ABSTRACT Proper functioning of the human body relies on the organization of cells in 3D space. A cell’s function and fate are determined by its biomolecule composition and its 3D environment. The spatially identification of proteins, RNAs, and DNAs in a tissue thus provides a powerful map to decipher how cells build tissues and become diseased. Through the use of single-cell omics, it’s been possible to reveal rare cell types that benchmark development, oncogenesis, and brain functions. However, the cell isolation process in single-cell analysis unavoidably causes loss of spatial information. To obtain spatial information, spatial transcriptomics based on imaging or sequencing have emerged to give insight into the heterogeneous expression patterns in tumors, brain, and wound tissues. Unfortunately, most spatial transcriptomics methods can only examine thin tissue sections, and are incompatible with proteomics. To address these drawbacks, the goal of the project is to develop a conceptually novel 3D spatial multiomics technology featuring gel-based optical isolation (GO3D). The proposed GO3D technology is distinct from all current spatial omics, and will enable the profiling of proteins, RNAs, and DNAs of whole-mount tissues with subcellular resolution, high coverage and high throughput, simultaneously. This innovative design is based on the gel-based label-retention expansion microscopy (LR- ExM) that the PI published recently. GO3D will drastically transform our understanding of many critical biomedical questions, which we lack of tools to address currently. For example, how are cells in a highly dynamic skin migrate in 3D to heal wounds? How do specialized neurons build brain? And where do microbes interact with what cell types in gut?

NARRATIVE The project is relevant to NIH’s mission because the 3D spatial organization of cells is intimately linked to human health and disease. The proposed GO3D spatial multiomics will provide an ideal technology to map all major biomolecules of whole tissues in 3D. Applications of GO3D will transform our understanding of many critical biomedical processes, such as would healing mechanism, tumor structure and its microenvironment, neuron interactions in brain, and microbiome-host relationship.