Rabbit anti olig2

The organoids were fixed and processed for immunostaining as is or after cryosectioning. Primary antibodies used for immunostaining were as follows: rabbit anti-SOX2 (Millipore), mouse anti-ISL1 (DSHB), rabbit anti-OLIG2 (IBL), mouse anti-NeuN (Chemicon), goat anti-NKX6.1 (R&D Systems), guinea pig anti-CHX10^{17 (link)}, rabbit anti-TUJ1 (BioLegend), mouse anti-TUJ1 (Millipore), rabbit cleaved-caspase3 (Cell Signaling Technology), goat anti-T (R&D Systems), rabbit anti-GFP (Abcam), goat anti-CHAT (Chemicon), and mouse anti-SMI-32 (BioLegend). After cell washes, samples were incubated with secondary antibodies and counterstained with Alexa Fluor® 488 phalloidin (Invitrogen) for axon labeling and Hoechst 33342 (Thermo Fisher Scientific) for nuclei staining.

Bielschowsky silver staining and immunohistochemistry were carried out according to standard procedures. The following primary antibodies were used: rabbit anti-MBP (1:1,000, Dako, Denmark), mouse anti-MBP peptide_70–89 (SMI94; 1:3,000, Covance, Princeton, New Jersey, USA), mouse anti-KiM1P (1:5,000) [43 (link)], rabbit anti-Olig2 (1:100, IBL, Spring Lake Park, Minnesota, USA), mouse anti-NogoA (1:10,000, mAb 11C7, a generous gift from M.E. Schwab, Brain Research Institute, ETH and University of Zurich, Switzerland) and mouse anti-Ki67 (1:50, clone Mib-1, Dako, Denmark). To identify Olig2⁺ or NogoA⁺ cells in control non-MS, normal-appearing MS and demyelinated MS neocortex, we performed double-immunolabeling combining either rabbit anti-Olig2 with mouse anti-MBP_70–89 (SMI94) or mouse anti-NogoA with rabbit anti-MBP.
Double-labeling immunohistochemistry was performed combining DAB (first primary antibody) and Fast Blue (second primary antibody with APAAP; Dako). For fluorescence double-labeling, Cy3- or Cy2-conjugated goat anti-mouse IgG and goat anti-rabbit IgG (both from Jackson ImmunoResearch Europe Ltd.) were used.