The Optoelectronic Systems (OS) are largely used in gait analysis to evaluate the motor performances of healthy subjects and patients. The accuracy of marker trajectories reconstruction depends on several aspects: the number of cameras, the dimension and position of the calibration volume, and the chosen calibration procedure. In this paper we propose a methodology to evaluate the effects of the mentioned sources of error on the reconstruction of marker trajectories. The novel contribution of the present work consists in the dimension of the tested calibration volumes, which is comparable with the ones normally used in gait analysis; in addition, to simulate trajectories during clinical gait analysis, we provide non-default paths for markers as inputs. Several calibration procedures are implemented and the same trial is processed with each calibration file, also considering different cameras configurations. The RMSEs between the measured trajectories and the optimal ones are calculated for each comparison. To investigate the significant differences between the computed indices, an ANOVA analysis is implemented. The RMSE is sensible to the variations of the considered calibration volume and the camera configurations and it is always inferior to 43 mm.