Rational Design of “Three-in-One” Ratiometric Nanoprobes: Protein-Caged Dityrosine, CdS Quantum Dots, and Gold Nanoclusters

Recently, multiplexed ratiometric fluorescence sensors for detecting several analytes have received much interest because of their multifunctionality. Here, we fabricate a novel trinity fluorescent nanoprobe in which one small-molecule fluorophore, blue-emissive dityrosine (diTyr) residues, and two nanomaterial fluorophores, green-emissive CdS quantum dots (CdSQDs) and red-emissive gold nanoclusters (AuNCs), are cocaged in a bovine serum albumin (BSA) molecule. The large differences of Stokes shifts among diTyr residues, CdSQDs, and AuNCs ensure their emission at a single excitation wavelength. The nanoprobes can be facilely integrated using two-step synthetic reactions. DiTyr residues and AuNCs are formed and bound to the protein cage through the redox reaction between Au3+ and tyrosine residues of BSA, and the CdSQDs are followed to be conjugated to the modified BSA cage-templated CdS combination reaction. With established benign biocompatibility, the nanoprobes can ratiometrically detect intracellular glutathione by significantly enhancing the green emission of the conjugated CdSQDs. Likewise, the ratiometric sensing of solution alkalinity and tris(2-carboxyethyl)phosphine can be achieved using blue-emitted diTyr residues and red-emitted AuNCs as the responsive units, respectively, and the corresponding other two fluorophores as the reference signals. This study addresses a concept of trinity fluorescence ratiometric sensing system with multiple targets and optional references, which should be a promising pathway to meet the challenges from complexing biochemical environments and multivariate analysis.