Color-Changing Hydrogels for Visual pH Monitoring of Confined Living Systems

Maintaining pH homeostasis is essential for cell viability, yet confined culture systems such as bioreactors are susceptible to pH drift, even under controlled nutrient supply. We developed a noninvasive method for continuous pH monitoring of microbial cultures using dual-dye colorimetric sensors covalently bound to pH-responsive hydrogels. A 3D-printed dual-chamber device spatially separates the sensors from active cultures, allowing only proton and metabolite diffusion across the semipermeable membrane. This eliminates the risk of sensor obscuration from cell turbidity. Quantitative pH measurements were obtained through image-based color analysis and numerical processing of the hydrogel response, resolving pH within ±0.1 units of those performed using conventional sensors. Our material strategy offers an alternative approach for in situ monitoring of confined culture environments, enabling live tracking of pH dynamics in biological systems.