Unveiling the Dynamic Electrocatalytic Activity of Online Synthesized Bimetallic Nanocatalysts via Electrochemiluminescence Microscopy.

Effective bimetallic nanoelectrocatalysis demands precise control of composition, structure, and understanding catalytic mechanisms. To address these challenges, we employ a two-in-one approach, integrating online synthesis with real-time imaging of bimetallic Au@Metal core-shell nanoparticles (Au@M NPs) via electrochemiluminescence microscopy (ECLM). Within 120 s, online electrodeposition and in situ catalytic activity screening alternate. ECLM captures transient faradaic processes during potential switches, visualizes electrochemical processes in real-time, and tracks catalytic activity dynamics at the single-particle level. Analysis using ECL photon flux density eliminates size effects and yields quantitative electrocatalytic activity results. Notably, a nonlinear activity trend corresponding to the shell metal to Au surface atomic ratio is discerned, quantifying the optimal surface component ratio of Au@M NPs. This approach offers a comprehensive understanding of catalytic behavior during the deposition process with high spatiotemporal resolution, which is crucial for tailoring efficient bimetallic nanocatalysts for diverse applications.

Endogenous AND Logic DNA Nanomachine for Highly Specific Cancer Cell Imaging.

Intracellular cancer-related biomarker imaging strategy has been used for specific identification of cancer cells, which was of great importance to accurate cancer clinical diagnosis and prognosis studies. Localized DNA circuits with improved sensitivity showed great potential for intracellular biomarkers imaging. However, the ability of localized DNA circuits to specifically image cancer cells is limited by off-site signal leakage associated with a single-biomarker sensing strategy. Herein, we integrated the endogenous enzyme-powered strategy with logic-responsive and localized signal amplifying capability to construct a self-assembled endogenously AND logic DNA nanomachine (EDN) for highly specific cancer cell imaging. When the EDN encountered a cancer cell, the overexpressed DNA repairing enzyme apurinic/apyrimidinic endonuclease 1 (APE1) and miR-21 could synergistically activate a DNA circuit via cascaded localized toehold-mediated strand displacement (TMSD) reactions, resulting in amplified fluorescence resonance energy transfer (FRET) signal. In this strategy, both endogenous APE1 and miR-21, served as two "keys" to activate the AND logic operation in cancer cells to reduce off-tumor signal leakage. Such a multiplied molecular recognition/activation nanomachine as a powerful toolbox realized specific capture and reliable imaging of biomolecules in living cancer cells.

Logic Signal Amplification System for Sensitive Electrochemiluminescence Detection and Subtype Identification of Cancer Cells.

Achieving sensitive detection and accurate identification of cancer cells is vital for diagnosing and treating the disease. Here, we developed a logic signal amplification system using DNA tetrahedron-mediated three-dimensional (3D) DNA nanonetworks for sensitive electrochemiluminescence (ECL) detection and subtype identification of cancer cells. Specially designed hairpins were integrated into DNA tetrahedral nanostructures (DTNs) to perform a catalytic hairpin assembly (CHA) reaction in the presence of target microRNA, forming hyperbranched 3D nanonetworks. Benefiting from the "spatial confinement effect," the DNA tetrahedron-mediated catalytic hairpin assembly (DTCHA) reaction displayed significantly faster kinetics and greater cycle conversion efficiency than traditional CHA. The resulting 3D nanonetworks could load a large amount of Ru(phen)32+, significantly enhancing its ECL signal, and exhibit detection limits for both miR-21 and miR-141 at the femtomolar level. The biosensor based on modular logic gates facilitated the distinction and quantification of cancer cells and normal cells based on miR-21 levels, combined with miR-141 levels, to further identify different subtypes of breast cancer cells. Overall, this study provides potential applications in miRNA-related clinical diagnostics.

AI-assisted mass spectrometry imaging with in situ image segmentation for subcellular metabolomics analysis.

Subcellular metabolomics analysis is crucial for understanding intracellular heterogeneity and accurate drug-cell interactions. Unfortunately, the ultra-small size and complex microenvironment inside the cell pose a great challenge to achieving this goal. To address this challenge, we propose an artificial intelligence-assisted subcellular mass spectrometry imaging (AI-SMSI) strategy with in situ image segmentation. Based on the nanometer-resolution MSI technique, the protonated guanine and threonine ions were respectively employed as the nucleus and cytoplasmic markers to complete image segmentation at the subcellular level, avoiding mutual interference of signals from various compartments in the cell. With advanced AI models, the metabolites within the different regions could be further integrated and profiled. Through this method, we decrypted the distinct action mechanism of isomeric drugs, doxorubicin (DOX) and epirubicin (EPI), only with a stereochemical inversion at C-4'. Within the cytoplasmic region, fifteen specific metabolites were discovered as biomarkers for distinguishing the drug action difference between DOX and EPI. Moreover, we identified that the downregulations of glutamate and aspartate in the malate-aspartate shuttle pathway may contribute to the higher paratoxicity of DOX. Our current AI-SMSI approach has promising applications for subcellular metabolomics analysis and thus opens new opportunities to further explore drug-cell specific interactions for the long-term pursuit of precision medicine.

