Background-Accurate measures are critical when attempting to distinguish normal from pathological changes in cardiac function during exercise, yet imaging modalities have seldom been assessed against invasive exercise standards. We sought to validate a novel method of biventricular volume quantification by cardiac MRI (CMR) during maximal exercise. Methods and Results-CMR was performed on 34 subjects during exercise and free-breathing with the use of an ungated real-time (RT-ungated) CMR sequence. ECG and respiratory movements were retrospectively synchronized, enabling compensation for cardiac cycle and respiratory phase. Feasibility of RT-ungated imaging was compared with standard exercise CMR imaging with ECG gating (gated); accuracy of RT-ungated CMR was assessed against an invasive standard (direct Fick); and reproducibility was determined after a second bout of maximal exercise. Ventricular volumes were analyzed more frequently during high-intensity exercise with RT-ungated compared with gated CMR (100% versus 47%; P<0.0001) and with better interobserver variability for RT-ungated (coefficient of variation=1.9% and 2.0% for left and right ventricular stroke volumes, respectively) than gated (coefficient of variation=15.2% and 13.6%; P<0.01). Cardiac output determined by RT-ungated CMR proved accurate against the direct Fick method with excellent agreement (intraclass correlation coefficient, R=0.96), which was highly reproducible during a second bout of maximal exercise (R=0.98). Conclusions-When RT-ungated CMR is combined with post hoc analysis incorporating compensation for respiratory motion, highly reproducible and accurate biventricular volumes can be measured during maximal exercise. (Circ Cardiovasc Imaging. 2013;6:329-338.)