This study aimed to investigate the reliability of 1) the key parameters of cycling ergospirometry (maximum power output [Pmax] and oxygen uptake [V̇O2peak], ventilatory thresholds 1 [VT 1] and 2 [VT 2], and cycling efficiency [CE] and gross efficiency [GE]), 2) the commonly used parameters to quantify exhaustion (maximum heart rate [HFmax], respiratory quotient [RQmax], blood lactate concentration [BLAmax], and ratings of perceived exhaustion [RPEmax]), and 3) the kinetics of exercise induced gas exchange measurements (oxygen uptake [V̇O2], carbon dioxide output [V̇CO2], and minute ventilation [V̇E]) using the PowerCube-Ergo metabolic system in consideration of international statistical recommendations. 12 women and 12 men (28 ± 4 years; 23.2 ± 2.4 kg/m(2)) performed two cycling tests (20 watt/min) separated by one week. The reliability was calculated based on differences in means (t test and effect sizes), retest correlation (intraclass correlation coefficient [ICC]), and within-subject variation (standard error of measurement [SEM]). Of the key parameters of cycling ergospirometry, an excellent reliability (ICC ≥ 0.969; p = 0.000) and high accuracy (%SEM ≤ 4.6) were found for Pmax, V̇O2peak, and VT 1. Of the most commonly used parameters to quantify exhaustion, an excellent reliability (ICC = 0.922; p = 0.000) and high accuracy (%SEM = 1.0) existed only for HFmax. The gas exchange measurements (V̇O2, V̇CO2 und V̇E) of the PowerCube-Ergo were all excellently reliable (ICC ≥ 0,991; p = 0.000) and the accuracy of V̇O2 (SEM = 0.10 l/min) and V̇E (SEM = 3.13 l/min) fulfilled the quality guidance of exercise physiology laboratories. For future studies and practical purposes, the results are vital for the decision as to whether a difference between two tests represents a true intervention effect or just a measurement error and for the estimation of required sample sizes.