Aggregation-Enabled Electrochemistry in Confined Nanopore Capable of Complementary Faradaic and Non-Faradaic Detection.

Electrochemistry that empowers innovative nanoscopic analysis has long been pursued. Here, the concept of aggregation-enabled electrochemistry (AEE) in a confined nanopore is proposed and devised by reactive oxygen species (ROS)-responsive aggregation of CdS quantum dots (QDs) within a functional nanopipette. Complementary Faradaic and non-Faradaic operations of the CdS QDs aggregate could be conducted to simultaneously induce the signal-on of the photocurrents and the signal-off of the ionic signals. Such a rationale permits the cross-checking of the mutually corroborated signals and thus delivers more reliable results for single-cell ROS analysis. Combined with the rich biomatter-light interplay, the concept of AEE can be extended to other stimuli-responsive aggregations for electrochemical innovations.

Electrogenerated chemiluminescence imaging of plasmon-induced electrochemical reactions at single nanocatalysts.

This study explores plasmon-induced electrochemical reactions on single nanoparticles using electrogenerated chemiluminescence microscopy (ECLM). Under laser irradiation, real-time screening showed lower plasmon-induced reaction efficiency for bimetallic Au@Pt nanoparticles compared to monometallic Au nanoparticles. ECLM offers a high-throughput imaging and precise quantitative approach for analyzing photo-electrochemical conversion at single nanoparticle level, valuable for both theoretical exploration and optimization of plasmonic nanocatalysts.

A hollow Ag/AgCl nanoelectrode for single-cell chloride detection.

This work reports the construction of a miniaturized Ag/AgCl nanoelectrode on a nanopipette, which is capable of dual-functions of single-cell drug infusion and chloride detection and is envisioned to promote the study of chloride-correlated therapeutic effects.

θ-Nanopore Ratiometry.

Developing nanoscale ratiometric techniques capable of biochemical response should prove of significance for precise applications with stringent spatial and biological restrictions. Here we present and devise the concept of θ-nanopore ratiometry, which uses ratiometric signals that could well address the serious concerns about device deviation in fabrication and nonspecific adsorption in the detection. As exemplified by a 200 nm θ-nanopore toward miRNA detection, the ±20 nm aperture drift could be mitigated and the issue of nonspecific adsorption could be minimized in the complex cytosolic environment. Practical application of this θ-nanopore ratiometry realizes the measurements of cytosolic miRNA-10b. This work has not only established a nanoscopic ratiometric technique but also enriched the extant armory of nanotools for single-cell studies and beyond.

Iontronic Photoelectrochemical Biorecognition Probing.

Herein, the first iontronic photoelectrochemical (PEC) biorecognition probing is devised by rational engineering of a dual-functional bioconjugate, i.e., a light-sensitive intercalated structural DNA, as a smart gating module confined within a nanotip, which could respond to both the incident light and biotargets of interest. Light stimulation of the bioconjugate could intensify the negative charge at the nano-orifice to sustain enhanced ionic current. The presence of proteins (e.g., acetylcholinesterase, AChE) or nucleic acids (e.g., microRNA (miR)-10b) could lead to bioconjugate release with altered ionic signaling. The practical applicability of the methodology is confirmed by AChE detection in human serum and miR-10b detection in single cells.

Simulation-Assisted DNA Nanodevice Serve as a General Optical Platform for Multiplexed Analysis of Micrornas.

Small frame nucleic acids (FNAs) serve as excellent carrier materials for various functional nucleic acid molecules, showcasing extensive potential applications in biomedicine development. The carrier module and function module combination is crucial for probe design, where an improper combination can significantly impede the functionality of sensing platforms. This study explores the effect of various combinations on the sensing performance of nanodevices through simulations and experimental approaches. Variances in response velocities, sensitivities, and cell uptake efficiencies across different structures are observed. Factors such as the number of functional molecules loaded, loading positions, and intermodular distances affect the rigidity and stability of the nanostructure. The findings reveal that the structures with full loads and moderate distances between modules have the lowest potential energy. Based on these insights, a multisignal detection platform that offers optimal sensitivity and response speed is developed. This research offers valuable insights for designing FNAs-based probes and presents a streamlined method for the conceptualization and optimization of DNA nanodevices.

