Photoexcited Ru single-atomic sites for efficient biomimetic redox catalysis.

The unsatisfactory catalytic activity of nanozymes owing to their inefficient electron transfer (ET) is the major challenge in biomimetic catalysis-related biomedical applications. Inspired by the photoelectron transfers in natural photoenzymes, we herein report a photonanozyme of single-atom Ru anchored on metal-organic frameworks (UiO-67-Ru) for achieving photoenhanced peroxidase (POD)-like activity. We demonstrate that the atomically dispersed Ru sites can realize high photoelectric conversion efficiency, superior POD-like activity (7.0-fold photoactivity enhancement relative to that of UiO-67), and good catalytic specificity. Both in situ experiments and theoretical calculations reveal that photoelectrons follow the cofactor-mediated ET process of enzymes to promote the production of active intermediates and the release of products, demonstrating more favorable thermodynamics and kinetics in H2O2 reduction. Taking advantage of the unique interaction of the Zr-O-P bond, we establish a UiO-67-Ru-based immunoassay platform for the photoenhanced detection of organophosphorus pesticides.

Atomic-level design of metalloenzyme-like active pockets in metal-organic frameworks for bioinspired catalysis.

Natural metalloenzymes with astonishing reaction activity and specificity underpin essential life transformations. Nevertheless, enzymes only operate under mild conditions to keep sophisticated structures active, limiting their potential applications. Artificial metalloenzymes that recapitulate the catalytic activity of enzymes can not only circumvent the enzymatic fragility but also bring versatile functions into practice. Among them, metal-organic frameworks (MOFs) featuring diverse and site-isolated metal sites and supramolecular structures have emerged as promising candidates for metalloenzymes to move toward unparalleled properties and behaviour of enzymes. In this review, we systematically summarize the significant advances in MOF-based metalloenzyme mimics with a special emphasis on active pocket engineering at the atomic level, including primary catalytic sites and secondary coordination spheres. Then, the deep understanding of catalytic mechanisms and their advanced applications are discussed. Finally, a perspective on this emerging frontier research is provided to advance bioinspired catalysis.

Nanozyme Metabolism Controls Air Pollution over an Atomic Potassium Cyano Site.

Increasing threats of air pollution prompt the design of air purification systems. As a promising initiative defense strategy, nanocatalysts are integrated to catalyze the detoxification of specific pollutants. However, it remains a grand challenge to tailor versatile nanocatalysts to cope with diverse pollutants in practice. Here, we report a nanozyme metabolism system to realize broad-spectrum protection from air pollution. Atomic K-modified carbon nitride featuring flavin oxidase-like and peroxidase-like activities was synthesized to initiate nanozyme metabolism. In situ experiments and theoretical investigations collectively show that K sites optimize the geometric and electronic structure of cyano sites for both enzyme-like activities. As a proof of concept, the nanozyme metabolism was applied to the mask against volatile organic compounds, persistent organic pollutants, reactive oxygen species, bacteria, and so on. Our finding provides a thought to tackle global air pollution and deepens the understanding of nanozyme metabolism.

Deep learning-based pathological prediction of lymph node metastasis for patient with renal cell carcinoma from primary whole slide images.

BACKGROUND: Metastasis renal cell carcinoma (RCC) patients have extremely high mortality rate. A predictive model for RCC micrometastasis based on pathomics could be beneficial for clinicians to make treatment decisions. METHODS: A total of 895 formalin-fixed and paraffin-embedded whole slide images (WSIs) derived from three cohorts, including Shanghai General Hospital (SGH), Clinical Proteomic Tumor Analysis Consortium (CPTAC) and Cancer Genome Atlas (TCGA) cohorts, and another 588 frozen section WSIs from TCGA dataset were involved in the study. The deep learning-based strategy for predicting lymphatic metastasis was developed based on WSIs through clustering-constrained-attention multiple-instance learning method and verified among the three cohorts. The performance of the model was further verified in frozen-pathological sections. In addition, the model was also tested the prognosis prediction of patients with RCC in multi-source patient cohorts. RESULTS: The AUC of the lymphatic metastasis prediction performance was 0.836, 0.865 and 0.812 in TCGA, SGH and CPTAC cohorts, respectively. The performance on frozen section WSIs was with the AUC of 0.801. Patients with high deep learning-based prediction of lymph node metastasis values showed worse prognosis. CONCLUSIONS: In this study, we developed and verified a deep learning-based strategy for predicting lymphatic metastasis from primary RCC WSIs, which could be applied in frozen-pathological sections and act as a prognostic factor for RCC to distinguished patients with worse survival outcomes.

