Anti doublecortin

Sections were washed 3 times in PBS before proceeding with immunostaining. BrdU detection required antigen retrieval (steaming at 95°C in 0.01 mol/L citrate buffer for 10 min) and incubation in 2N HCl (30 min) before antibody incubation. For all other stains (including double staining), sections underwent antigen retrieval, then incubation in 33% normal serum in PBS (1 h) and primary antibody (overnight) at room temperature in 0.3% Triton X-100 and 1% normal serum in PBS. The primary antibodies used were: mouse monoclonal anti-BrdU (1:100; Becton-Dickinson, San Jose, CA), rabbit monoclonal anti-cleaved Caspase 3 (1:300; Cell Signaling, Beverly, MA), rabbit polyclonal anti-Ki67 (1:500; Abcam, ab15580, Cambridge, England), rabbit polyclonal anti-Sox2 (1:1000; Abcam, ab97959), rabbit polyclonal anti-Tbr2 (1:300, Abcam, ab115986) and guinea pig polyclonal anti-Doublecortin (1:1000; Millipore, ab5910, Temecula, CA). For fluorescent staining, the secondary antibodies used were: Alexa Goat anti-Mouse 488, Alexa Goat anti-Rabbit 594, and Alexa Goat anti-Guinea pig 594 (Molecular Probes, Eugene, OR).

Adult mice were anesthetized, perfused with 4% PFA. Brain tissue was dissected out, equilibrated in 30% sucrose, and sectioned into 40 μm-thick serial sections. The brain sections were washed in PBS for 15 min three times, and then blocked in blocking solution (3% BSA+0.3%Triton X-100+0.2% sodium azide) at room temperature for 1 h. The primary antibodies we used are as follows: anti-H3K27me3 (1:1,000, 07449, Millipore), anti-Map2 (1:1,000, Mab3418, Millipore), anti-GFP (1:1,000, A10262, Life technology), anti-NeuN (1:1,000, millipore), anti-Doublecortin (1:500, millipore). After incubation in primary antibody solution at 4°C overnight, the brain sections were washed with TBS for 30 min three times and then incubated with the secondary antibodies conjugated to Alexa Fluor 488 or 594 with a concentration of 1:500 at room temperature. The sections were finally stained with DAPI and mounted using adhesion anti-fade medium.