Localization of Multiple Hydrogels with MultiCUBE Platform Spatially Guides 3D Tissue Morphogenesis In Vitro

Localization of multiple hydrogels is expected to develop the structure of 3D tissue models in a location‐specific manner. Here, 3D tissue morphogenesis is spatially guided by localizing different hydrogel conditions at different parts of a tissue. To achieve the localization, a unit‐based scaffold is developed with a unique frame design to trap hydrogel solutions inside their designated units. An optimal range of unit dimensions and surface wettabilities enables a solution trapping up to several cubic millimeters without any need for chemical additives. This capability allows spatial organization of biomolecular compositions and physical conditions of hydrogels, as well as the relative position of biological samples (cells, spheroids, and reconstituted tissues) within the scaffold. Successful localization of branching development on reconstituted human epithelial tissues is achieved by localizing growth factors or cross‐linked matrix proteins within hydrogels, demonstrating a direct dependence on local hydrogel conditions. Unlike 3D‐bioprinting or microfluidic techniques, this scaffold‐based localization of hydrogels requires only a manual pipetting and no specialized tools, making it ready‐to‐use for researchers from any field. This localization technique provides a new promising route to spatially control morphogenesis, differentiation, and other developmental processes within 3D organoids or tissue models for practical biomedical applications in the future. The unit‐based scaffold (MultiCUBE) is developed to localize multiple hydrogel solutions. Localization of different matrix composition, growth factor, or cross‐linking condition within hydrogels leads to a location‐specific tissue development such as 3D angiogenesis toward tumor spheroid and 3D localized branching morphogenesis of reconstituted bronchial tissues. This technology can be potentially applied to 3D organoid culture for future biomedical applications.