Infiltrated immune cells isolated from 5 chemosensitivet breast cancer patients
CD45+ immune cells in tumor suspensions were isolated through FACS with dead-cell exclusion and doublets discard, at least 5,000 cells were collected per patient and cells from 5 patients with the same chemotherapeutic efficacy were mixed as a sequencing sample. The cells were then resuspended at approximately 20000 cells/mL with a final viability of > 90% as determined. Single-cell capture, lysis, cDNA library construction, Generation of gel beads in Emulsion (GEM), barcoding, post GEM-reverse transcription cleanup and cDNA amplification were performed using Chromium system from 10X Genomics, with the following kits: Chromium Single Cell 3 Library & Gel Bead Kit v2 kit and Chromium Single Cell A Chip Kits. Amplified cDNA was purified using SPRIselect beads and sheared to 250-400 bp.Qualification was performed using Qubit 3.0 Fluorometer. Libraries were sequenced on an Illumina NovaSeq 6000 system.