The 3 min step test is a widely used method to evaluate physical fitness, but whether this method is valid when performed at altitude is unknown. The purpose of this study was to examine the effect of altitude on the fitness score of the 3 min step test, and the role of ambient temperature in this effect. In study I, 11 healthy volunteers (aged 18.1 +/- 1.1 years) performed a 3 min step test at sea level and at altitude (1950 m). Plasma lactate and stress hormones, as indicators of metabolic stress, were measured before and after the test. To determine the role of ambient temperature, we performed study II at sea level with the same step test simulating the altitude temperature condition (24 degrees C at high altitude versus 32 degrees C at sea level) with 23 subjects (aged 20.4 +/- 0.4 y). In study I, plasma lactate of the subjects was elevated during the step test at sea level and to a greater extent at high altitude. Plasma cortisol and testosterone levels were elevated only at high altitude. However, the heart rate (HR) recovery after the step test was faster at high altitude than at sea level, producing a better physical fitness index. Furthermore, in study II, we demonstrated that the subjects who performed the 3 min step test at 24 degrees C exhibited faster HR recovery than at 32 degrees C. The current study therefore suggests that environmental conditions leading to temperature variation have strong confounding effects on the fitness score of the 3 min step test.