Standard Hematoxilin & Eosin, Cresyl Violet and Luxol Fast Blue histochemical stainings were performed on paraffin sections and digitized using Mirax (Carl Zeiss) or Aperio (Leica) slide scanners as described previously [79 (link)]. Immunofluorescent detection was performed on free-floating sections as described [80 (link)] using the following antibodies: rabbit anti-ASPA serum (in house), mouse anti-NeuN (Merck), rat anti-MBP (Abcam), mouse anti-GFAP (Cell Signaling), chicken anti-β-Gal (Abcam), mouse anti-Flag (Cell Signaling) and rabbit anti-Neurofilament 200 (Sigma) followed by the appropriate Alexa-488/594 conjugated secondary antibodies (Thermo Fisher). Immunoperoxidase detection was performed on free-floating sections as described in [15 (link)] using mouse anti-APC (Merck) and rabbit anti-NeuN (Cell Signaling), the appropriate biotinolated secondary antibody (Dianova) and a Vectastain Elite ABC kit (Vector Labs). Immunoblotting was performed as described previously [78 (link)] with the following antibodies: rabbit anti-ASPA serum, mouse anti-GAPDH (Sigma), rat anti-MBP (Abcam), rat anti-PLP aa3 and rat anti-NG2 (gift of J. Trotter) followed by the appropriate HRP-conjugated secondary antibodies (Dianova). qRT-PCR for Aspa, Nat8l, NAAG synthetase-I and II (Rimklb and Rimkla) and hypoxanthine phosphoribosyltransferase (Hprt) was performed as described [26 (link)].
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