OBJECTIVE: Multi-component T2 relaxation allows for assessing the myelin water fraction in nervous tissue, providing a surrogate marker for demyelination. The assessment of the number and distribution of different T2 components for devising exact models of tissue relaxation has been limited by T2 sampling with conventional MR methods. MATERIALS AND METHODS: A T2-prepared UTE sequence was used to assess multicomponent T2 relaxation at 9.4 T of fixed mouse and rat spinal cord samples and of mouse spinal cord in vivo. For in vivo scans, a cryogenically cooled probe allowed for 78-µm resolution in 1-mm slices. Voxel-wise non-negative least square analysis was used to assess the number of myelin water-associated T2 components. RESULTS: More than one myelin water-associated T2 component was detected in only 12 % of analyzed voxels in rat spinal cords and 6 % in mouse spinal cords, both in vivo and in vitro. However, myelin water-associated T2 values of individual voxels varied between 0.1 and 20 ms. While in fixed samples almost no components below 1 ms were identified, in vivo, these contributed 14 % of the T2 spectrum. No significant differences in MWF were observed in mouse spinal cord in vivo versus ex vivo measurements. CONCLUSION: Voxel-wise analysis methods using relaxation models with one myelin water-associated T2 component are appropriate for assessing myelin content of nervous tissue.