Quantitative assessment of regional cardiac function can improve the accuracy of detecting wall motion abnormalities due to heart disease. While recently developed fast cine displacement-encoded with stimulated echoes (DENSE) MRI is a promising modality for the quantification of regional myocardial function, it has not been validated for clinical applications. The purpose of this study, therefore, was to validate the accuracy of fast cine DENSE MRI with numerical simulation and in vivo experiments. A numerical phantom was generated to model physiologically relevant deformation of the heart, and the accuracy of fast cine DENSE was evaluated against the numerical reference. For in vivo validation, 12 controls and 13 heart-disease patients were imaged using both fast cine DENSE and myocardial tagged MRI. Numerical simulation demonstrated that the echo-combination DENSE reconstruction method is relatively insensitive to clinically relevant resonance frequency offsets. The strain measurements by fast cine DENSE and the numerical reference were strongly correlated and in excellent agreement (mean difference = 0.00; 95% limits of agreement were 0.01 and -0.02). The strain measurements by fast cine DENSE and myocardial tagged MRI were strongly correlated (correlation coefficient = 0.92) and in good agreement (mean difference = 0.01; 95% limits of agreement were 0.07 and -0.04).