Hybrid Oxygen-Sensing Bio-Scaffolds for 3D Micro-Tissue Models

Culturing cells and micro-tissue samples in 3D bio-scaffolding structures is gaining popularity; however, precise control of tissue micro-environment in such systems remains challenging. We describe a family of new hybrid bio-scaffolds with 3D O₂-sensing ability, produced by simple means from readily available bio-scaffolding and O₂-sensing materials. Three different types of phosphorescent O₂-sensing materials-polymeric microparticles (MPs), supramolecular probe MitoXpress and nanoparticulate probes NanO2 and Nano-IR (NPs)-were integrated in Matrigel and agarose scaffolding materials and evaluated. Key working characteristics of such hybrid scaffolds, including heterogeneity, stability, cytotoxicity, optical signals and O₂-sensing properties, ease of fabrication and use, were compared. The results show superiority of the Matrigel hybrids with NanO2 and Nano-IR probes. Demonstration experiments were conducted with HCT116 cells and individual spheroids derived from these cells, culturing them in the Matrigel-NP hybrid scaffolds and monitoring oxygenation and local O₂ gradients on a time-resolved fluorescence plate reader and by phosphorescence lifetime imaging microscopy (PLIM).