Purpose: To validate a method called bi-ventricular strain unwrapped phase (BiSUP) for reconstructing three-dimensional plus time (3D+t) biventricular strain maps from phase-unwrapped harmonic phase (HARP) images derived from tagged cardiac magnetic resonance imaging (MRI). Materials and Methods: A set of 30 human subjects were imaged with tagged MRI. In each study, HARP phase was computed and unwrapped in each short-axis and long-axis image. Inconsistencies in unwrapped phase were resolved using branch cuts manually placed with a graphical user interface. The 3D strain maps were computed independently in each imaged time frame through systole and mid diastole in each study. The BiSUP strain and displacements were compared with those estimated by a 3D feature-based (FB) technique and a 2D+t HARP technique. Results: The standard deviation of the difference between strains measured by the FB and the BiSUP methods was less than 4% of the average of the strains from the two methods. The correlation between peak minimum principal strain measured using the BiSUP and HARP techniques was over 83%. Conclusion: The BiSUP technique can reconstruct full 3D+t strain maps from tagged MR images through the cardiac cycle in a reasonable amount of time and user interaction compared with other 3D analysis methods. Copyright © 2011 Wiley-Liss, Inc.