Stress is a word used to describe human reactions to emotionally, cognitively and physically challenging experiences. A hallmark of the stress response is the activation of the autonomic nervous system, resulting in the "fight-freeze-flight" response to a threat from a dangerous situation. Consequently, the capability to objectively assess and track a controller's stress level while dealing with air traffic control (ATC) activities would make it possible to better tailor the work shift and maintain high safety levels, as well as to preserve the operator's health. In this regard, sixteen controllers were asked to perform a realistic air traffic management (ATM) simulation during which subjective data (i.e. stress perception) and neurophysiological data (i.e. brain activity, heart rate, and galvanic skin response) were collected with the aim of accurately characterising the controller's stress level experienced in the various experimental conditions. In addition, external supervisors regularly evaluated the controllers in terms of manifested stress, safety, and efficiency throughout the ATM scenario. The results demonstrated 1) how the stressful events caused both supervisors and controllers to underestimate the experienced stress level, 2) the advantage of taking into account both cognitive and hormonal processes in order to define a reliable stress index, and 3) the importance of the points in time at which stress is measured owing to the potential transient effect once the stressful events have ceased. Stress in air traffic management. The air traffic management (ATM) system is currently going through a major restructuring process, covering the entire architecture of process, personnel, and management environments. The increase in the level of automation as support tools for air traffic control (ATC) activities is one of the main technological challenges which will be faced in the future 1,2. Automation, digitalisation of information and virtualisation of the ATC operations will require a transformation of tasks and working methods. For example, with the prospect of an increasing in the volume of air traffic to be managed, controllers will be expected to delegate more activities and decisions to automation. In ATC, high demand is usually associated with a high mental workload and level of responsibility, while stress is linked to available time, team support, adherence to procedures and control over the planning and execution of tasks. The concepts of mental workload and stress are often used ambiguously, especially because of their relationship with performance when they reach very low or high values. In such cases, performance drops dramatically. However, certain levels of mental workload and