Real-time detection of plasmin activity in milk using an ordered porous layer interferometry

In this study, a novel sensing system based on ordered porous layer interferometry (OPLI) was developed for real-time and label-free detection of plasmin activity in milk. A fibrin-functionalized silica colloidal crystalline film was used as interference sensing layers for plasmin recognition. Plasmin-triggered fibrinolysis within the interference sensing layer caused a shift in the interference fringes, represented as an optical thickness change (ΔOT) that could be tracked in real time. The OPLI system showed no response to milk matrix or other proteases (final ΔOT about 0 nm), effectively overcoming the interference from the complex milk matrix. The detection limit for plasmin was 0.5 μg/mL, with a wide linear range of about 0.5-100 μg/mL. Moreover, plasmin activity in raw milk, pasteurized milk and UHT milk was successfully detected without sample pretreatment. The results were validated by a concurrent enzyme-linked immunosorbent assay (ELISA).