CAR-T cells, treated with Ponatinib
FACS-sorted CD8+ CAR-T cells (CD3CD8EGFR) were isolated from the peripheral blood of healthy donors and prepared for single-cell RNA sequencing (scRNA-seq). Cells were treated with either 100 nM Ponatinib or DMSO as a control. scRNA-seq libraries were constructed using the Chromium Next GEM Single Cell 3 Reagent Kit v3.1 (10x Genomics) according to the manufacturers protocol. Each sample was processed separately without pooling before library preparation. Unique molecular identifiers (UMIs) and barcodes were incorporated using microfluidics. Sequencing was performed on an Illumina NovaSeq X Plus platform with paired-end reads. Raw base call files were demultiplexed into FASTQ format using the "Cellranger mkfastq" pipeline, and the "Cellranger count" pipeline was used for read filtering, alignment to the hg41 reference genome, and gene expression quantification.