Using thiol–ene click chemistry to engineer 3D printed plasmonic hydrogel scaffolds for SERS biosensing

3D cell culture models allow the study of the biomolecular processes underlying pathophysiological conditions by mimicking tissues and organs. Despite significant progress in creating such 3D architectures, studying cell behaviour in these systems still poses some challenges due to their heterogeneity and complex geometry. In this context, surface-enhanced Raman spectroscopy (SERS) can be implemented for molecular detection in biological settings with high sensitivity. The incorporation of SERS sensors in 3D models can thus lead to powerful platforms to study cellular response to therapeutics, metabolic pathways, signaling, and cell-cell communication events. Here, we introduce a library of plasmonic hydrogels that can be orthogonally photo-crosslinked <i>via</i> thiol-ene click chemistry and identify the main physicochemical factors accounting for their SERS performance. Using hydrogel-forming polymers such as gelatin, alginate, and carboxymethylcellulose modified with complementary thiol and norbornene groups, we created hydrogels with tailored chemical backbones. We identified swelling, porosity, and chemical composition as crucial factors determining their potential to detect different molecules by SERS. We additionally assessed their biocompatibility and printability, to ensure that these hydrogels meet the requirements for their use as 3D cellular scaffolds, showing their potential for real-time and <i>in situ</i> detection of biorelevant metabolites.