From Free‐Radical to Radical‐Free: A Paradigm Shift in Light‐Mediated Biofabrication

In recent years, the development of novel photocrosslinking strategies and photoactivatable materials has stimulated widespread use of light‐mediated biofabrication techniques. However, despite great progress toward more efficient and biocompatible photochemical strategies, current photoresins still rely on photoinitiators (PIs) producing radical‐initiating species to trigger the so‐called free‐radical crosslinking/polymerization. In the context of bioprinting, where cells are encapsulated in the bioink, the presence of radicals raises concerns of potential cytotoxicity. In this work, a universal, radical‐free (RF) photocrosslinking strategy to be used for light‐based technologies is presented. Leveraging RF uncaging mechanisms and Michael addition, cell‐laden constructs are photocrosslinked by means of one‐ and two‐photon excitation with high biocompatibility. A hydrophilic coumarin‐based group is used to cage a universal RF photocrosslinker based on 4‐arm‐PEG‐thiol (PEG4SH). Upon light exposure, thiols are uncaged and react with an alkene counterpart to form a hydrogel. RF photocrosslinker is shown to be highly stable, enabling potential for off‐the‐shelf products. While PI‐based systems cause a strong upregulation of reactive oxygen species (ROS)‐associated genes, ROS are not detected in RF photoresins. Finally, optimized RF photoresin is successfully exploited for high resolution two‐photon stereolithography (2P‐SL) using remarkably low polymer concentration (<1.5%), paving the way for a shift toward radical‐free light‐based bioprinting.