Fabrication of High-Negatively Charged Bicelle-Mediated Supported Lipid Bilayer.

Supported lipid bilayers (SLBs), two-dimensional lipid films formed on a solid-supporting substrate, serve as models for biomembranes and exhibit remarkable potential in chemistry, biology, and medicine. However, preparing SLBs with highly negatively charged contents on the negatively charged surface by overcoming electrostatic repulsion remains a challenge. Here, a creative bicelle-mediated and divalent cation-free SLB preparation method with the assistance of phosphate-buffered saline (PBS) solution was proposed, which can form the SLBs containing 50% DOPS or 30% CL on the silica surface monitored by a quartz crystal microbalance with dissipation (QCM-D). Results of molecular dynamics (MD) simulation indicate that electrostatic repulsion can be overcome by the increased number of hydrogen bonds caused by the adsorption of dihydrogen phosphate ions onto the headgroups of lipids. In addition, the negatively charged SLB formation was identified to be a three-step kinetic process, which differs from a two-step mechanism in the case of amphoteric SLB. The extra kinetic step can be attributed to the reduction in the number of intermolecular hydrogen bonds and the ordering of water molecules in the hydration layer. This investigation resolves the challenge of fabricating SLB over negatively charged surfaces and offers a fresh perspective on the SLB assembly methodology.

Amino-Ligand-Coordinated Dicopper Active Sites Enable Catechol Oxidase-Like Activity for Chiral Recognition and Catalysis.

Developing highly active and selective advanced nanozymes for enzyme-mimicking catalysis remains a long-standing challenge for basic research and practical applications. Herein, we grafted a chiral histidine- (His-) coordinated copper core onto Zr-based metal-organic framework (MOF) basic backbones to structurally mirror the bimetal active site of natural catechol oxidase. Such a biomimetic fabricated process affords MOF-His-Cu with catechol oxidase-like activity, which can catalyze dehydrogenation and oxidation of o-diphenols and then transfer electrons to O2 to generate H2O2 by the cyclic conversion of Cu(II) and Cu(I). Specifically, the elaborate incorporation of chiral His arms results in higher catalytic selectivity over the chiral catechol substrates than natural enzyme. Density functional theory calculations reveal that the binding energy and potential steric effect in active site-substrate interactions account for the high stereoselectivity. This work demonstrates efficient and selective enzyme-mimicking catalytic processes and deepens the understanding of the catalytic mechanism of nanozymes.

High-Indexed Intermetallic Pt3Sn Nanozymes with High Activity and Specificity for Sensitive Immunoassay.

Great efforts have been made to expand the application fields of nanozymes, which puts forward requirements for nanozymes with both superior catalytic activity and specificity. Herein, we reported the high-indexed intermetallic Pt3Sn (H-Pt3Sn) with high peroxidase-like activity and specificity. The resultant H-Pt3Sn exhibits a specific activity of 345.3 U/mg, which is 1.82 times higher than Pt. Moreover, H-Pt3Sn possesses negligible oxidase-like and catalase-like activities, achieving superior catalytic specificity toward H2O2 activation. Experimental and theoretical calculations reveal both the splitting energy for adsorbed H2O2 and the energy barrier for the rate-determining step of H-Pt3Sn are significantly decreased compared with Pt3Sn and Pt. Finally, a nanozyme-linked immunosorbent assay is successfully developed, achieving the sensitive and accurate colorimetric detection for carcinoembryonic antigen with a low detection limit of 0.49 pg/mL and showing practical feasibility in serum sample detection.

Harmonizing Enzyme-like Cofactors to Boost Nanozyme Catalysis.

Engineering isolated metal sites resembling the primary coordination sphere of metallocofactors enables atomically dispersed materials as promising nanozymes. However, most existing nanozymes primarily focus on replicating specific metallocofactors while neglecting other supporting cofactors within active pockets, leading to reduced electron transfer (ET) efficiency and thus inferior catalytic performances. Herein, we report a metal-organic framework UiO-67 nanozyme with atomically dispersed iron sites, which involves multiple tailored enzyme-like nanocofactors that synergistically drive the ET process for enhanced peroxidase-like catalysis. Among them, the linker-coupled atomic iron site plays a critical role in substrate activation, while bare linkers and zirconia nodes facilitate the ET efficiency of intermediates. The synergy of three nanocofactors results in a 4.29-fold enhancement compared with the single effort of isolated metal site-based nanocofactor, holding promise in immunoassay for sensitive detection of chlorpyrifos. This finding opens a new way for designing high-performance nanozymes by harmonizing various nanocofactors at the atomic and molecular scale.

