ABSTRACT Objectives: to assess the effect of an educational intervention based on clinical simulation on nursing professionals’ compliance with practices to prevent peripherally inserted central venous catheter-associated primary bloodstream infections in a Neonatal Intensive Care Unit. Methods: a quasi-experimental study, with preand post-intervention assessment with a single group. The population consisted of 41 nursing professionals, with 31 observations being made before and after the intervention. Analyzes were performed using descriptive statistics and the McNemar non-parametric test. A significance level of 5% was adopted. Results: after the intervention, there was an increase in compliance with prevention practices of surgical antisepsis and professional hand hygiene, skin antisepsis with chlorhexidine, waiting for the time of the effect of alcoholic chlorhexidine and compliance with the sterile technique. Conclusions: the educational intervention showed an effect on increasing compliance with catheter-associated infection prevention practices.