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PurposeTo identify long-term reproducible texture features (TFs) of spinal computed tomography (CT), and characterize variations with regard to gender, age and vertebral level using our automated quantification framework.MethodsWe performed texture analysis (TA) on baseline and follow-up CT (follow-up duration: 30–90 days) of 21 subjects (8 females, 13 males, age at baseline 61.2 ± 9.2 years) to determine long-term reproducibility. TFs with a long-term reproducibility error Δrelglobal measures the spread of the gray-level distribution in an image while Entropy reflects the uniformity of gray-levels. Short-run emphasis (SRE), Long-run emphasis (LRE), Run-length non-uniformity (RLN) and Run percentage (RP) contain information on consecutive voxels of a particular grey-level, or grey-level range, in a particular direction. Long runs (LRE) represent coarse texture while short runs (SRE) represent fine texture. RLN reflects similarities in the length of runs while RP reflects distribution and homogeneity of runs with a specific direction.ResultsSix of the 24 extracted TFs had Δrelglobal, Entropy, SRE, LRE, RLN, RP), and were analyzed further in 4716 thoracolumbar vertebrae. Five TFs (Varianceglobal,SRE,LRE, RLN,RP) showed a significant difference between genders (pglobal and Entropy showed a significant increase from the thoracic to the lumbar spine (pConclusionLong-term reproducible CT-based TFs of the thoracolumbar spine were established and characterized in a predominantly older adult study population. The gender-, age- and vertebral-level-specific values may serve as foundation for osteoporosis diagnostics and facilitate future studies investigating vertebral microstructure.