NIR-to-Vis Handheld Platforms for Detecting miRNA Level and Mutation Based on Sub-10 nm Sulfide Nanodots and HCR Amplification

Abstract

Sub-10 nm monodisperse alkaline-earth sulfide nanodots (ASNDs) with bright near-infrared (NIR)-excitation fluorescence and adjustable emission wavelength were prepared by a thermal decomposition method for the first time. The ASNDs exhibited high NIR-to-vis conversion efficiency and served as multicolor fluorescent labels in the proposed miR-224 assay. Targeted detection of the miR-224 level and single-nucleotide variation in miR-224 was carried out on a smartphone-based platform using a hybridization chain reaction (HCR) amplification strategy. In the presence of miR-224, the ASND-labeled HCR probes self-assembled on the surface of the diagnosis kits, generating strong fluorescent signals linearly proportional to miR-224 contents in the range of 10− 2000 fM. Significantly, mutations in miR-224 led to the variation in the fluorescence intensity ratio in RGB channels. Simultaneously, evident changes of fluorescent brightness and color were easily visualized by the naked eye, which enabled on-site discrimination of miR-224 with different mutant loci. This work provides a novel preparation approach for ultrasmall NIR excitation sulfide nanodots and reveals the potential of the as-synthesized ASNDs in point-of-care (POC) nucleic acid testing. Further, it may provide a handheld platform for miRNA single-nucleotide polymorphism analysis.

Recent advances in enzyme-free and enzyme-mediated single-nucleotide variation assay in vitro

“…Reprinted with permission from ref. [ 43 ]. (c) Schematic diagram of Cas9-immobilized gFET for SNP detection and typical results.…”

“…Thus, the occurrence (or not) of the ligation reaction can be served as an indicator to determine the presence of SNV [ 42 ]. Usually, ligation-based SNV detection has been successfully operated with various amplification strategies, such as HCR [ 43 ], PCR [ 44 ], and RCA [ 45 ], to obtain amplified ligation products and signals. HCR is an isothermal amplification method, in which two hairpins cross-open each other with the assistance of a trigger, thus producing a long and nicked double-helix chain and enabling eminent amplification efficiency [ 46 ].…”

Endogenous mRNA-Powered and Spatial Confinement-Derived DNA Nanomachines for Ultrarapid and Sensitive Imaging of Let-7a

“…However, the reliance on external additions of fuel DNA strands or protein enzymes limits their applicability in vivo. In contrast, the field of DNA nanotechnology provides promising approaches, particularly through nonenzymatic signal amplification methods. , DNA amplifiers based on a hybridization chain reaction (HCR), − toehold strand displacement reaction (TSDR), , and catalytic hairpin assembly (CHA) − have emerged as strong contenders, leveraging their nonenzymatic and isothermal characteristics for amplified detection of intracellular miRNA. Despite their potential, most of these DNA amplifiers face challenges such as inefficient penetration of the cell membrane, susceptibility to intracellular nucleases, and slow reaction dynamics, which limit their usefulness in imaging low-abundance miRNAs. , …”

Double Signal Amplification Strategy for Dual-Analyte Fluorescent Aptasensors for Visualizing Cancer Biomarker Proteins

“…Trichromatic fluorescent ASNDs, namely, red-emitting ASNDs (rASNDs), green-emitting ASNDs (gASNDs), and blue-emitting ASNDs (bASNDs), were obtained through the modified thermal decomposition method (Figure S1a). 21 Further modification and encapsulation with 1-octadecanethiol (ODT) and distearoyl phosphoethanolamine-polyethylene glycol (NH 2 -DSPE-PEG) made ASNDs with good monodispersity and colloidal stability (Figure 1a). The lipidencapsulated rASNDs and bASNDs (i.e., Lip@rASNDs and Lip@bASNDs) were subsequently used as fluorescent labels to construct a ratiometric fluorescent sensor for VEGF and sIL-6R assays.…”