Aperio scanscope at turbo system

Formalin-fixed, paraffin-embedded tissue blocks were cut into 6 µm-thick adjacent sections for immunohistochemistry using optimized methods.^{21 (link)} Briefly, adjacent sections were baked at 60°C for 20 min, deparaffinized in xylene and rehydrated with successive ethanol baths before removal of endogenous peroxidase and appropriate antigen retrieval procedures. Adjacent sections were incubated with primary antibodies for: myelin (proteolipid protein, PLP), microglial/macrophage inflammation (CD68/PG-M1), T-lymphocytes (CD3), B-lymphocytes (CD20), complement deposition (C9neo), acute axonal injury (beta-amyloid precursor protein, β-APP), glial fibrillary acid protein (GFAP) and aquaporin-4 channel (AQP4), as specified in Supplementary Table 1, with subsequent labelling with secondary antibody and 3,3′-diaminobenzidine (DAB) visualization.^{21 (link)} Sections were counter-stained with haematoxylin. Omission of the primary antibody was performed to confirm specificity of the immunoreaction. Observers were blind to disease category. Immunohistochemistry sections were imaged and visualized using AxioVision software (v4.9.1, Zeiss) and digitally scanned using Aperio ScanScope AT Turbo system (Leica Biosystems) at ×400 magnification for downstream analyses.