Background: Stapedotomy is the most efficient treatment for otosclerosis. The anatomical structure of the operation area is complex, but it has a great impact on the postoperative effect. We measure the anatomical parameters of the stapes and its surrounding structures to provide an anatomical reference for stapes surgery in otosclerosis. Materials and methods: Fifteen adult cadaver heads (30 samples) were scanned using micro-CT. The stapes, facial nerve and external auditory canal were reconstructed by image processing. The stapes parameters and relationships between the stapes and surrounding structures were measured using a three-dimensional reconstruction model. Results: The length, width and thickness of the stapes footplate were 2.93 ± 0.17 mm, 1.46 ± 0.08 mm and 0.30 ± 0.11 mm, respectively. The distance between the stapes footplate and long process of the incus was 3.79±0.39 mm. The angle of the incudostapedial joint was 88.29 ± 11.58°. The distance from the center of the stapes footplate to the facial canal was 1.60 ± 0.34 mm. In simulated stapes surgery, the minimum depth of the external auditory canal to be removed was 2.17 ± 0.91 mm, and no significant difference was found between the left and right sides and between men and women (P > 0.05). Conclusions: A three-dimensional model of the stapes bone and its surrounding anatomical structures was established based on Micro-CT imaging. Anatomical parameters of the stapes bone and its surrounding structures were measured using the model. In stapedotomy, the implanted piston diameter should be around 0.6mm, with a length of approximately 4.6mm. Care should be taken to protect the facial nerve canal during the surgery. These data provide reference for otologists.