title: Single-cell transcriptomes of sorted CD45+ cells in brains were performed by Single Cell Genomics Facility in Cincinnati Children's Hospital Medical Center following established protocols. The final concentration of sorted single cells was about 1,500 cells/uL. Single-cell suspensions were processed using the 10x Genomics Chromium platform (10x Genomics) to generate Gel Beads-in-Emulsion (GEMs), enabling the encapsulation of individual cells with barcoded gel beads and reverse transcription reagents. GEM formation and reverse transcription were performed according to the manufacturers instructions using GEM-X Chromium Single Cell 5 Library & Gel Bead Kit v3. Sequencing libraries were generated through enzymatic fragmentation, end repair, A-tailing, adapter ligation, and sample index PCR, following the standard 10x Genomics protocol. The final Illumina-ready libraries were sequenced on a NovaSeq 6000 system (Illumina, San Diego, CA, USA), targeting approximately 50,000 reads per cell.
design: Single-cell transcriptomes of sorted CD45+ cells in brains were performed by Single Cell Genomics Facility in Cincinnati Children's Hospital Medical Center following established protocols. The final concentration of sorted single cells was about 1,500 cells/uL. Single-cell suspensions were processed using the 10x Genomics Chromium platform (10x Genomics) to generate Gel Beads-in-Emulsion (GEMs), enabling the encapsulation of individual cells with barcoded gel beads and reverse transcription reagents. GEM formation and reverse transcription were performed according to the manufacturers instructions using GEM-X Chromium Single Cell 5 Library & Gel Bead Kit v3. Sequencing libraries were generated through enzymatic fragmentation, end repair, A-tailing, adapter ligation, and sample index PCR, following the standard 10x Genomics protocol. The final Illumina-ready libraries were sequenced on a NovaSeq 6000 system (Illumina, San Diego, CA, USA), targeting approximately 50,000 reads per cell.