The assessment of asymmetry has always represented one of the most relevant field of research in anatomy, with several applications in reconstructive and aesthetic surgery. This study aims at exposing an innovative protocol for the assessment of asymmetry of cranial structures based on 3D-3D superimposition and mirroring procedures. Five patients who underwent a head CT scan were selected. Three anatomical structures were automatically segmented through ITK-SNAP free software: maxillary sinuses, zygomatic bones and the petrous portion of both temporal bones. The left structure was then reflected automatically into the right image and superimposed over the contralateral one according to the least point-to-point distance on the entire surface. RMS (root mean square) distance was then automatically calculated for each superimposition. In addition, a chromatic map of superimposition was generated, showing the most variable areas between the two sides. Possible statistically significant differences in RMS value according to the three anatomical structures were assessed through one-way ANOVA test (p<0.05). On average, RMS values for maxillary sinuses, zygomatic bones and petrous part of temporal bones were respectively 1.00 mm (SD: 0.27 mm), 0.70 mm (SD: 0.34 mm) and 1.26 mm (SD: 0.17 mm). A significant difference among the symmetry of the three analysed structures was found (p<0.01, ANOVA). A novel approach for the assessment of anatomical asymmetry is suggested, based both on a morphological and a quantitative evaluation of differences between the right and left side skull bones.