OBJECTIVE The purpose of this study was to quantify the reproducibility, temporal stability, and functional correlation of diffusion MR characteristics in the spinal cord in patients with cervical stenosis with or without myelopathy. The association between longitudinal diffusion tensor imaging (DTI) measurements and serial neurological function assessment was explored at both the group and individual level. METHODS Sixty-six nonoperatively treated patients with cervical stenosis were prospectively followed (3 months to > 5 years) using synchronous serial MRI and functional outcome assessment. A total of 183 separate MRI examinations were performed, separated by at least 3 months, and each patient had a minimum of 2 MRI scans (range 2-5 scans). Anatomical and DTI measurements were performed within the spinal cord at the C1-2 region as well as at the area of highest compression. Coefficients of variance (COVs) were compared across measurements in both reference tissue and areas of compression for anatomical measurements, fractional anisotropy (FA), and mean diffusivity (MD). The correlation between diffusion MR measures at the site of compression and evaluations of neurological function assessed using the modified Japanese Orthopaedic Association (mJOA) scale at multiple time points was evaluated. RESULTS The COVs for anatomical measurements (Torg ratio and canal diameter) were between 7% and 10%. The median COV for FA measurements at the site of compression was 9%, and for reference tissue at C1-2 it was 6%. The median COV for MD at the site of compression was approximately 12%, and for reference tissue at C1-2 it was 10%. The FA and MD measurements of C1-2 averaged 0.61 and 0.91 μm/msec, respectively, whereas the FA and MD measurements at the site of compression averaged 0.51 and 1.26 μm/msec, respectively. Both FA (slope = 0.037; R = 0.3281, p < 0.0001) and MD (slope = -0.074; R = 0.1101, p = 0.0084) were significantly correlated with the mJOA score. The FA decreased by approximately 0.032 units per mJOA unit decrease (R = 0.2037, p < 0.0001), whereas the MD was increased by approximately 0.084 μm/msec for every mJOA unit decrease (R = 0.1016, p < 0.0001). CONCLUSIONS Quantitative DTI measurements of the spinal cord in patients with cervical stenosis with or without myelopathy have a median COV of 5%-10%, similar to anatomical measurements. The reproducibility of these measurements and significant correlation with functional outcome status suggest a potential role in the evaluation and longitudinal surveillance of nonoperatively treated patients. With respect to the specific DTI measurements, FA within the spinal cord appears slightly more sensitive to neurological function and more stable than measures of MD. Therefore, DTI of the spinal cord may be a clinically feasible imaging technique for longitudinally monitoring patients with cervical spondylotic myelopathy.