To enable an accurate assessment of disease-related abnormalities in the SC of marmosets with EAE.Five SCs were collected following perfusion fixation for healthy (n=1) and EAE animals (n=4) immunized with 200 mg fresh-frozen human white matter homogenate. Both upper and lower spine tissues were stored in 4% formaldehyde (Figure 1A). Each SC sample was inserted inside a 50 ml Falcon tube filled with Fomblin and imaged on a 7T animal scanner (30 cm USR/AVIII Bruker Biospin) using a 35 mm inner diameter (ID) volume coil (Bruker Biospin). The imaging protocol included a 3D T2*-weighted gradient-echo (TR/TE: 80/15 ms, FA: 20°) using 100 μm isotropic voxel size. To obtain high-quality images of the entire spine, the total acquisition time was ~30 h. After this first imaging session, the SC tissues were extracted from the spine samples by removing the surrounding tissue, including vertebrae and muscles (Figure 1B). A 3D-printed SC holder was created² (netfabb Professional 5.0, Projet 6000 HD printer with Visijet Sl Tough material) (Figure 1C) and was used to insert all the extracted SC tissues in a 15 ml Falcon tube filled with Fomblin (Figure 1D). These extracted SC tissues were imaged on the same scanner with a 25 mm ID volume coil (Bruker Biospin). Images were acquired using a 3D T2* weighted gradient-echo (TR/TE: 50/10 ms, FA: 12°) with 70 μm isotropic in-plane resolution and 200 μm slice thickness. The total scan time for the entire spine was 12 h in one session. All images were processed offline (MIPAV and ImageJ, NIH) to generate volumes of the entire SC.At equivalent signal-to-noise level (SNR_{non-extracted} = 39.7 versus SNR_extracted = 35.5) and similar voxel size (1 nL for non-extracted versus 0.98 nL for extracted), it was possible to image the extracted SC tissues in a substantially shorter acquisition time (30 hours for non-extracted versus 12 hours for extracted). Due to the presence of various tissues (nerve roots, vertebrae, muscles) around the spinal cord, images of the non-extracted SC displayed non-negligible signal dropout on the edges of the SC white matter (Figure 2, top). These image artifacts were absent from the extracted SC images (Figure 2, bottom), thus enabling better visualization of the SC white matter. Another benefit of the extraction was to correct for the curvature of the SC (Figure 3, sagittal views). SC images from the healthy control displayed excellent gray-to-white matter contrast and homogeneous signal intensity across the white matter (Figure 3, left). SC images from EAE animals also showed excellent lesion-to-white matter contrast, which allowed for accurate detection of focal area of abnormal signal and confluent rim-like abnormal signal rims along the edges of the SC (Figure 3, right).Using our proposed methodology, we were able to obtain high-quality artifact-free images of the entire ex vivo marmoset spinal cord with an overnight scan. This methodology opens the door for accurate evaluation of the spinal cord lesion distribution in this animal model of MS.