Near-infrared II aggregation-induced electrochemiluminescence of organic dots.

For the first time, we report novel aggregation-induced electrochemiluminescence (AIECL) of organic dots in aqueous media, with near-infrared II (NIR-II) luminescence peaked at 906 nm. Furthermore, a hybrid mechanism of ECL generation is revealed by various experiments in conjunction with theoretical calculations. This work opens a window for exploring efficient organic dye-based NIR-II AIECL emitters.

An Ultrasensitive and Efficient microRNA Nanosensor Empowered by the CRISPR/Cas Confined in a Nanopore.

This work presents a clustered regularly interspaced short palindromic repeat (CRISPR)/Cas-nanopipette nano-electrochemistry (Cas = CRISPR-associated proteins) capable of ultrasensitive microRNA detection. Nanoconfinement of the CRISPR/Cas13a within a nanopipette leads to a high catalytic efficacy of ca. 169 times higher than that in bulk electrolyte, contributing to the amplified electrochemical responses. CRISPR/Cas13a-enabled detection of representative microRNA-25 achieves a low limit of detection down to 10 aM. Practical application of this method is further demonstrated for single-cell and real human serum detection. Its general applicability is validated by addressing microRNA-141 and the SARS-CoV-2 RNA gene fragment. This work introduces a new CRISPR/Cas-empowered nanotechnology for ultrasensitive nano-electrochemistry and bioanalysis.

Chemical Redox Cycling in an Organic Photoelectrochemical Transistor: Toward Dual Chemical and Electronic Amplification for Bioanalysis.

The organic photoelectrochemical transistor (OPECT) has been proven to be a promising platform to study the rich light-matter-bio interplay toward advanced biomolecular detection, yet current OPECT is highly restrained to its intrinsic electronic amplification. Herein, this work first combines chemical amplification with electronic amplification in OPECT for dual-amplified bioanalytics with high current gain, which is exemplified by human immunoglobulin G (HIgG)-dependent sandwich immunorecognition and subsequent alkaline phosphatase (ALP)-mediated chemical redox cycling (CRC) on a metal-organic framework (MOF)-derived BiVO4/WO3 gate. The target-dependent redox cycling of ascorbic acid (AA) acting as an effective electron donor could lead to an amplified modulation against the polymer channel, as indicated by the channel current. The as-developed bioanalysis could achieve sensitive HIgG detection with a good analytical performance. This work features the dual chemical and electronic amplification for OPECT bioanalysis and is expected to stimulate further interest in the design of CRC-assisted OPECT bioassays.

High-Precision Single-Cell microRNA Therapy by a Functional Nanopipette with Sensitive Photoelectrochemical Feedback.

This work proposes the concept of single-cell microRNA (miR) therapy and proof-of-concept by engineering a nanopipette for high-precision miR-21-targeted therapy in a single HeLa cell with sensitive photoelectrochemical (PEC) feedback. Targeting the representative oncogenic miR-21, the as-functionalized nanopipette permits direct intracellular drug administration with precisely controllable dosages, and the corresponding therapeutic effects can be sensitively transduced by a PEC sensing interface that selectively responds to the indicator level of cytosolic caspase-3. The experimental results reveal that injection of ca. 4.4 × 10-20  mol miR-21 inhibitor, i.e., 26488 copies, can cause the obvious therapeutic action in the targeted cell. This work features a solution to obtain the accurate knowledge of how a certain miR-drug with specific dosages treats the cells and thus provides an insight into futuristic high-precision clinical miR therapy using personalized medicine, provided that the prerequisite single-cell experiments are courses of personalized customization.

Tracking Ion Transport in Nanochannels via Transient Single-Particle Imaging.

The transport behavior of ions in the nanopores has an important impact on the performance of the electrochemical devices. Although the classical Transmission-Line (TL) model has long been used to describe ion transport in pores, the boundary conditions for the applicability of the TL model remain controversial. Here, we investigated the transport kinetics of different ions, within nanochannels of different lengths, by using transient single-particle imaging with temporal resolution up to microseconds. We found that the ion transport kinetics within short nanochannels may deviate significantly from the TL model. The reason is that the ion transport under nanoconfinement is composed of multi basic stages, and the kinetics differ much under different stage domination. With the shortening of nanochannels, the electrical double layer (EDL) formation would become the "rate-determining step" and dominate the apparent ion kinetics. Our results imply that using the TL model directly and treating the in-pore mobility as an unchanged parameter to estimate the ion transport kinetics in short nanopores/nanochannels may lead to orders of magnitude bias. These findings may advance the understanding of the nanoconfined ion transport and promote the related applications.

