PURPOSE: To investigate the clinical importance and feasibility of a 3-D portal image analysis method in comparison with a standard 2-D portal image analysis method for pelvic irradiation techniques. METHODS AND MATERIALS: In this study, images of 30 patients who were treated for prostate cancer were used. A total of 837 imaged fields were analyzed by a single technologist, using automatic 2-D and 3-D techniques independently. Standard deviations (SDs) of the random, systematic, and overall variations, and the overall mean were calculated for the resulting data sets (2-D and 3-D), in the three principal directions (left-right [L-R], cranial-caudal [C-C], anterior-posterior [A-P]). The 3-D analysis included rotations as well. For the translational differences between the three data sets, the overall SD and overall mean were computed. The influence of out-of-plane rotations on the 2-D registration accuracy was determined by analyzing the difference between the 2-D and 3-D translation data as function of rotations. To assess the reliability of the 2-D and 3-D methods, the number of times the automatic match was manually adjusted was counted. Finally, an estimate of the workload was made. RESULTS: The SDs of the random and systematic components of the rotations around the three orthogonal axes were 1. 1 (L-R), 0.6 (C-C), 0.5 (A-P) and 0.9 (L-R), 0.6 (C-C), 0.8 (A-P) degrees, respectively. The overall mean rotation around the L-R axis was 0.7 degrees, which deviated significantly from zero. Translational setup errors were comparable for 2-D and 3-D analysis (ranging from 1.4 to 2.2 mm SD and from 1.5 to 2.5 mm SD, respectively). The variation of the difference between the 2-D and 3-D translation data increased from 1.1 mm (SD) for zero rotations to 2.7 mm (SD) for out-of-plane rotations of 3 degrees, due to a reduced 2-D registration accuracy for large rotations. The number of times the analysis was not considered acceptable and was manually adjusted was 44% for the 2-D analysis, and 6% for the 3-D analysis. CONCLUSION: True 3-D analysis of setup errors for a group of 30 patients with prostate cancer demonstrated that setup rotations are rather small. The deformation of the projected anatomy in portal images caused by out-of-plane rotations leads to a reduced 2-D registration accuracy. For rotations larger than 3 degrees this effect can be quite pronounced, making 3-D registration the preferred method. Furthermore, the automatic 3-D registration has a higher success rate, most likely because this technique uses more information compared to the 2-D method.