Breaking Through Bead-Supported Assay: Integration of Optical Tweezers Assisted Fluorescence Imaging and Luminescence Confined Upconversion Nanoparticles Triggered Luminescent Resonance Energy Transfer (LRET)

Abstract

Herein, a conceptual approach for significantly enhancing a bead-supported assay is proposed. For the fluorescence imaging technology, optical tweezers are introduced to overcome the fluid viscosity interference and immobilize a single tested bead at the laser focus to guarantee a fairly precise imaging condition. For the selection of fluorescent materials and the signal acquisition means, a type of innovative luminescence confined upconversion nanoparticle with a unique sandwich structure is specially designed to act as an efficient energy donor to trigger the luminescent resonance energy transfer (LRET) process. By further combining the double breakthrough with a molecular beacon model, the newly developed detection strategy allows for achieving a pretty high LRET ratio (≈ 88%) to FAM molecules and offering sound assay performance toward miRNA analysis with a detection limit as low as the sub-fM level, and is capable of well identifying single-base mismatching. Besides, this approach not only is able to accurately qualify the low-abundance targets from as few as 30 cancer cells but also can be employed as a valid cancer early warning tool for performing liquid biopsy.

Biomimetic Chip Enhanced Time-Gated Luminescent CRISPR-Cas12a Biosensors under Functional DNA Regulation

“…Like the CEA sensing, the above flexible transduction pattern is ascertained successively by the visible discrepancies that emerge in electrophoresis image (Figure 4B) and luminescence spectra (Figure 4C). Following the same verification process for the analysis performance, the target concentration dependent luminescence recovery manner (Figure 4D) can be also attributed to a wide linear relationship (R 2 : 0.9973, Figure 4E) ranging from 80 nM to 5000 μM, and the LOD is estimated to be as low as 63 nM (which is slightly higher than our previous work 21 due to the different signal collectors), exhibiting an apparent sensitivity advantage over the frequently used Na + meter (LOD ∼ 10 μM) and the recently reported optical biosensors (Table S2). More than that, profiting from the specific identification of the metal ions driven DNAzyme, the analytical methodology can hardly respond to other nonspecific metal cations from monovalent to trivalent or a representative nonmetallic cation as well as some biomacromolecules (Figure 4F).…”

“…For another, the acquisition of optical signals for these luminescent biosensors is mostly focused on the commonly applied Stokes emission by selecting a source within visible light, forcing a poor capacity to avoid the interference of background signals in complicated biological media such as human plasma. , Although our previous work has employed a kind of anti-Stokes luminescence mode (upconversion luminescence) to partly address this concern, the complex synthesis process (a high-temperature thermal decomposition based seed-mediated growth) of the upconversion nanoparticle and the harsh nonlinear multiphoton absorption behavior under a very high photodensity condition compel it to be short of user friendliness. Inspiringly, another option, which is the long-persistent nanophosphor (LPN), can perfectly conquer these problems.…”

Nanotechnology in emerging liquid biopsy applications

“…Luminescent upconverting nanoparticles were used in combination with luminescence energy transfer by a molecular beacon, which was hindered after miRNA hybridization. This system offered an LOD 762 aM and an analysis time of 10 min [209]. The excellent catalytic properties of ZnO nanostars were also exploited along with a luminol-O 2 system for an ultrasensitive electrochemiluminescent detection of micro-RNA (LOD 18.6 aM) [210].…”

Spatially Selective Imaging of Mitochondrial MicroRNAs via Optically Programmable Strand Displacement Reactions

“…Notably, UCNPs have been widely used in the design of FRET-based sensors for fluorescent imaging of RNA, in which UCNPs act as the energy donors and dye-or quencher-labeled DNA are used as the energy acceptors. [18] Recently, we and others have explored using UCNPs as light transducers to achieve NIR light-activated RNA imaging in living cells. [11b,c, 19] However, engineering of biosensors for spatially-selective imaging of mitomiRs has not yet been achieved yet.…”