Figure S5.

Multiplexed Ion Beam Imaging Acquisition and Analysis, Related to Figure 5

(A) Workflow for image acquisition and standard processing to achieve marker quantification for single cells. (B) Heatmap representing scRNA-seq expression for genes exhibiting similar expression patterns in MIBI and scRNA-seq. (C) Plot of cell positions and tumor or normal status in large, tiled image from Patient 8 (2mm x 2mm). Regions outlined by squares are analyzed for composition of non-tumor cells (similar to Figure 5C). (D) Distribution of mean distance to Tregs for each CD4 T cell in blue. In red, CD4 cell identities were permuted among all non-tumor, non-CD4, non-Treg cells and the mean distance to Treg was re-calculated. (E) Similar to (D), analyzing macrophage to Treg distances. (F) Comparison of CD8 versus Treg differential expression and correlation with FOXP3 across ST spots for patients 2 and 4. (G) Spatial feature plots of Treg marker FOXP3 and CD8 effector/chemokine genes demonstrating co-localization. (H) Similar to Figure 5K. Cells were labeled as stromal (tumor-excluded), leading edge (tumor-stroma border), and infiltrated in tumor parenchyma.