OBJECTIVES Using both in vitro and in vivo techniques, we investigated the extent to which cardiac motion alters Doppler-measured blood flow velocity and thus potentially can alter the calculation of valve areas or pressure gradients. BACKGROUND Blood flow velocity measured by Doppler ultrasound represents the net motion of the blood relative to the transducer. It is widely assumed that the measured velocity represents the actual flow. It has been demonstrated that cardiac motion generates regularly occurring low velocity Doppler signals that are commonly treated as artifact. METHODS We used an in vitro model that allowed us to measure and independently control the flow of a liquid through a chamber and the motion of the chamber relative to the Doppler beam. A cornstarch-water slurry was driven by a pulsatile pump through tubing to simulate the blood flow within the heart, and the tubing was cyclically moved by a piston to simulate the heart motion. We also measured cardiac motion using M-mode and two-dimensional echocardiography and compared the results with the Doppler signal derived from cardiac motion in subjects without cardiac disease. RESULTS In the in vitro model, alteration in the motion of the tubing resulted in apparent changes in the measured maximal velocity of the fluid. The Doppler spectrum of the combined motion of the tubing and the fluid was the algebraic sum of their Doppler signals. In human subjects, the maximal slope of the M-mode tracing of the aortic annular motion and the peak Doppler signal due to cardiac motion were compared and were highly correlated. CONCLUSIONS Cardiac motion alters the Doppler signal derived from blood flow. This effect can be demonstrated in vitro and in vivo.