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Study design A randomized controlled trial. Objective To compare the radiation exposure with the scrub nurse, assistant surgeon, and anesthetist during minimally invasive transforaminal lumbar interbody fusion using conventional 2-dimensional (2D) fluoroscopy or 3D fluoroscopy-based navigation. Summary of background data Minimally invasive spinal fusion techniques are related to higher radiation exposures compared with open techniques. Especially the routinely exposed surgical staff faces the risks of increased radiation exposure. Methods In total, 41 patients with planned monosegmental minimally invasive transforaminal lumbar interbody fusion were randomized into the intraoperative imaging techniques 2D fluoroscopy or 3D navigation. Eye lens and film dosemeters were attached to defined locations of the scrub nurse, assistant surgeon, and anesthetist. Mann-Whitney U and Wilcoxon-matched pairs signed-rank test were used to compare dosemeter readings. This study was registered with the German Clinical Trials Register (DRKS00004514). Results The radiation exposure per surgery was low for the scrub nurse, assistant surgeon, and anesthetist in both the 2D fluoroscopy and 3D navigation groups. The maximum average value of 0.057±0.031 mSv was measured on the unprotected chest of the assistant surgeon and was thus slightly above the lower detection limit of the dosemeters (0.044 mSv). The annual occupational dose limit would be exceeded at the earliest after 571 operations for the unprotected eye lens of the assistant surgeon. Conclusions Minimally invasive lumbar fusion surgery is possible with comparatively low radiation exposure to the assisting operating room personnel without exceeding the annual maximum occupational radiation exposure. However, there is no definite dose value below which ionizing radiation poses no risk. Consequently, radiation sparing work routines should be strictly followed.