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Digital spatial profiling platform allows for spatially-resolved, multiplexed measurement of solid tumor protein distribution and abundance in FFPE tissue sections

      Normal 0 false false false EN-GB X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin-top:0in; mso-para-margin-right:0in; mso-para-margin-bottom:8.0pt; mso-para-margin-left:0in; line-height:107%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri",sans-serif; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-ansi-language:EN-GB;} Characterization of the spatial distribution and abundance of proteins within tissues enables a better understanding of biological systems in many research areas, including immunology and oncology. However, it has proven difficult to perform such studies in a highly multiplexed manner. To address this unmet need, we have developed a novel optical-barcode based microscope and tissue-sampling platform designed to simultaneously analyze hundreds of proteins on a single FFPE section (Digital Spatial Profiling, DSP). DSP probes are not multiplex-limited by spectral resolution. Instead, “colors” are determined using barcode indexing oligos that are conjugated to antibodies. These indexing oligos are conjugated via UV-cleavable linkers and, following UV light exposure, are released and siphoned off the tissue surface via a microcapillarytip. This UV-cleavage is precisely controlled by a digital micromirror device that can illuminate discrete regions ranging from entire tissue microenvironments to single-cells. For quantification of signal, the photocleaved oligos are hybridized to Nano String barcodes, providing digital counts of the protein in each region-of-interest using Nano String nCounter instruments. Using this novel approach, we spatially resolve over 30 solid tumor targets, simultaneously, on Her2+ breast cancer samples and Diffuse Large B-Cell Lymphoma samples. Solid tumor targets were probed with antibodies that recognize phosphorylated or total protein content (e.g. Her2, Ki67, P-AKT, P-ERK). We demonstrate multiplexed detection from discrete regions within a tumor and the adjacent normal tissue, enabling systematic interrogation of immune activity in FFPE samples. We also show the ability to profile either whole tissue sections or tissue microarrays. Finally, we validate that indexing oligo conjugation and high multiplexing of antibodies do not interfere with specificity. The simplicity of the DSP platform allows high-resolution, high-multiplexed, spatially-resolved protein characterization in any lab capable of performing immunohistochemistry procedures, providing a potential method that can bridge the gap between translational research discovery and clinical applications. Continued work on the platform will expand the library of protein targets accessible for profiling and future assay development will demonstrate multiplexing up to 800 targets.
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