scRNA-seq was performed on ESCC tissues obtained from epithelial conditional knockout CD276 (K14CreER, Cd276flox/flox) mice and control mice (Cd276wt/wt) to elucidate the main mechanism by which CD276 regulates immune escape in ESCC. To prepare the single-cell suspension, esophageal tissues were microdissected and disaggregated into single cells using enzymatic digestion with mouse Tumor Dissociation Kit (130-096-730, Macs Miltenyi Biotec, China) at 37 °C for 45 min using gentleMACS Dissociator (130-093-235, Macs Miltenyi Biotec, China). For scRNA-seq library preparation, we used the Chromium Single Cell 3′ v3 (10× Genomics, Pleasanton, California, USA) following the manufacturer’s instructions.
Seurat R package was employed to analyze and visualize the scRNA-seq data.37 (link) Briefly, we identified high-variable genes, capturing heterogeneity and key regulatory factors. Next, principal component analysis reduced dimensionality, enabling visualization of cellular heterogeneity, and identifying cell clusters based on gene expression patterns.
Seurat R package was employed to analyze and visualize the scRNA-seq data.37 (link) Briefly, we identified high-variable genes, capturing heterogeneity and key regulatory factors. Next, principal component analysis reduced dimensionality, enabling visualization of cellular heterogeneity, and identifying cell clusters based on gene expression patterns.
