OBJECTIVE This study evaluated the use of gated single-photon emission computed tomographic (SPECT) myocardial perfusion images for determination of left ventricular ejection fraction. BACKGROUND Gated SPECT has expanded the applications of myocardial perfusion imaging to include the evaluation of left ventricular size, regional wall motion and regional systolic thickening. Accurate automated or semi-automated methods for quantitation of left ventricular ejection fraction from tomographic perfusion images would provide additional valuable clinical information. METHODS Rest gated SPECT was performed on the stress distribution of technetium-99m sestamibi, using eight frames per cardiac cycle. Mid-horizontal long-axis and vertical long-axis gated tomographic perfusion images were analyzed after digital matrix inversion, which enhances edge detection, for ejection fraction determination. These ejection fractions were compared with those determined by contrast ventriculography (n = 54, including 45 biplane and 9 single plane) and first-pass radionuclide angiography (n = 38) in patients with coronary artery disease. RESULTS Myocardial perfusion SPECT image inversion-derived ejection fractions were slightly lower (2.7 ejection fraction units, p < 0.01), and first-pass ejection fractions were much lower (8.0 ejection fraction units, p < 0.001) than those obtained with contrast ventriculography. There was excellent correlation between SPECT and contrast ventriculographic ejection fractions (r = 0.93) over a wide range of ejection fractions (14% to 89%). Good correlation was also observed between first-pass radionuclide angiography and both contrast ventriculography (r = 0.83) and SPECT (r = 0.87). Reproducibility of SPECT image inversion ejection fractions was excellent (intraobserver r = 0.99, interobserver r = 0.93). CONCLUSIONS Semiautomated ejection fractions can be obtained from gated SPECT technetium-99m sestamibi perfusion images using the image inversion technique. These results are reproducible and correlate well with results of first-pass radionuclide angiography but are closer in value to those obtained with contrast ventriculography.