A 3D-Printed Scaffolded Hydrogel Microneedle Array Biosensor for Real-Time, Continuous Monitoring.

Hydrogel-based biosensors offer a promising platform for designing microneedles capable of continuously tracking biomarkers in real time. However, such biosensors have been limited by the mechanical properties of hydrated hydrogels, which are generally ineffective at penetrating the skin to access interstitial fluid (ISF). As a solution, we have developed a microneedle-array biosensor (MAB) patch that enables continuous, reversible sensing by coupling fluorescent deoxyribonucleic acid (DNA) aptamer switches to a hydrated hydrogel mesh within a 3D-printed scaffold. This scaffold provides essential mechanical support for skin insertion while preserving the apatmer-hydrogel's sensing functionality in the ISF. We demonstrate this design by tuning both aptamer switch design and hydrogel mesh size to detect exogenous levels of stress hormone cortisol and the metabolite adenosine triphosphate. We subsequently incorporated our cortisol-sensing hydrogel into the MAB scaffold and coupled this system to a custom-designed portable optical detector. Following in vitro validation, we demonstrated the biocompatibility and in vivo utility of our system by conducting continuous, real-time measurements of exogenous cortisol in the ISF of live rats. These results demonstrate, for the first time, submicromolar detection using a sensor-embedded hydrogel microneedle system, highlighting the MAB platform as a versatile solution for real-time, continuous in vivo biosensing.