Purpose: To determine the intra- and inter-day reliability of a 2x4-minute performance test on a cycle ergometer (Wattbike) separated by 30-minutes of passive recovery (2x4MMP). Methods: Twelve highly-trained cyclists (mean ᠓D; age = 20 ᠲ years; predicted VO2max = 59.0 ᠳ.6 mL竧-1筩n-1) completed six 2x4MMP cycling tests on a Wattbike ergometer separated by seven days. Mean power was measured to determine intra-day (T1 to T2) and inter-day reliability (week 1-6) over the repeated trials. Results: The mean intra-day reliability (T1 to T2) of the 2x4MMP test, as expressed by the typical error of measurement (TEM; watts) and coefficient of variation (CV; %) over the six weeks was 10.0 watts (95% confidence limits, CL; 8.2-11.8), and 2.6% (95%CL 2.1-3.1), respectively. The mean inter-day reliability TEM and CV for T1 over the six weeks was 10.4 watts (95%CL 8.7-13.3) and 2.7% (95%CL 2.3-3.5), respectively, and 11.7 watts (95%CL 9.8-15.1) and 3.0% (95%CL 2.5-3.9) for T2. Conclusion: The testing protocol performed on a Wattbike cycle ergometer in the current study is reproducible in highly-trained cyclists. The high intra- and inter-day reliability makes it a reliable method for monitoring cycling performance and for investigating factors that affect performance in cycling events.