O4 apc vio770

Generation of single-cell suspension was performed as described in Llorens-Bobadilla et al.⁶ (link) Cells were stained with the following antibodies: O4-allophycocyanin (APC) and O4-APC-Vio770 (Miltenyi; diluted 1:50), Ter119-APC-Cy7 (BioLegend; 1:100), CD45-APC-Cy7 (Becton Dickinson [BD]; 1:200), GLAST (ACSA-1)-phycoerythrin (PE; Miltenyi: 1:20), CD9-eFluor450 (eBioscience; 1:300), Alexa647::EGF (Life Technologies; 1:100), polysialylated neuronal cell adhesion molecule (PSA-NCAM)-PE-Vio770 (Miltenyi; 1:75), Prominin1- peridinin-chlorophyll-protein PerCP-eFluor 710 (eBioscience; 1:75), CD24-PE-Cy7 (eBioscience; 1:75), and Sytox Blue (Life Technologies; 1:1,000). For RNA-seq, cells were directly sorted into 100 μL of the PicoPure RNA Isolation Kit (Thermo Fisher Scientific) extraction buffer. For ex vivo transduction, NSCs were sorted into growth factor-free Neurobasal medium (NBM).

FACS analysis for testing the transduction efficiency of the candidate viruses was performed by two methods. The first method consisted of injecting 5-month-old TiCY mice with the AAV1_P5_Cre virus, and after 8 days, SVZ and OB cells were FACS analyzed (Figures 4G and 4H). In the second method, we injected 2-month-old C57BL/6N mice with AAV1_P5_YFP and AAV9_A2_YFP viruses and analyzed them after 6 days (Figures S3C and S3D).
For FACS quantification of AAV-injected NSC/progeny, cells were sorted with the following antibodies: O4-APC-Vio770 (Miltenyi; diluted 1:100), CD45-APC-Cy7 (BD; 1:200), Ter119-APC-Cy7 (BioLegend; 1:100), GLAST (ACSA-1)-PE (Miltenyi; 1:50), Prominin1-APC (eBioscience; 1:75), PSA-NCAM-PE-Vio770 (Miltenyi; 1:50), Texas-Red::EGF (Life Technologies; 1:75).