Vigilance degradation usually causes significant performance decrement. It is also considered the major factor causing the out-of-the-loop phenomenon (OOTL) occurrence. OOTL is strongly related to a high level of automation in operative contexts such as the Air Traffic Management (ATM), and it could lead to a negative impact on the Air Traffic Controllers’ (ATCOs) engagement. As a consequence, being able to monitor the ATCOs’ vigilance would be very important to prevent risky situations. In this context, the present study aimed to characterise and assess the vigilance level by using electroencephalographic (EEG) measures. The first study, involving 13 participants in laboratory settings allowed to find out the neurophysiological features mostly related to vigilance decrements. Those results were also confirmed under realistic ATM settings recruiting 10 professional ATCOs. The results demonstrated that (i) there was a significant performance decrement related to vigilance reduction; (ii) there were no substantial differences between the identified neurophysiological features in controlled and ecological settings, and the EEG-channel configuration defined in laboratory was able to discriminate and classify vigilance changes in ATCOs’ vigilance with high accuracy (up to 84%); (iii) the derived two EEG-channel configuration was able to assess vigilance variations reporting only slight accuracy reduction.