BACKGROUND No evidence-based recommendation exists regarding how far clinicians should insert a rectal thermistor to obtain the most valid estimate of core temperature. Knowing the validity of temperatures at different rectal depths has implications for exertional heat-stroke (EHS) management. OBJECTIVE To determine whether rectal temperature (Trec) taken at 4 cm, 10 cm, or 15 cm from the anal sphincter provides the most valid estimate of core temperature (as determined by esophageal temperature [Teso]) during similar stressors an athlete with EHS may experience. METHODS Cross-sectional study. METHODS Laboratory. METHODS Seventeen individuals (14 men, 3 women: age = 23 ± 2 years, mass = 79.7 ± 12.4 kg, height = 177.8 ± 9.8 cm, body fat = 9.4% ± 4.1%, body surface area = 1.97 ± 0.19 m2). METHODS Rectal temperatures taken at 4 cm, 10 cm, and 15 cm from the anal sphincter were compared with Teso during a 10-minute rest period; exercise until the participant's Teso reached 39.5°C; cold-water immersion (∼10°C) until all temperatures were ≤38°C; and a 30-minute postimmersion recovery period. The Teso and Trec were compared every minute during rest and recovery. Because exercise and cooling times varied, we compared temperatures at 10% intervals of total exercise and cooling durations for these periods. METHODS The Teso and Trec were used to calculate bias (ie, the difference in temperatures between sites). RESULTS Rectal depth affected bias (F2,24 = 6.8, P = .008). Bias at 4 cm (0.85°C ± 0.78°C) was higher than at 15 cm (0.65°C ± 0.68°C, P < .05) but not higher than at 10 cm (0.75°C ± 0.76°C, P > .05). Bias varied over time (F2,34 = 79.5, P < .001). Bias during rest (0.42°C ± 0.27°C), exercise (0.23°C ± 0.53°C), and recovery (0.65°C ± 0.35°C) was less than during cooling (1.72°C ± 0.65°C, P < .05). Bias during exercise was less than during postimmersion recovery (0.65°C ± 0.35°C, P < .05). CONCLUSIONS When EHS is suspected, clinicians should insert the flexible rectal thermistor to 15 cm (6 in) because it is the most valid depth. The low level of bias during exercise suggests Trec is valid for diagnosing hyperthermia. Rectal temperature is a better indicator of pelvic organ temperature during cold-water immersion than is Teso.