Single-Cell Analysis and Classification according to Multiplexed Proteins via Microdroplet-Based Self-Driven Magnetic Surface-Enhanced Raman Spectroscopy Platforms Assisted with Machine Learning Algorithms.

A microdroplet-based surface-enhanced Raman spectroscopy (microdroplet SERS) platform was constructed to envelop individual cells in microdroplets, followed by the SERS detection of their extracellular vesicle-proteins (EV-proteins) via the in-drop immunoassays by use of immunomagnetic beads (iMBs) and immuno-SERS tags (iSERS tags). A unique phenomenon is found that iMBs can start a spontaneous reorientation on the probed cell surface based on the electrostatic force-driven interfacial aggregation effect, which leads EV-proteins and iSERS tags to be gathered from a liquid phase to a cell membrane interface and significantly improves SERS sensitivity to the single-cell analysis level due to the formation of numbers of SERS hotspots. Three EV-proteins from two breast cancer cell lines were collected and further analyzed by machine learning algorithmic tools, which will be helpful for a deeper understanding of breast cancer subtypes from the view of EV-proteins.

Direct Virus Gene Detection: A CRISPR/dCas9-Mediated Surface-Enhanced Raman Scattering Strategy with Enzyme-Catalyzed Signal Amplification.

The aberrant growth of cervical cells caused by the infection of human papillomavirus (HPV) may cause cervical cancer. In order to effectively prevent the occurrence of cervical cancer and for better follow-up treatment after surgery, a rapid and reliable detection method of HPV DNA is essential. Here, a surface-enhanced Raman scattering (SERS) detection method was developed based on the clustered regularly interspaced short palindromic repeats (CRISPR)/dCas9 technique and the enzyme catalysis amplification reaction, which achieved a simple and rapid detection of low-content HPV genes. The CRISPR/dCas9/sgRNA complex was anchored above a magnetic bead, which can precisely capture target DNA sequences, exhibiting high selectivity for HPV genes. When the biotinylated target DNAs exist, they can bridge a streptavidin-modified horse radish peroxidase (HRP) to the magnetic bead, producing an HRP-decorated conjugate. This conjugate allows an HRP-catalyzed reaction for its substrate (3,3',5,5'-tetramethylbenzidine, TMB). Gold nanostars with a silica shell exhibiting the lightning rod effect of SERS were employed to measure the SERS spectra of the oxidative product of TMB. Enzyme catalysis and SERS co-contribute to the SERS signal output, ensuring a high detection sensitivity. This method is a proof of concept for detecting HPV DNAs in a complex system. The current method can be applied to other target DNAs simply by changing the sgRNA sequence. Many superiors portend that the CRISPR/dCas9-based SERS method is promising for further clinical application.

Iron Single-Atom Catalyst-Enabled Peroxydisulfate Activation Enhances Cathodic Electrochemiluminescence of Tris(bipyridine)ruthenium(II).

The tris(bipyridine)ruthenium(II) (Ru(bpy)32+)-tripropylamine anodic electrochemiluminescence (ECL) system has been widely applied in commercial bioanalysis. However, the presence of amine compounds in the biological environment results in unavoidable anodic interference signals, which hinder further extensive use of the system. In contrast, the cathodic Ru(bpy)32+ ECL system can overcome these limitations. The Ru(bpy)32+/peroxydisulfate (S2O82-, PDS) ECL system has been extensively employed due to its ability to produce a sulfate radical anion (SO4•-) with strong oxidation ability, which enhances the ECL signal. However, the symmetrical molecular structure of PDS makes it challenging to be activated and causes low luminescence efficiency. To address this issue, we propose an efficient Ru(bpy)32+-based ternary ECL system that uses the iron-nitrogen-carbon single-atom catalyst (Fe-N-C SAC) as an advanced accelerator. Fe-N-C SAC can efficiently activate PDS into reactive oxygen species at a lower voltage, which significantly boosts the cathodic ECL emission of Ru(bpy)32+. Benefiting from the outstanding catalytic activity of Fe-N-C SAC, we successfully established an ECL biosensor that detects alkaline phosphatase activity with high sensitivity, demonstrating the feasibility of practical application.

Nanozymatic Biofuel Cell-Enabled Self-Powered Sensing System for a Sensitive Immunoassay.

