Mouse anti 6e10

The cells were fixed with 4% paraformaldehyde for 15 min. After several washes, they were blocked with 0.01M phosphate-buffered saline supplemented with 1% BSA and 0.3% Triton X-100 for 30 min. Primary antibodies (mouse anti-LAP2α, 1:500, Abcam, UK; mouse anti-betaIII tubulin, 1:500, Abcam; mouse anti-NeuN, 1:1000, Abcam; rabbit anti-NeuN, 1:200, Abways; rabbit anti-Tbr1, 1:500, Abcam; mouse anti-γH2AX, 1:250, Millipore, USA; mouse anti-Lamin A/C, 1:200, Abcam; rabbit anti-H3K9me3, 1:4000, Abcam; mouse anti-6E10, 1:200, Abcam) were diluted with blocking buffer and applied overnight at 4° C. The cells were stained with suitable Alexa Fluor–labeled secondary antibodies (Invitrogen) in blocking buffer at a concentration of 1:500 for 1.5 h at room temperature. 4', 6-Diamidino-2-phenylindole (Thermo Fisher, IL, USA) was applied to counterstain cells for visualizing nuclei at 1:1000 in Milli Q water (Biocel, Millipore, USA). The images were obtained with an Olympus IX71 microscope using a Hamamatsu ORCA CCD camera and Leica TCS SP5.

After being anesthetized with 4% isoflurane, mice were perfused transcardially with saline and 4% paraformaldehyde (PFA). Brains were removed and fixed with 4% PFA, and then cut into 30 μm thick coronal sections. The sections were treated with 3% H₂O₂ for 10 min and incubated overnight at 4 °C with the primary antibodies mouse anti-A_2AR (1:500, Abcam, Berlin, Germany) or mouse anti-6E10 (1:500, Abcam, Berlin, Germany), diluted in 3% bovine serum albumin (BSA) and 0.3% Triton X-100. Subsequently, the sections were incubated with biotinylated goat anti-mouse secondary antibody (1:1000, Invitrogen, Carlsbad, CA) for 2 h at room temperature. After incubation, the sections were developed using a DAB kit (Vector Laboratories Inc.). The immunohistochemistry was visualized using a microscope (Olympus, Tokyo, Japan).