This dataset includes 3D scanning electron microscopy (SEM) images of a female mouse corpus callosum and MATLAB code for random walker (RaW) segmentations of myelinated axons. The code additionally characterizes inner axonal diameters and fiber orientation dispersion within segments of the intra-axonal spaces and myelin sheaths.

In the study, a female 8-week-old C57BL/6 mouse was perfused trans-cardiacally using a fixative solution. The genu of corpus callosum was later excised from the midsagittal slice of the dissected brain, and the tissue was sampled from the central region of the genu and was fixed in the same fixative solution. The tissue was then stained and an En Bloc lead staining was performed to enhance membrane contrast. The brain sample was dehydrated in alcohol and acetone, and embedded in Durcupan ACM resin. Then, the tissue sample was analyzed with SEM.

The SEM dataset includes 4 files:

• datac.nii (a stack of SEM data, 200 slices; resolution: 24nm by 24nm by 100nm; volume: 36μm by 48μm by 20μm)
• maskc.nii (foreground mask)
• myelin_mask.nii (myelin mask generated by pixel-wise classifier in ilastik)
• fibers.nii (intra-axonal space segmented by using random-walker-based approach)