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Until now, the use of computed tomography (CT) in the diagnosis and evaluation of otosclerosis has been based on correlation of radiologic findings to patient histories, intraoperative examinations, and audiologic data. The purpose of this study was to compare CT findings in otosclerosis to histopathology.Prospective blinded.Radiology department in a tertiary referral hospital and otopathology laboratory.Temporal bones from patients with otosclerosis and other otologic diseases (used as controls).Blinded review of specimen CT scans by radiologists and comparison of CT findings to histopathology of the same bones.Ability of CT to diagnose otosclerosis, identify otosclerotic foci in defined zones of the otic capsule, determine endosteal layer involvement, oval window (OW) obliteration, and round window (RW) obliteration.In a randomized blinded evaluation, radiologists identified 8 of 10 bones with otosclerosis and made 3 false-positive diagnoses from the 36 control bones. Radiologic examination correctly identified otosclerosis anterior to the oval window, in the pericochlear area, and in the round window niche in 17 of 17, 9 of 11, and 3 of 6 bones, respectively. CT correctly determined involvement of the endosteal layer, OW obliteration, and RW obliteration in 5 of 8, 2 of 2, and 2 of 2 temporal bones.High-resolution CT is highly sensitive and specific for the diagnosis of otosclerosis when compared with histopathology. Very small and subtle otosclerotic foci seen on pathology may be missed on CT. Although CT was able to positively identify cochlear endosteal margin involvement, the false-negative rate on CT was significant.