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Decrypting the dynamics of receptor dimerization on cell membranes bears great importance in identifying the mechanisms regulating diverse cellular activities. In this regard, long-term monitoring of single-molecule behavior during receptor dimerization allows deepening insight into the dimerization process and tracking of the behavior of individual receptors, yet this remains to be realized. Herein, real-time observation of the receptor tyrosine kinases family (RTKs) at single-molecule level based on plasmon rulers was achieved for the first time, which enabled precise regulation and dynamic monitoring of the dimerization process by DNA programming with excellent photostability. Additionally, those nanoprobes demonstrated substantial application in the regulation of RTKs protein dimerization/phosphorylation and activation of downstream signaling pathways. The proposed nanoprobes hold considerable potential for elucidating the molecular mechanisms of single-receptor dimerization as well as the conformational transitions upon dimerization, providing a new paradigm for the precise manipulation and monitoring of specific single-receptor crosslink events in biological systems.

Achieving accurate measurements of G-quadruplexes (G4s), especially the characterization of their complicated non-covalent interactions with various components (such as metal ions and ligands) under physiological conditions, is of fundamental significance in unveiling their biological roles and developing antitumor drugs. By employing a nanopore ion emitter (∼30 nm), we demonstrated for the first time that G4 ions, which are free of non-specific adduction and meanwhile maintaining their pre-existing specific bindings with metal ions or ligands, can be directly liberated from common biochemical buffers (consisting of concentrated non-volatile salts of150 mM) for mass spectrometric examination. Notably, the intermediate complexes of G4s with mixed di-cation coordination formed during the Na

Methicillin-resistant

The use of functional DNA nanostructures as carriers to ship proteins through solid-state nanopores has recently seen substantial growth in single-protein-molecule detection (SPMD), driven by the potential of this methodology and implementations that it may enable. Ultrasmall nanopores have exhibited obvious advantages in spatiotemporal biological detection due to the appropriate nanoconfined spaces and unique properties. Herein, a 6.8 nm DNA tetrahedron (TDN) with a target-specific DNA aptamer (TDN-apt) was engineered to carry the representative target of acetylcholinesterase (AChE) through an ultrasmall nanopipet with a 30 nm orifice, underpinning the advanced SPMD of AChE with good performance in terms of high selectivity, low detection limit (0.1 fM), and especially superior signal-to-noise ratio (SNR). The kinetic interaction between TDN-apt and AChE was studied and the practical applicability of the as-developed SPMD toward real samples was validated using serum samples from patients with Alzheimer's disease. This work not only presented a feasible SPMD solution toward low-abundance proteins in complex samples and but also was envisioned to inspire more interest in the design and implementation of synergized DNA nanostructure-ultrasmall nanopore systems for future SPMD development.

Energy carrier evolution is crucial for material performance. Ultrafast microscopy has been widely applied to visualize the spatiotemporal evolution of energy carriers. However, direct imaging of small amounts of energy carriers on nanoscale remains difficult due to extremely weak transient signals. Here we present a method for ultrasensitive and high-throughput imaging of energy carrier evolution in space and time. This method combines femtosecond pump-probe techniques with interferometric scattering microscopy (iSCAT), named Femto-iSCAT. The interferometric principle and unique spatially-modulated contrast enhancement increase the transient image contrast by >2 orders of magnitude and enable the exploration of new science. We address three important and challenging problems: transport of different energy carriers at various interfaces, heterogeneous hot electron distribution and relaxation in single plasmonic resonators, and distinct structure-dependent edge state dynamics of carriers and excitons in optoelectronic semiconductors. Femto-iSCAT holds great potential as a universal tool for ultrasensitive imaging of energy carrier evolution in space and time.

