PURPOSE: This study was conducted to determine the effect of different ambient conditions on thermo-physiological responses of CBRN ensemble wearers performing work in order to provide information regarding exposure limits and rest schedules. METHODS: Eight healthy males (VO2max >/= 50ml/kg/min) completed two trials of a heat stress test (4.8 km/h, 3% grade) while wearing a CBRN ensemble in neutral (22°C, 50%RH) and hot/humid (35°C, 65% RH) ambient conditions. The study variables included body core (Tre) and skin temperatures (Tsk), heart rate (HR), respiratory parameters (VO2, VE), and microclimate temperature/humidity (Tmicro, RHmicro). Two-way repeated measures ANOVA (Conditions × Time) was carried out for statistical analysis. RESULTS: The study results indicated there was a significant main effect for ambient conditions and time in all thermoregulatory and respiratory variables (p<0.001). A pairwise comparison showed significant final measurement values in Tre (38.16+/-0.29 VS. 39.73+/-0.30 °C), Tsk (36.42+/-0.30 VS. 39.12+/-0.37 °C), HR (152.9+/-18.2 vs. 187.1+/-2.1 beats/min), VO2 (1.59+/-0.2 VS. 2.03+/-0.2 L/min), VE (47.7+/-6.0 VS. 59.3+/-10.1 L/min) between neutral and hot/humid conditions, respectively (p<0.001). Tmicro was also significantly elevated in the hot/humid environment (35.1 VS. 38.6 °C), whereas RHmicro reached >/= 95% at 15min exercise in both conditions, significantly hindering evaporative heat loss to the environment. CONCLUSIONS: Working while wearing vapor resistant CBRN ensemble imposes a significant thermo-physiological stress on a wearer. This study showed ambient conditions also have a significant impact on the level of thermal strain, mainly due to diminished evaporative capacity and elevated Tmicro. Environmental conditions need to be taken into account to determine work exposure limits and rest cycles for individuals wearing a CBRN ensemble.