It is stated that cardiac MRI imaging can provide accurate estimation of left ventricular (LV) volumes and ejection fraction (EF). The purpose of this study was to evaluate the accuracy of gated myocardial perfusion SPECT for assessment of LV end-diastolic volume (EDV), end-systolic volume (ESV) and EF, using cardiac MRI as the reference methods/(methodology). Gated myocardial perfusion SPECT images were analyzed with two different quantification software, QGS and 4D-MSPECT. Thirty-four consecutive patients were studied. Myocardial perfusion SPECT and cardiac MRI had excellent intra/interobserver reproducibility. Correlation between the results of gated myocardial perfusion SPECT and cardiac MRI were high for EDV and EF. However, ESV and EDV were significantly underestimated by gated myocardial perfusion SPECT compared to cardiac MRI. Moreover, gated myocardial perfusion SPECT overestimated EF for small heart. One reason for the difference in volumes and EF is the delineation of the endocardial border. Cardiac MRI has higher spatial resolution. We should understand the differences of volumes and EF as determined by gated myocardial perfusion SPECT and cardiac MRI.