The development of uranyl ion detection technology has exhibited its significance in public security and environmental fields for the radioactivity and chemical toxicity of uranyl ion. The WHO standard of uranyl ion makes it necessary to develop highly sensitive uranyl rapid warning system in drinking water monitoring. Herein, a visualized rapid warning system for trace uranyl ion is carried out based on electrochemiluminescence (ECL) imaging technology to give an ultra-low limit of detection (LOD) and high selectivity. Amidoxime, a bi-functional group with both uranyl ion capturing and co-reactive functions, is modified on a conjugated polymer backbone with strong ECL signal to be prepared into three-in-one polymer nanoparticles (PNPs) with self-enhanced ECL property. The captured uranyl ion can enhance the ECL signal of PNPs via resonance energy transfer process to give the LOD as 0.5 ng/L, which is much lower than the known luminescent uranyl sensors. Furthermore, ECL imaging technology is introduced into realizing visualized uranyl rapid warning, and can be successfully applied on natural water samples. This study provides a novel strategy for uranyl rapid warning, and shows its potential meaning in public security and environmental fields.

Myocardial miRNAs in peripheral blood are closely related to the pathogenic process of myocardial infarction. Rapid identification and accurate quantification of myocardial miRNAs are of great significance to clinical interventions for treating cardiovascular lesions. Therefore, a ratiometric electrochemiluminescence (ECL) biosensor integrating DNAzyme with a resonance energy transfer (RET) system was designed to detect myocardial miRNA. The dual-signal system was composed of rA marked substrate strand functionalized CdTe quantum dots (QDs) as reductive-oxidative (R-O) emitters and Cy5-labeled strand-functionalized Ru(bpy)

DNA methylation is catalyzed by a family of DNA methyltransferases that play crucial roles in various biological processes. Therefore, an ultrasensitive methyltransferase assay is highly desirable in biomedical research and clinical diagnosis. However, conventional assays for the detection of DNA methyltransferase activity often involve radioactive labeling, costly equipment, and laborious operation. In this study, an ultrasensitive and label-free method for detecting DNA adenine methyltransferase (Dam) and CpG methyltransferase (M.SssI) was developed using the nanopore technique coupled with DNA cascade signal amplification reactions. A hairpin DNA (H

A unique poly(amido amine) (PAMAM)/polymer dots (Pdots)/NiO heterostructure is developed for photocathodic detection of tyrosinase (TYR), which is applicable to TYR detection in actual human serum samples.

Understanding the pH evolution during endocytosis is essential for our comprehension of the fundamental processes of biology as well as effective nanotherapeutic design. Herein, we constructed a plasmonic Au@PANI core-shell nanoprobe, which possessed significantly different scattering properties under acidic and basic conditions. Encouragingly, the scattering signal of Au@PANI nanoprobes displayed a positive linear correlation with the pH value not only in PBS but also in nigericin-treated cells. Ultimately, benefiting from the excellent response properties as well as the excellent biocompatibility and stability, the Au@PANI nanoprobes have successfully enabled a dynamic assessment of the evolving pH in the endosomal package as the endosome matured from early to late, and eventually to the lysosome, by reporting scattering signal changes.

By assembling nanotweezers with ATP-splitting aptamers on gold nanorods (AuT123L), we constructed a near-infrared-activated ATP sensing device that could time-controllably image ATP levels in living cells. By replacing the aptamers on the nanotweezers, the nanoplatform can be applied to other important biomolecules, opening up more possibilities for the study of time controllable nanodevices.

A high-performance organic photoelectrochemical transistor (OPECT) is developed by synergizing a light-sensitive aptamer/CdS QDs/Ti wire gate photoanode with a PEDOT:PSS-based device towards detection of the tear biomarker lysozyme.

Single-cell protein therapeutics is expected to promote our in-depth understanding of how does a specific protein with therapeutic dosing treat the cell without population averaging. However, it has not yet been tackled by current single-cell nanotools. We address this challenge by utilization of a double-barrel nanopipette, in which one lumen was used for electroosmotic cytosolic protein delivery and the other was customized for ionic evaluation of the consequence. Upon injection of protein DJ-1 via delivery lumen, upregulation of the antioxidant protein could protect neural PC-12 cells against oxidative stress from phorbol myristate acetate exposure, which could be transduced by targeting the cytosolic hydrogen peroxide by the detecting lumen. This work has devised a nanotool for single-cell protein therapeutics and provided a perspective for futuristic single-cell therapeutics involving different therapeutic modalities e.g. peptides, enzymes and nucleic acids.

