Bioluminescent Peptide-Based Biosensors for Early Enamel Demineralization: An In Vitro Proof-of-Concept

BackgroundEarly enamel demineralization corresponding to ICDAS 0-1 is difficult to detect through routine visual-tactile examination, as initial mineral loss often precedes visible surface change. Existing optical adjuncts improve detection but frequently require specialized equipment, high costs, or ionizing radiation, limiting widespread clinical use. ObjectiveTo develop a low-cost, luciferin-inspired fluorogenic peptide biosensor capable of selectively binding early enamel porosity and producing a quantifiable green luminescent signal under standard dental blue-light activation. MethodsA calcium-affinitive peptide (P-Ca) was synthesized and functionalized with an inexpensive fluorogenic ester-quencher pair that becomes fluorescent upon conformational stabilization on partially demineralized enamel. Thirty extracted molars, collected as anonymized biowaste from orthodontic procedures, were sectioned and assigned to sound enamel, mild demineralization (pH-cycling, 48 h), or moderate demineralization (96 h). After 60 s incubation with P-Ca, specimens were illuminated using a dental curing light (450-470 nm). Emission spectra ({lambda}_max 515 {+/-} 5 nm) and fluorescence intensities were quantified and compared with quantitative light-induced fluorescence (QLF). Cytocompatibility was evaluated using an immortalized human gingival fibroblasts cell line (HGF-1). ResultsFluorescence intensity increased in accordance with demineralization severity (p < 0.001), and luminescent output strongly correlated with QLF {Delta}F values (p < 0.001). HGF-1 viability remained above 95% after 24 h exposure. ConclusionThis in vitro study supports the feasibility of a fluorogenic peptide biosensor as an inexpensive, non-radiographic adjunct for early enamel demineralization detection, with clear potential for future chairside translation.