Owing to the unique optophysical properties of nanomaterials and their self-calibration characteristics, nanomaterial-based (e.g., polymer dots (Pdots) quantum dots (QDs), silicon nanorods (SiNRs), and gold nanoparticle (AuNPs), etc.) ratiometric fluorescent sensors play an essential role in numerous biosensing and cell imaging applications. The dual-emission ratiometric fluorescence technique has the function of effective internal referencing, thereby avoiding the influence of various analyte-independent confounding factors. The sensitivity and precision of the detection can therefore be greatly improved. In this review, the recent progress in nanomaterial-based dual-emission ratiometric fluorescent biosensors is systematically summarized. First, we introduce two general design approaches for dual-emission ratiometric fluorescent sensors, involving ratiometric fluorescence with changes of one response signal and two reversible signals. Then, some recent typical examples of nanomaterial-based dual-emission ratiometric fluorescent biosensors are illustrated in detail. Finally, probable challenges and future outlooks for dual-emission ratiometric fluorescent nanosensors for biosensing and cell imaging are rationally discussed.

With the development of nanotechnology, fluorescent silicon nanomaterials have been synthesized and applied in various areas. Among them, silicon quantum dots (SiQDs) are a new class of zero-dimensional nanomaterials with outstanding optical properties, benign biocompatibility, and ultra-small size. In recent years, SiQDs have been gradually utilized for constructing high-performance fluorescent sensors for chemical or biological analytes. Herein, we focus on reviewing recent advances in SiQD-based fluorescent biosensors from a broad perspective and discussing possible future trends. First, the representative progress for synthesizing water-soluble SiQDs in the past decade is systematically summarized. Then, the latest achievement of the design and fabrication of SiQD-based fluorescent biosensors is introduced, with a particular focus on analyte-induced photoluminescence (fluorescence) changes, hybrids of SiQDs with other materials or molecules, and biological ligand-modification methods. Finally, the current challenges and prospects of this field are highlighted.

A ratiometric DNA sensor was developed based on fluorescent silicon nanodots (SiNDs) and Ru(bpy)₂(dppx)²⁺. The absorption spectrum of Ru(bpy)₂(dppx)²⁺ has significant overlap with both the excitation and emission spectra of SiNDs. Therefore, fluorescence quenching of Ru(bpy)₂(dppx)²⁺ toward SiNDs can occur on account of the strong inner filter effect. The effect of quenching is not influenced by the specific binding between Ru(bpy)₂(dppx)²⁺ and DNA. Fluorescence turn-on detection of DNA can be performed employing Ru(bpy)₂(dppx)²⁺ and SiNDs as the response and reference signals, respectively. Using SiND–Ru(bpy)₂(dppx)²⁺, a convenient, sensitive, rapid, and precise method could be developed for DNA detection. In aqueous solutions, the I₆₀₁/I₄₄₈ fluorescence intensity ratio of SiND–Ru(bpy)₂(dppx)²⁺ increases linearly in the DNA concentration range of 20–1500 nM. The limit of detection and precision of the method is 4.3 nM and 3.5% (50 nM, n = 13), respectively. The ratiometric sensor was tested for visual detection of trace DNA. Moreover, this method was found suitable for the ratiometric detection of DNA in a simulated sample and a human serum sample, and the recoveries were in the range of 98–119%.