Accurately obtaining information on the heterogeneity of CTCs at the single-cell level is a very challenging task that may facilitate cancer pathogenesis research and personalized therapy. However, commonly used multicellular population capture and release assays tend to lose effective information on heterogeneity and cannot accurately assess molecular-level studies and drug resistance assessment of CTCs in different stages of tumor metastasis. Herein, we designed a near-infrared (NIR) light-responsive microfluidic chip for biocompatible single-cell manipulation and study the heterogeneity of CTCs by a combination of the lateral flow microarray (LFM) chip and photothermal response system. First, immunomagnetic labeling and a gradient magnetic field were combined to distribute CTCs in different regions of the chip according to the content of surface markers. Subsequently, the LFM chip achieves high single-cell capture efficiency and purity (even as low as 5 CTCs per milliliter of blood) under the influence of lateral fluid and magnetic fields. Due to the rapid dissolution of the gelatin capture structure at 37 °C and the photothermal properties of gold nanorods, the captured single CTC cell can be recovered in large quantities at physiological temperature or released individually at a specific point by NIR. The multifunctional NIR-responsive LFM chip demonstrates excellent performance in capture and site release of CTCs with high viability, which provides a robust and versatile means for CTCs heterogeneity study at the single-cell level.

Cuproptosis is a new form of programmed cell death and exhibits enormous potential in cancer treatment. However, reducing the undesirable Cu ion release in normal tissue and maximizing the Cu-induced therapeutic effect in cancer sites are two main challenges. Herein, we constructed a photothermally triggered nanoplatform (Au@MSN-Cu/PEG/DSF) to realize the on-demand delivery for synergistic therapy. The released DSF chelated with Cu2+ in situ to generate the high cytotoxic bis(diethyldithiocarbamate)-copper (CuET) causing cell apoptosis and formed Cu+ species promoting toxic mitochondrial protein aggregation, leading to cell cuproptosis. Synergistic with photothermal therapy, the Au@MSN-Cu/PEG/DSF could effectively kill the tumor cell and inhibit tumor growth (inhibition rate up to 80.1%). These results provide a promising perspective for potential cancer treatment based on cuproptosis, and also inspire the design of advanced nano-therapeutic platforms.

Starvation therapy kills tumor cells via consuming glucose to cut off their energy supply. However, since glucose oxidase (GOx)-mediated glycolysis is oxygen-dependent, the cascade reaction based on GOx faces the challenge of a hypoxic tumor microenvironment. By decomposition of glycolysis production of H

Lysosomal acidification is essential for its degradative function, and the flux of H+ correlated with that of K+ in lysosomes. However, there is little research on their correlation due to the lack of probes that can simultaneously image these two ions. To deeply understand the role of K+ in lysosomal acidification, here, we designed and fabricated a nanodevice using a K+-aptamer and two pH-triggered nanoswitches incorporated into a DNA triangular prism (DTP) as a dual signal response platform to simultaneously visualize K+ and pH in lysosomes by a fluorescence method. This strategy could conveniently integrate two signal recognition modules into one probe, so as to achieve the goal of sensitive detection of two kinds of signals in the same time and space, which is suitable for the detection of various signals with the correlation of concentration. By co-imaging both K+ and H+ in lysosomes, we found that the efflux of K+ was accompanied by a decrease of pH, which is of great value in understanding lysosomal acidification. Moreover, this strategy also has broad prospects as a versatile optical sensing platform for multiplexed analysis of other biomolecules in living cells.

