Monoclonal anti nestin

Immunodetection of neuronal markers was carried out as previously described [53 (link), 54 (link)]. Primary antibodies used were: rabbit anti-doublecortin (Cell Signaling Technology Inc.), monoclonal anti-NeuN (Millipore), goat anti-FZD1 (R&D Systems), goat anti-FZD1 (LifeSpan Biosciences, Inc.), rabbit anti-SOX2 (Cell Signaling Technology Inc.), monoclonal anti-GFAP (Sigma-Aldrich), monoclonal anti-Nestin (Millipore), rabbit anti-Ki67 (Abcam). As secondary antibodies, Alexa (Molecular Probes) and DyLight (Abcam) conjugated antibodies were used. NucBlue (Life Technologies) was used as nuclear dye. Slices were mounted on gelatin-coated slides with Fluoromont-G (Electron Microscopy Sciences). Double-labeled sections were analyzed by confocal laser microscopy (Olympus FV 1000). Image analysis and z-projections were made with ImageJ software (NIH, USA).

Immunodetection of BrdU and neuronal markers in tissue sections was carried out as previously described in [26 (link)]. Primary antibodies used were rat anti-BrdU (Abcam), rabbit anti-doublecortin (Cell Signaling Technology Inc.), monoclonal anti-NeuN (Millipore, Billerica, MA, USA), rabbit anti-Ki67 (Abcam), goat anti-NeuroD1 (Santa Cruz Biotechnology, Inc.), monoclonal anti-Nestin (Millipore), Alexa (Molecular Probes, Life Technologies), and DyLight (Abcam) conjugated secondary antibodies were used. Slices were mounted on gelatin-coated slides with Fluoromount-G (Electron Microscopy Sciences).