Three-dimensional (3D) autotransplantation is a technique for surgical transposition of a tooth to another site within one patient, using 3D printed replicas of the donor tooth. In this study, we evaluated intraoperative experiences during 3D autotransplantation of teeth.A multicenter prospective clinical study was implemented. All procedures were performed using preoperative cone-beam computed tomography imaging and computer-assisted design with computer-assisted manufacturing resulting in a 3D replica of the donor tooth.The 100 autotransplantation procedures (79 patients) included canines, premolars, molars, and 1 supernumerary tooth. In 82% of the procedures, the transplantation was performed with an extra-alveolar time of less than 1 minute and an immediate good fit of the donor tooth. In 14%, the extra-alveolar time was 1 to 3 minutes or multiple fitting attempts were necessary. In 4%, the extra-alveolar time exceeded 3 minutes. Difficulties during the procedures were caused by movement artifacts on the preoperative cone-beam computed tomography imaging, a long interval between the imaging and the procedure, or insufficient bone volume at the recipient site.The use of a 3D printed donor tooth replica during autotransplantation procedures minimized the extra-alveolar time and intraoperative fitting attempts of transplants. This facilitated a quick and reliable treatment and enabled more difficult procedures.