This paper describes research that aims to provide the overall scientific basis for implementation of a Fatigue Risk Management System (FRMS) for French regional airlines. The current research has evaluated the use of different tools and indicators that would be relevant candidates for integration into the FRMS. For the Fatigue Risk Management component, results show that biomathematical models of fatigue are useful tools to help an airline to prevent fatigue related to roster design and for the management of aircrew planning. The Fatigue Safety assurance includes two monitoring processes that have been evaluated during this research: systematic monitoring and focused monitoring. Systematic monitoring consists of the analysis of existing safety indicators such as Air Safety Reports (ASR) and Flight Data Monitoring (FDM). Results show a significant relationship between the hours of work and the frequency of ASR. Results for the FDM analysis show that some events are significantly related to the fatigue risk associated with the hours of works. Focused monitoring includes a website survey and specific in-flight observations and data collection. Sleep and fatigue measurements have been collected from 115 aircrews over 12-day periods (including rest periods). Before morning duties, results show a significant sleep reduction of up to 40% of the aircrews’ usual sleep needs leading to a clear increase of fatigue during flights. From these results, specific guidelines are developed to help the airlines to implement the FRMS and for the airworthiness to oversight the implementation of the FRMS process.