A Diffusion-Driven Chromogenic Reagent-Free Viscosity-Based Sensor with Dual Signals for the Visual Detection of Hyaluronidase.

Point-of-care testing (POCT) requires simple and cost-effective strategies to enhance reliability and practicality. Here, a state-of-the-art, dual-mode viscosity-based sensor (DM vis-sensor) is developed by leveraging the diffusion-driven viscosity differentiation on the mesoporous membrane. The rapid and visual detection of hyaluronidase (HAase) is demonstrated as a model. The sensor exploits the enzymatic cleavage of hyaluronic acid (HA) by HAase, which reduces solution viscosity, thereby altering the solution diffusion pattern on the mesoporous membrane. High-viscosity HA solutions form small and dark diffusion spots with short diameters and low grayscale ratio (GR) values, while HAase-treated HA solutions yield large and light diffusion spots with long diameters and high GR values. This dual-mode sensor, which combines distance and color readouts, enables intuitive and instrument-free detection of HAase without the need of chromogenic agents. It demonstrates high sensitivity to HAase, with detection limits of 0.12 U/mL (distance-based readout) and 0.16 U/mL (grayscale-based readout). Additionally, it is successfully validated through the quantification of HAase in human urine samples. Notably, its simplicity, low cost, and dual-readout reliability make it a promising POCT tool for early diagnosis and monitoring, especially in resource-limited settings. Therefore, this work highlights the potential of the development of dual-mode viscosity-based sensors for POCT applications.