Ultrafast C-C and C-N bond formation reactions in water microdroplets facilitated by the spontaneous generation of carbocations.

Carbocations are important electrophilic intermediates in organic chemistry, but their formation typically requires harsh conditions such as extremely low pH, elevated temperature, strong oxidants and/or expensive noble-metal catalysts. Herein, we report the spontaneous generation of highly reactive carbocations in water microdroplets by simply spraying a diarylmethanol aqueous solution. The formation of transient carbocations as well as their ultrafast in-droplet transformations through carbocation-involved C-C and C-N bond formation reactions are directly characterized by mass spectrometry. The intriguing formation and stabilization of carbocations are attributed to the super acidity of the positively charged water microdroplets as well as the high electric fields at the water-air interfaces. Without the utilization of external acids as catalysts, we believe that these microdroplet reactions would pose a new and sustainable way for the construction of aryl-substituted compounds.

In Situ Probing the Short-Lived Intermediates in Visible-Light Heterogeneous Photocatalysis by Mass Spectrometry.

Visible-light-mediated heterogeneous photocatalysis has recently emerged as an environmentally friendly and energy-sustainable alternative for organic transformations. Despite the advancements in developing wide varieties of photocatalysts during the past decades, the accurate probing and identification of the photogenerated species, especially the short-lived radical intermediates, are still challenging. In this work, we reported a hybrid ion emitter that integrated with a pico-liter heterogeneous photocatalytic reactor, which was fabricated by depositing the photocatalyst (e.g., TiO2) into the front tip of a quartz micropipette. Benefited from the dual-function feature of the hybrid micropipette (i.e., a clog-free tip-confined pico-liter reactor for heterogeneous photocatalysis and an ion emitter for nanoelectrospray ionization), sensitized photoredox reactions at the catalyst-solution interface can be triggered upon visible-light irradiation using a cheap LED laser (453 nm), and the newly produced transient radical intermediates can be rapidly transformed into gaseous ions for mass spectrometric identification. Using this novel low-delay coupling device, photogenerated intermediates, including the cationic radicals produced during the photooxidation of anilines and the anionic radicals produced during the photoreduction of quinones, were successfully captured by mass spectrometry. We believe that our hybrid photochemical microreactor/ion emitter has provided a new and powerful tool for exploring the complicated heterogeneous photochemical processes, especially their ultrafast initial transformations.

θ-Nanopipette for Single-Cell Resistive-Pulse Profiling of DNA Repair Proteins Accompanied by Drug Evaluation.

Single-cell analysis of the DNA repair protein is important but remains unachieved. Exploration of nanopipettte technologies in single-cell electroanalysis has recently seen rapid growth, while the θ-nanopipette represents an emerging technological frontier with its potential largely veiled. Here a θ-nanopipette is first applied for single-cell resistive-pulse sensing (RPS) of the important DNA repair protein O6-alkylguanine DNA alkyltransferase (hAGT). The removal of alkyl mutations by hAGT could restore the damaged aptamer linking with a structural DNA carrier, allowing the selective binding of the aptamer to thrombin with precisely matched size to produce distinct RPS signals when passing through the orifice. Kinetic analysis of hAGT repair was studied. Meanwhile, the device shows the simultaneous on-demand infusion of inhibitors to inactivate the hAGT activity, indicative of its potential in drug screening for enhanced chemotherapy. This work provides a new paradigm for θ-nanopipette-based single-cell RPS of a DNA repair protein accompanied by drug evaluation.

Plasmonic Cavity-Catalysis by Standing Hot Carrier Waves.

Manipulating active sites of catalysts is crucial but challenging in catalysis science and engineering. Beyond the design of the composition and structure of catalysts, the confined electromagnetic field in optical cavities has recently become a promising method for catalyzing chemical reactions via strong light-matter interactions. Another form of confined electromagnetic field, the charge density wave in plasmonic cavities, however, still needs to be explored for catalysis. Here, we present an unprecedented catalytic mode based on plasmonic cavities, called plasmonic cavity-catalysis. We achieve direct control of catalytic sites in plasmonic cavities through standing hot carrier waves. Periodic catalytic hotspots are formed because of localized energy and carrier distribution and can be well tuned by cavity geometry, charge density, and excitation angle. We also found that the catalytic activity of the cavity mode increases several orders of magnitude compared with conventional plasmonic catalysis. We ultimately demonstrate that the locally concentrated long-lived hot carriers in the standing wave mode underlie the formation of the catalytic hotspots. Plasmonic cavity-catalysis provides a new approach to manipulate the catalytic sites and rates and may expand the frontier of heterogeneous catalysis.