We describe a novel software system that utilizes automated algorithms to perform edge detection and wall tracking of high-resolution B-mode arterial ultrasound images, combined with synchronized Doppler waveform envelope analysis, to calculate conduit arterial blood flow (BF) across the cardiac cycle. Furthermore, we describe changes in brachial arterial BF to the resting forearm during incremental cycle ergometry in eight subjects. During exercise, peak BF during the cardiac cycle increased at each workload (P < 0.001), because of increased velocity in the presence of unaltered cross-sectional area. In contrast, mean BF calculated across each cardiac cycle decreased at lower workloads before increasing at 100 and 160 W (P < 0.001). Differences in the pattern of peak and mean cardiac cycle flows were due to the influence of retrograde diastolic flow, which had a larger impact on mean flows at lower workloads. In conclusion, BF can be measured with high temporal resolution across the cardiac cycle in humans. Resting brachial arterial flow, including retrograde flow, increases during lower limb exercise.