Shelf‐Stable Human‐Derived Granular Extracellular Matrix (gECM) Hydrogel Wafers as 2.5D Substrates for Microphysiological Modeling

Understanding disease pathology and evaluating emerging therapeutics require in vitro models that accurately recapitulate human tissue environments. However, existing microphysiological systems often compromise either biomimicry or ease-of-use, limiting widespread adoption and scalability. Here, we present lyophilized granular extracellular matrix (gECM) hydrogel wafers as shelf-stable, humanized 2.5D substrates that enable physiologically relevant modeling while simplifying integration into experimental workflows. Derived from decellularized human cartilage and bone, our gECM hydrogel wafers retain tissue-specific architecture while introducing microporosity and surface topography through lyophilization. These wafers maintain swelling behavior, structural integrity, and mechanical properties over three months of room-temperature storage, allowing pre-fabrication and on-demand use without loss of function. gECM hydrogel wafers support direct cell seeding without encapsulation and sustain viability and proliferation of human adipose-derived mesenchymal stromal cells over 21 days. Furthermore, wafers can be readily integrated into proof-of-concept on-chip fabrication with in situ hydration and transport of large biomolecules. Together, this platform bridges the gap between conventional 2D culture ease-of-use and 3D biomaterial biomimicry, providing a scalable, physiologically-relevant, and humanized in vitro model approach that shows potential for future high-throughput disease studies and therapeutic screening.