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Tissue Doppler is used to evaluate mitral annulus (MA) dynamics. This technique has two major limitations: it is two-dimensional and angle-dependent. To solve these limitations, our aim was to develop and test a tool for automated 3D tracking and quantification of the MA motion throughout the cardiac cycle by using real-time 3D echocardiography (RT3DE). The tracking algorithm was developed using a combination of two optical flow techniques in the 3D space. Transthoracic RT3DE imaging was performed in 45 subjects, including 15 normal subjects (NL) and 30 patients with mitral regurgitation (MR) due to dilated (n=15, DCM- MR) or ischemic (n=15, ISC-MR) cardiomyopathy. Our technique allowed the evaluation of MA geometry and the quantification of MA function parameters from RT3DE images, which were able to discriminate between NL, DCM-MR and ISC-MR subjects.