The sensitivity of NASA-TLX scale, modified Cooper-Harper (MCH) scale and the mean inter-beat interval (IBI) of successive heart beats, as measures of pilot mental workload (MWL), were evaluated in a flight training device (FTD). Operational F/A-18C pilots flew instrument approaches with varying task loads. Pilots’ performance, subjective MWL ratings and IBI were measured. Based on the pilots’ performance, three performance categories were formed; high-, medium- and low-performance. Values of the subjective rating scales and IBI were compared between categories. It was found that all measures were able to differentiate most task conditions and there was a strong, positive correlation between NASA-TLX and MCH scale. An explicit link between IBI, NASA-TLX, MCH and performance was demonstrated. While NASA-TLX, MCH and IBI have all been previously used to measure MWL, this study is the first one to investigate their association in a modern FTD, using a realistic flying mission and operational pilots.