Tomographic multi-gated blood pool scintigraphy (TMUGA) is a widely available method which permits simultaneous assessment of right and left ventricular ejection fractions. However, the widespread clinical use of this technique is impeded by the lack of segmentation methods dedicated to an automatic analysis of ventricular activities. In this study we evaluated how a watershed algorithm succeeds in providing semi-automatic segmentation of ventricular activities in order to measure right and left ejection fractions by TMUGA. The left ejection fractions of 30 patients were evaluated both with TMUGA and with planar multi-gated blood pool scintigraphy (PMUGA). Likewise, the right ejection fractions of 25 patients were evaluated with first-pass scintigraphy (FP) and with TMUGA. The watershed algorithm was applied to the reconstructed slices in order to group together the voxels whose activity came from one specific cardiac cavity. First, the results of the watershed algorithm were compared with manual drawing around left and right ventricles. Left ejection fractions evaluated by TMUGA with the watershed procedure were not significantly different (p=0. 30) from manual outlines whereas a small but significant difference was found for right ejection fractions (p=0.004). Then right and left ejection fractions evaluated by TMUGA (with the semi-automatic segmentation procedure) were compared with the results obtained by FP or PMUGA. Left ventricular ejection fractions evaluated by TMUGA showed an excellent correlation with those evaluated by PMUGA (r=0. 93; SEE=5.93%; slope=0.99; intercept = 4.17%). The measurements of these ejection fractions were significantly higher with TMUGA than with PMUGA (P<0.01). The interoperator variability for the measurement of left ejection fractions by TMUGA was 4.6%. Right ventricular ejection fractions evaluated by TMUGA showed a good correlation with those evaluated by FP (r = 0.81; SEE = 6.68%; slope = 1.00; intercept = 0.85%) and were not significantly different (P = 0.42). The interoperator variability with TMUGA was 6.7% for the right ventricle. Thus, the watershed algorithm proposed is an efficient segmentation tool for the semi-automatic analysis of right and left ventricular ejection fractions by TMUGA. Further studies are necessary to check whether this procedure can be used to evaluate ventricular volumes and cardiac outflow.