Mabn377

In the immunofluorescence staining, the brain sections from monkey Middle age 2 were washed for 5 min in 0.01 mol/L PBS containing 5% bovine serum albumin (BSA) and 0.3% Triton X-100, and then incubated with a set of primary antibodies (in 0.01 mol/L PBS with 1% BSA and 0.3% Triton X-100) overnight at 4 °C.
In gene-editing effect tests, two sets of primary antibodies were used: (1) anti-EGFP (ab6673, Abcam), anti-NeuN (ABN78, Millipore), anti-Paris (75-195, NeuroMab); and (2) anti-EGFP (ab13970, Abcam), anti-NeuN (MABN377, Millipore), and anti-DJ-1 (ab201147, Abcam).
For glial cell co-staining, another set of primary antibodies was used: anti-GFAP (SMI-21R, Covance Research Products Inc.), anti-Iba1 (019-19741, Wako), and anti-EGFP (ab13970, Abcam).
All the sections were subsequently treated with the corresponding secondary antibodies (1:1000, Alexa-Fluor-conjugated, Invitrogen). DAPI was used to label nuclei and the sections were mounted with 75% glycerol for microscopy.

Coronal sections (80 μm) were obtained with a vibratome (Leica, Germany), permeabilized with 0.2% Triton X-100 (Sigma-Aldrich) and incubated with 3% bovine serum albumin (Sigma-Aldrich). After standard antigenic retrieval using sodium citrate, the free-floating sections were incubated overnight with the following primary antibodies: anti-Caspase-3 (1:300, Millipore, AB3623); rabbit anti-Ki67 (1:100, Millipore, AB9260); mouse anti-NS1 (1:10, Thermo Fisher Scientific, MA5-24585); rabbit anti-SOX2 (1:200, Sigma-Aldrich/Millipore, AB5603); rabbit anti-SOX9 (1:200, Abcam, ab185230); mouse anti-IBA1 (1:1000, Millipore, MABN92); mouse anti-NEUN (1:200, Millipore, MABN377); and rabbit anti-OLIG2 (1:300, Millipore, MABN50). Following the overnight incubation, sections were washed with PBS and incubated for 2 h with goat anti-rabbit Alexa Fluor 488 (1:500, Millipore, AP132JA4) and goat anti-mouse Alexa Fluor 546 (1:500, Invitrogen, A11003) secondary antibodies. The free-floating sections that were stained with isolectin B4 (IB4; 1:200, Invitrogen, 121411) were incubated with the stain together with the secondary antibodies. Nuclei were stained with DAPI (0.5 μg/ml) for 30 min. Sections were placed in glass slides and mounted with Fluoromount (Sigma-Aldrich, F4680).