background The need for interprofessional collaboration has been emphasized by health organizations. This study was part of a mixed-methods evaluation of interprofessional teamwork modules implementation in an emergency department (ED), where a major intervention was didactic training of team roles and behaviours in combination with practice scenarios. The aim of the study was to evaluate the implementation of interprofessional teamwork modules from a staff perspective and focus on how implementation fidelity may be sustained. Methods In this mixed-methods case study we triangulated staff data from structured observations, semi-structured interviews, and a questionnaire repeated at intervals over 5 years. A protocol of key team behaviours was used for the observations conducted in June 2016 and June 2018, 1½ and 3½ years after the initial implementation. A purposeful sample of central informants, including nursing and medical professionals and section managers, was interviewed from May to June 2018. The interview guide consisted of open-ended questions about the experiences of interprofessional teamwork modules and the implementation process. The questionnaire consisted of five statements about the perceived workload, interprofessional collaboration and patient satisfaction, where each was rated on a Likert scale. Results Good fidelity to four out of five key team behaviours was observed during the first year. However, fidelity was sustained only for one key team behaviour after 3 years. We conducted a qualitative content analysis of 18 individual interviews. The theme Enjoying working together, but feeling less efficient emerged of the interprofessional teamwork modules, despite shorter ED stays for the patients. Negative experiences of the staff included passive team leaders and slow care teams. The theme Stimulating to create, but challenging to sustain emerged of the implementation process, where barriers were not adressed and implementation fidelity not sustained. The staff questionnaire showed that the perceived work conditions was improved in periods of high fidelity, but deteriorated to pre-implementation levels as fidelity to the key team behaviours decayed in 2018. Conclusions Extensive planning and successful initial implementation were not enough to sustain the key behaviour changes in the study. The use of implementation frameworks can be helpful in future projects.