Spatially resolved whole transcriptome profiling in human and mouse tissue using Digital Spatial Profiling

Abstract

Emerging spatial profiling technology has enabled high-plex molecular profiling in biological tissues, preserving the spatial and morphological context of gene expression. Here we describe expanding the chemistry for the Digital Spatial Profiling platform to quantify whole transcriptomes in human and mouse tissues using a wide range of spatial profiling strategies and sample types. We designed multiplexed in situ hybridization probes targeting the protein-coding genes of the human and mouse transcriptomes, referred to as the human or mouse Whole Transcriptome Atlas (WTA). Human and mouse WTAs were validated in cell lines for concordance with orthogonal gene expression profiling methods in regions ranging from ~10-500 cells. By benchmarking against bulk RNA-seq and fluorescence in situ hybridization, we demonstrate robust transcript detection down to ~100 transcripts per region. To assess the performance of WTA across tissue and sample types, we applied WTA to biological questions in cancer, molecular pathology, and developmental biology. Spatial profiling with WTA detected expected gene expression differences between tumor and tumor microenvironment, identified disease-specific gene expression heterogeneity in histological structures of the human kidney, and comprehensively mapped transcriptional programs in anatomical substructures of nine organs in the developing mouse embryo. Digital Spatial Profiling technology with the WTA assays provides a flexible method for spatial whole transcriptome profiling applicable to diverse tissue types and biological contexts.

Ultra High-Plex Spatial Proteogenomic Investigation of Giant Cell Glioblastoma Multiforme Immune Infiltrates Reveals Distinct Protein and RNA Expression Profiles

“…In contrast, for the assay under slightly acidic HIER conditions, the FPR increased to 30%. The high FPR associated with slightly acidic HIER conditions is consistent with the previous observation: an increase in non-specific hybridization when epitope retrieval was performed under slightly acidic conditions (25).…”

“…RNA only control slides were processed according to the RNA FFPE BOND RX Slide Preparation Protocol in the GeoMx-NGS Slide Preparation User Manual for FFPE (MAN-10151) and associated published materials (22,23,25).…”

“…For the spatial proteogenomic slides, GeoMx Digital Spatial Profiling was carried out according to GeoMx-DSP Spatial Proteogenomic Protocol (MAN-10158); whereas the control slides were processed according to GeoMx-NGS DSP Instrument User Manual (MAN-10152) and as described by Merritt, et al (23,25). For CPAs, two geometric regions of interest (ROIs) of 200 µm in diameter were profiled per cell line.…”

“…The GeoMx DSP enables spatially resolved, high-plex digital quantitation of proteins (> 100-plex) and RNA (whole transcriptome, > 18,000-plex) from human and mouse tissues (22)(23)(24)(25). Previous work has detailed proteomic or transcriptomic single-analyte workflows using the GeoMx DSP technology with unique affinity reagents (antibodies for protein or in situ hybridization (ISH) probes for RNA) coupled to photocleavable oligonucleotide barcodes (22,23,25) (Fig. 1B).…”

Multi-omic spatial profiling reveals the unique virus-driven immune landscape of COVID-19 placentitis

“…Virus positivity of each region was also assigned by a registered pathologist (M.P.). The NanoString Whole Transcriptome Atlas (WTA) gene panel was used 52 . Oligonucleotides were cleaved off the antibodies and gathered within each ROI as per the GeoMx DSP protocol.…”