Planar gated blood-pool imaging (GBPI) is a standard method for non-invasive assessment of left ventricular (LV) function. Gated blood-pool single photon emission computed tomographic (GBPS) data acquisition can be accomplished in the same time as GBPI, with the benefit of enabling visualization of all cardiac chambers simultaneously. The purpose of this investigation was to evaluate the degree to which automated and manual LVEF calculations agree with one another and with conventional GBPI LVEF measurements. GBPI studies were performed in 22 consecutive, unselected patients, followed by GBPS data acquisition. GBPS left ventricular ejection fraction (LVEF) calculations were performed by available software (NuSMUGA, Northwestern University, Chicago, IL) automatically and manually, using all LV gated short axis slices. Automatic LVEF assessed by GBPS correlated well with conventional planar GBPI (r = 0.88, P < 0.001). Mean planar GBPI LVEF was 50% +/- 12%, and mean GBPS automatic LVEF was significantly lower at 45% + 14% (P = 0.001), with a mean difference of 6% +/- 5%. Manual GBPS LVEF also correlated well with conventional planar GBPI (r = 0.90, P < 0.0001). Mean LVEF measurement by manual GBPS versus GBPI was significantly higher at 59% +/- 13%, with a mean difference of 10% +/- 6% (P < 0.001). Manual GBPS LVEF values were also significantly higher than automatically determined GBPS LVEF values (P < 0.001). It is concluded that LVEF values assessed by NuSMUGA GBPS software were reproducible, and automatic and manual values correlated well with conventional GBPI values. However, both automatic and manual GBPS calculations were significantly different from one another and from GBPI values, so that GBPI and NuSMUGA calculations cannot be considered to be equivalent.