The purpose of the study was to investigate which systematic effects of prosthetic misalignment could be observed with the use of the SYBAR system. The alignment of the prosthesis of five well-trained unilateral trans-tibial amputees was changed 15° in magnitude in varus, valgus, flexion, extension, endorotation, exorotation, dorsal flexion, and plantar flexion. Subjects walked over a distance of 8 m at a self-selected walking speed with the alignment of the prosthesis as it was at the start of the experiment (reference) and with each changed alignment. Two video cameras (frontal and sagittal) and a force plate of the SYBAR system (Noldus Information Technology, The Netherlands) were used to capture gait characteristics of the subjects. Temporal and spatial characteristics, the magnitude and timing of the ground reaction force (GRF), and the external joint moments were derived from these data. Despite the substantial perturbations to prosthetic alignment, only a few effects were observed in the temporal and spatial characteristics of gait, the magnitude and timing of the GRF, and the external joint moments. Only the pattern of the ground reaction force in the mediolateral direction and the joint moment around the ankle in the frontal plane during terminal stance showed a systematic effect when the alignment was set into varus and valgus or exorotation. It was concluded that using the SYBAR system in this study revealed little effect of perturbations in prosthetic alignment, for this group of patients, and for the selected parameters. It was questioned whether this is due to the relatively low resolution of the SYBAR system or the capacity of the well-trained subjects to compensate for the disturbance in alignment. It was suggested that the usability of the SYBAR system in clinical settings should be further explored. © 2005 ISPO.