CODEX multiplexed imaging of immunotherapy in human and mouse melanomas

Our research used CODEX (Co-Detection by Indexing) multiplexed imaging to gain insights into melanoma tumors in both murine models and human samples. CODEX imaging involves an iterative process of annealing and stripping fluorophore-labeled oligonucleotide barcodes, complementing the barcodes attached to over 40 antibodies used for tissue staining. Subsequently, images underwent standard CODEX image processing (tile stitching, drift compensation, cycle concatenation, background subtraction, deconvolution, and determination of best focal plane), single cell segmentation, and column marker z-normalization by tissue. Our datasets comprise individual cells as rows, each characterized by 40+ antibody fluorescence values quantified from various markers evaluated for each study. These markers correspond to the antibodies targeting specific proteins within the tissue, quantified at the single-cell level. The values represent per-cell/area-averaged fluorescent intensities, z-normalized along eac..., For a detailed description of each of the steps of protocols and processes to obtain this data see the detailed materials and methods in the associated manuscript. Briefly, murine melanomas were extracted from sacrificed mice at indicated timepoint post therapy and frozen in OCT. These were assembled into an array of 4-10 tissues, cut into 7 um slices, and stained with a panel of 40+ CODEX DNA-oligonucleotide barcoded antibodies. Tissues were imaged with a Keyence microscope at 20x objective and then processed using image stitching, drift compensation, deconvolution, and cycle concatenation. For human melanoma samples, we took available tumor tissue and cut with a 3µm section thickness and mounted onto Superfrost PLUS slides for further staining with a panel of 58 CODEX DNA-oligonucleotide barcoded antibodies imaged with the Phenocycler Fusion machine and processed using manufacturer’s v1.6.0 (e.g., background-subtracted, stitched, and shading correction). Processed data were then segme..., , # CODEX multiplexed imaging of immunotherapy in human and mouse melanomas Our research used CODEX (Co-Detection by Indexing) multiplexed imaging to gain insights into melanoma tumors in both murine models and human samples. CODEX imaging involves an iterative process of annealing and stripping fluorophore-labeled oligonucleotide barcodes, complementing the barcodes attached to over 40 antibodies used for tissue staining. Subsequently, images underwent standard CODEX image processing (tile stitching, drift compensation, cycle concatenation, background subtraction, deconvolution, and determination of best focal plane), single cell segmentation, and column marker z-normalization by tissue. ## Description of the data and file structure Our datasets comprise individual cells as rows, each characterized by 40+ antibody fluorescence values quantified from various markers evaluated for each study. These markers correspond to the antibodies targeting specific proteins within the tissue, qua...