A protein structure-dependent fluorescent probe for real-time β-lactoglobulin detection

Real-time detection of β-lactoglobulin, a prominent bioactive protein in milk-based beverages, enables the acceleration of analytical workflows, thereby facilitating immediate on-site beverage quality analysis. Conventional analytical methods often require specialized equipment or prolonged times, limiting their utility for instant on-site applications. Herein, targeting the unique central calyx binding site within β-lactoglobulin, a novel protein structure-dependent fluorescent probe (DNP) was designed for the protein detection. Without requiring enzymatic reactions, DNP demonstrated a highly selective and sensitive fluorescence "turn-on" responsiveness to β-lactoglobulin through direct non-covalent interactions. This characteristic enabled the visualization of exogenous β-lactoglobulin in living cells. Additionally, DNP enabled fluorescence-based real-time preliminary assessments of β-lactoglobulin content in beverage samples. It also enabled quantitative detection of β-lactoglobulin, accounting for sample matrix effects. These characteristics render DNP a promising tool for real-time β-lactoglobulin detection, offering the potential to immediate analysis in beverage analysis.