Real-time sensing of food spoilage using a recombinant mVenus–Tolles FRET pH biosensor

Abstract Monitoring food spoilage is essential for enhancing food safety and reducing waste. pH changes serve as a valuable indicator of microbial activity, and real-time pH monitoring can provide an accurate and non-invasive indication of food spoilage. The pHlameleon chimera proteins were developed for pH sensing by Förster resonance energy transfer (FRET) and were extensively used in various biomedical applications. Herein, we evaluate the mVenus–Tolles as a FRET-based biosensor for detecting pH changes as a proxy for food spoilage. The protein was fused to an N-terminal vesicle-nucleating peptide (VNP) tag and recombinantly expressed and purified to homogeneity. Experimental validation demonstrated pH-responsive FRET signal in an array of buffers as well as in a complex food matrix such as chickpea paste, correlating with increasing acidity and microbial growth in food. These findings suggest that this protein-based FRET biosensor holds promise for safe integration into food or packaging for real-time freshness monitoring.