title: Transcriptomic profiling of BrO, DMGO and GD2 CAR T cells
design: Transcriptomic profiling of BrO, DMGO and GD2 CAR T cells
library_construction_protocol: BrOs and DMGOs containing microglia and optionally treated with GD2 CAR T cells were dissociated 21 days after initial microglia incorporation with the Neural Tissue Dissociation Kit (P) (Miltenyi Biotec, Cat. #130-092-628), as described above for preparation of single cell RNA and tracker seq libraries. Dissociated cells, control PMPs and unexposed GD2 CAR T cells were washed and stained in FC buffer with CD3-BV421 (1:100; BD Biosciences, clone SK7) and LIVE/DEAD Fixable Near-IR Dead Cell Stain (1:1000; Thermo Fisher) for 30 min at 4ËšC. CD3+ T cells and mScarlet+ microglia/ PMP were sorted into 386-well plates containing well-specific barcoded primers (Single Cell Discoveries), one cell per well, on a Sony SH800s (Sony Biotechnology). Plates containing sorted cells were immediately snap frozen on dry ice and processed for SORT-seq by Single Cell Discoveries.  Nuclei acid extraction was provided by Single Cell Discoveries (Utrecht, Netherlands)  Library generation was provided by Leiden Genome Technology Center (Leiden, The Netherlands) Library generation was provided by Single Cell Discoveries (Utrecht, Netherlands) 