Bioxolography Using Diphenyliodonium Chloride and N-Vinylpyrrolidone Enables Rapid High-Resolution Volumetric 3D Printing of Spatially Encoded Living Matter

Abstract

Light-based volumetric bioprinting enables fabrication of cubic centimeter-sized living materials with micrometer resolution in minutes. Xolography is a light sheet-based volumetric printing technology that offers unprecedented volumetric generation rates and print resolutions for hard plastics. However, the limited solubility and reactivity of current dual-color photoinitiators (DCPIs) in aqueous media have hindered their application for high-resolution bioprinting of living matter. Here, we present a novel three-component formulation that drastically improves photoreactivity and thereby enables high-resolution, rapid, and cytocompatible Xolographic biofabrication of intricately architected yet mechanically robust living materials. To achieve this, various relevant additives are systematically explored, which revealed that diphenyliodonium chloride and N-vinylpyrrolidone strongly enhance D-mediated photoreactivity, as confirmed by dual-color photo-rheology. This enables Xolographic bioprinting of gelatin methacryloyl-based bioresins, producing >1 cm³ constructs at ≈20 µm positive and 125 µm negative resolution within minutes. Multimaterial printing, molecular patterning, and grayscale-mediated mechanical patterning are explored to programmably create intricate, biomimetic, and concentration-controlled architectures. We demonstrate the Bioxolographic printing of various cell types, showing excellent cell viability, compatibility with long-term culture, and ability for nascent protein deposition. These results position Bioxolography as a transformative platform for rapid, scalable, high-resolution fabrication of functional living materials with encoded chemical and mechanical properties.