2.4  Immunohistochemistry
DAB-immunostaining was performed according to the manufacturer׳s instruction (Thermo-Scientific; Ultravision LP Detection System). The wild type samples were sectioned at 10 μm. Antigen retrieval was performed with citrate buffer (10 mM sodium citrate, 0.05% Tween 20, pH 6.0) at 68 °C for 20 min and 0.3% Triton at room temperature for 30 min. The endogenous peroxidase activity was minimized with hydrogen peroxide incubation for 15 minutes for DAB staining. Two mouse monoclonal anti-SGO1 antibodies (abcam; ab58023 or abnova: M01_clone 3C11; 1/350 in 10% goat serum) were used separately and compared in representative slides to support the specificity of the SGO1 localisation. TUBB3 and RBPMS antibodies were acquired as the courtesy of Alyson Fournier׳s lab at Montreal Neurological Institute. The primary antibodies were targeted with rabbit anti-mouse IgG secondary antibody conjugated to horseradish peroxidase for DAB staining and goat anti-mouse Alexa 488 for fluorescent staining. DAB slides were counterstained with methyl green and mounted with Permount medium. Fluorescent slides were mounted with DAPI incorporated anti-fade medium.