Objective Explosive ordnance disposal (EOD) often requires technicians to wear multiple protective garments in challenging environmental conditions. The accumulative effect of increased metabolic cost coupled with decreased heat dissipation associated with these garments predisposes technicians to high levels of physiological strain. It has been proposed that a perceptual strain index (PeSI) using subjective ratings of thermal sensation and perceived exertion as surrogate measures of core body temperature and heart rate, may provide an accurate estimation of physiological strain. Therefore, this study aimed to determine if the PeSI could estimate the physiological strain index (PSI) across a range of metabolic workloads and environments while wearing heavy EOD and chemical protective clothing. Methods Eleven healthy males wore an EOD and chemical protective ensemble while walking on a treadmill at 2.5, 4 and 5.5 km·h− 1 at 1% grade in environmental conditions equivalent to wet bulb globe temperature (WBGT) 21, 30 and 37 °C. WBGT conditions were randomly presented and a maximum of three randomised treadmill walking trials were completed in a single testing day. Trials were ceased at a maximum of 60-min or until the attainment of termination criteria. A Pearson's correlation coefficient, mixed linear model, absolute agreement and receiver operating characteristic (ROC) curves were used to determine the relationship between the PeSI and PSI. Results A significant moderate relationship between the PeSI and the PSI was observed [r = 0.77; p < 0.001; mean difference = 0.8 ± 1.1 a.u. (modified 95% limits of agreement − 1.3 to 3.0)]. The ROC curves indicated that the PeSI had a good predictive power when used with two, single-threshold cut-offs to differentiate between low and high levels of physiological strain (area under curve: PSI three cut-off = 0.936 and seven cut-off = 0.841). Conclusions These findings support the use of the PeSI for monitoring physiological strain while wearing EOD and chemical protective clothing. However, future research is needed to confirm the validity of the PeSI for active EOD technicians operating in the field.