Multiplex Single-Cell Bioprinting for Engineering of Heterogeneous Tissue Constructs with Subcellular Spatial Resolution.

Tissue development, function, and disease are largely driven by the spatial organization of individual cells and their cell-cell interactions. Precision engineered tissues with single-cell spatial resolution, therefore, have tremendous potential for next generation disease models, drug discovery, and regenerative therapeutics. Despite significant advancements in biofabrication approaches to improve feature resolution, strategies to fabricate tissues with the exact same organization of individual cells in their native cellular microenvironment have remained virtually non-existent to date. Here we report a method to spatially pattern single cells with up to eight cell phenotypes and subcellular spatial precision. As proof-of-concept we first demonstrate the ability to systematically assess the influence of cellular microenvironments on cell behavior by controllably altering the spatial arrangement of cell types in bioprinted precision cell-cell interaction arrays. We then demonstrate, for the first time, the ability to produce high-fidelity replicas of a patient's annotated cancer biopsy with subcellular resolution. The ability to replicate native cellular microenvironments marks a significant advancement for precision biofabricated in-vitro models, where heterogenous tissues can be engineered with single-cell spatial precision to advance our understanding of complex biological systems in a controlled and systematic manner.