Purpose <p>The free fibular flap (FFF) is a standard procedure for the oral rehabilitation of segmental bone defects in the mandible caused by diseases such as malignant processes, osteonecrosis, or trauma. Digital guides and computer-assisted surgery (CAS) can improve precision and reduce the time and cost of surgery. This study evaluates how different designs of slot cutting guides, guiding heights, and cutting instruments affect surgical accuracy during mandibular reconstruction.</p> Methods <p>Ninety model operations in a three-part fibular transplant for mandibular reconstruction were conducted according to digital planning with three guide designs (standard, flange, and anatomical slots), three guide heights (1 mm, 2 mm, 3 mm), and two osteotomy instruments (piezoelectric instrument and saw). The cut segments were digitized using computed tomography and digitally evaluated to assess surgical accuracy.</p> Results <p>For vestibular and lingual segment length, the anatomical slot and the flange appear to be the most accurate, with the flange slightly under-contoured vestibularly and the standard slot over-contoured lingually and vestibularly (<i>p</i> &lt; 0.001). There were only minor differences between the use of saw and piezoelectric instrument for lingual (<i>p</i> = 0.005) and vestibular (<i>p</i> &lt; 0.001) length and proximal angle (<i>p</i> = 0.014). The U-distance after global reconstruction for flanges resulted in a median deviation of 0.0468 mm (IQR 8.15), but was not significant (<i>p</i> = 0.067).</p> Conclusion <p>Anatomical slots and flanges are recommended for osteotomy, with guiding effects relying on both haptic and visual control. Unilateral guided flanges also work accurately at high guidance heights. The results of piezoelectric instrument (PI) and saw showed comparable results in the assessment of individual segments and U-reconstruction in this in vitro study without soft tissue, so that the final decision is left to the expertise of the surgeons.</p>