Electrogenerated chemiluminescence microscopy (ECLM) provides a real-time imaging approach to visualize the surface-dependent catalytic activity of nanocatalysts, which helps to rationalize the design of catalysts. In this study, we first propose super-resolution ECLM that could measure the facet- and site-specific activities of a single nanoparticle with nanometer resolution. The stochastic nature of the ECL emission makes the generation of photons obey Poisson statistics, which fits the requirement of super-resolution radial fluctuation (SRRF). By processing an SRRF algorithm, the spatial resolution of ECL images achieved ca. 100 nm, providing more abundant details on electrocatalytic reactivities at the subparticle level. Beyond conventional wide-field ECL imaging, super-resolution ECLM provided the spatial distribution of catalytic activities at a Au nanorod and nanoplate with scales of a few hundred nanometers. It helped uncover the facet- and defect-dependent surface activity, as well as the dynamic fluctuation of reactivity patterns on single nanoparticles. The super-resolution ECLM provides high spatiotemporal resolution, which shows great potential in the field of catalysis, biological imaging, and single-entity analysis.

Intracellular potassium ions (K+) play pivotal roles in many physiological processes. Several K+ sensors have been developed for probing cellular K+ fluctuations. Nevertheless, the existing solutio...

Over the past 2 years, many DNA motors have been synthesized and run in living cells, but there are still challenges in designing integrated DNA motors self-powered to enable autonomous intracellul...

Rational utilization of the rich light-bio-matter interplay taking place in single-cell analysis represents a new technological direction in the field. The light-fueled operation is expected to achieve advanced photoelectrochemical (PEC) single-cell analysis with unknown possibilities. Here, a PEC nanoreactor capable of single-cell sampling and near zero-background Faradaic detection of intracellular microRNA (miR) is devised by the construction of a small reaction chamber accommodating the target-triggered hybridization chain reaction for binding the metallointercalator of [Ru(bpy)

We report the mass spectrometric identification of short-lived intermediates generated under alternating-current electrolysis, via combination of bipolar electrochemistry with nanoelectrospray ionization in a hybrid ultramicroelectrode/ion emitter.

A novel mass spectrometric method for probing the flash chemistry of electrogenerated reactive intermediates was developed based on rapid collision mixing of electrosprayed microdroplets by using a theta-glass capillary. The two individual microchannels of the theta-glass capillary are asymmetrically or symmetrically fabricated with a carbon bipolar electrode to produce intermediates in situ . Microdroplets containing the newly formed intermediates collide with those of the invoked reactants at sub-10 microsecond level, making it a powerful tool for exploring their ultrafast initial transformations. As a proof-of-concept, we present the identification of the key radical cation intermediate in the oxidative dimerization of 8-methyl-1,2,3,4-tetrahydroquinoline and also the first revealment of previously hidden nitrenium ion involved reaction pathway in the C-H/N-H cross-coupling between N,N'-dimethylaniline and phenothiazine.

Aggregation-induced emission (AIE) has attracted considerable attention in designing high-performance emitters for both photoluminescence (PL) and electrochemiluminescence (ECL). In this study, to prepare AIE-active polymer dots (Pdots) with efficient PL and ECL emission, we demonstrate a molecular engineering strategy by tuning the side chain of fluorenyl–tetraphenylethylene polymers from ester to carboxyl groups. By introducing the carboxyl group at a moderate fraction, significant PL enhancement was achieved, benefiting from the more quantities and the better compactness of the luminophores inside the Pdots. The ECL emission of these carboxylic Pdots has also been promoted to a highest quantum yield (ΦECL) of 54.7%, making it a promising ECL emitter for visual detection for bioanalysis. Unlike the traditional oxidative–reductive mechanism, the ECL of carboxylic Pdots exhibited a unique low-oxidation-potential (LOP) route that originated from the enhanced proton-coupled electron transfer (PCET) within the carboxylic nanoparticles.

Screening appendants on membrane proteins to understand their varied regulation effects is desirable for finding the potential candidates of the membrane-protein-targeted drugs. However, most artif...