In Situ Hydrogelation of Cellular Monolayers Enables Conformal Biomembrane Functionalization for Xeno‐Free Feeder Substrate Engineering

Abstract The intricate functionalities of cellular membranes have inspired strategies for deriving and anchoring cell‐surface components onto solid substrates for biological studies, biosensor applications, and tissue engineering. However, introducing conformal and right‐side‐out cell membrane coverage onto planar substrates requires cumbersome protocols susceptible to significant device‐to‐device variability. Here, a facile approach for biomembrane functionalization of planar substrates is demonstrated by subjecting confluent cellular monolayer to intracellular hydrogel polymerization. The resulting cell–gel hybrid, herein termed GELL (gelated cell), exhibits extraordinary stability and retains the structural integrity, membrane fluidity, membrane protein mobility, and topology of living cells. In assessing the utility of GELL layers as a tissue engineering feeder substrate for stem cell maintenance, GELL feeder prepared from primary mouse embryonic fibroblasts not only preserves the stemness of murine stem cells but also exhibits advantages over live feeder cells owing to the GELL's inanimate, non‐metabolizing nature. The preparation of a xeno‐free feeder substrate devoid of non‐human components is further shown with HeLa cells, and the resulting HeLa GELL feeder effectively sustains the growth and stemness of both murine and human induced pluripotent stem cells. The study highlights a novel bio‐functionalization strategy that introduces new opportunities for tissue engineering and other biomedical applications.