Given the importance of ensuring athletes train and compete in a non-fatigued state, reliable tests are required in order to regularly monitor fatigue. The purpose of this study was to investigate the reliability of a cycle ergometer to measure peak power during short maximal sprint cycle efforts in running-based team sport athletes. Fourteen professional male Australian rules footballers performed a sprint cycle protocol during three separate trials, with each trial separated by seven days. The protocol consisted of a standardized warm-up, a maximal 6 s sprint cycle effort, a 1-minute active recovery, and a second maximal 6 s sprint cycle effort. Peak power was recorded as the highest power output of the two sprint cycle efforts. Absolute peak power (mean ± SD) was 1502 ± 202 W, 1498 ± 191 W, and 1495 ± 210 W for trials 1, 2, and 3, respectively. The mean coefficient of variation, intraclass correlation coefficient, and standard error of measurement for peak power between trials was 3.0% (90% Confidence Intervals = 2.5-3.8%), 0.96 (90% Confidence Intervals = 0.91-0.98), and 39 W, respectively. The smallest worthwhile change for relative peak power was 6.0%, which equated to 1.03 W·kg. The cycle ergometer sprint test protocol described in this study is highly reliable in elite Australian rules footballers and can be used to track meaningful changes in performance over time, making it a potentially useful fatigue-monitoring tool.