Self-powered sensing system (SPSS) integrating the enzymatic biofuel cell and biosensing platform has attracted tremendous interest. However, natural enzymes suffer from the intrinsic drawbacks of enzymes and enzymatic proteins. Nanozymes with enzyme-like activities are the ideal alternatives to enzymes, and it is greatly challenging to explore high-performance nanozymatic biofuel cell for SPSS. Herein, the advanced nanozymatic biofuel cell-enabled SPSS is developed for the sensitive detection of the prostate-specific antigen (PSA), where Ir single atoms supported by nitrogen-doped carbon and Au nanozymes serve as the cathode and anode, respectively. Based on the excellent electrochemical activity and stability, the resultant nanozymatic biofuel cell exhibits a higher power output and open-circuit potential than the Pt/C-based counterpart, which is beneficial for the application of SPSS. As a proof of concept, the nanozymatic biofuel cell-enabled SPSS shows a wide detection range of 0.2-500 ng mL-1 with a detection limit of 62 pg mL-1 for PSA, which provides new insight into broadening the application scenarios of nanozymes.

Photothermal-Switched Single-Atom Nanozyme Specificity for Pretreatment and Sensing.

Various applications lead to the requirement of nanozymes with either specific activity or multiple enzyme-like activities. To this end, intelligent nanozymes with freely switching specificity abilities hold great promise to adapt to complicated and changeable practical conditions. Herein, a nitrogen-doped carbon-supported copper single-atom nanozyme (named Cu SA/NC) with switchable specificity is reported. Atomically dispersed active sites endow Cu SA/NC with specific peroxidase-like activity at room temperature. Furthermore, the intrinsic photothermal conversion ability of Cu SA/NC enables the specificity switch by additional laser irradiation, where photothermal-induced temperature elevation triggers the expression of oxidase-like and catalase-like activity of Cu SA/NC. For further applications in practice, a pretreatment-and-sensing integration kit (PSIK) is constructed, where Cu SA/NC can successively achieve sample pretreatment and sensitive detection by switching from multi-activity mode to specific-activity mode. This study sets the foundation for nanozymes with switchable specificity and broadens the application scope in point-of-care testing.

A SERS/fluorescence dual-mode immuno-nanoprobe for investigating two anti-diabetic drugs on EGFR expressions.

A surface-enhanced Raman scattering (SERS)/fluorescence dual-mode nanoprobe was proposed to assess anti-diabetic drug actions from the expression level of the epidermal growth factor receptor (EGFR), which is a significant biomarker of breast cancers. The nanoprobe has a raspberry shape, prepared by coating a dye-doped silica nanosphere with a mass of SERS tags, which gives high gains in fluorescence imaging and SERS measurement. The in situ detection of EGFR on the cell membrane surfaces after drug actions was achieved by using this nanoprobe, and the detection results agree with the enzyme-linked immunosorbent assay (ELISA) kit. Our study suggests that rosiglitazone hydrochloride (RH) may be a potential drug for diabetic patients with breast cancer, while the anti-cancer effect of metformin hydrochloride (MH) is debatable since MH slightly promotes the EGFR expression of MCF-7 cells in this study. This sensing platform endows more feasibility for highly sensitive and accurate feedback of pesticide effects at the membrane protein level.

Nanobiocatalysis: a materials science road to biocatalysis.

With high activity and specificity to conduct catalysis under mild conditions, enzymes show great promise in many fields. However, they are not acclimatized to environments in practice after leaving the familiar biological conditions. Aiming at this issue, nanobiocatalysis, a fresh area integrating nanotechnology and enzymatic catalysis, is expected to design biocatalysis based on materials science. Specifically, nano-integrated biocatalysis and bio-inspired nanocatalysis are considered as two effective nanobiocatalytic systems to meet different design needs. Notably, both systems are not entirely separated, and the combination of both further sparks more possibilities. This review summarizes the type, construction, and function of nanobiocatalytic systems, analyzing the pros and cons of different strategies. Moreover, the corresponding applications in bioassay, biotherapy, and environmental remediation are highlighted. We hope that the advent of nanobiocatalysis will help in grasping the inherence of biocatalysis and propel biocatalytic applications.

Influence of Rotational Speed on Isothermal Piston Compression System.

An isothermal piston is a device that can achieve near-isothermal compression by enhancing the heat transfer area with a porous media. However, flow resistance between the porous media and the liquid is introduced, which cannot be neglected at a high operational speed. Thus, the influence of rotational speed on the isothermal piston compression system is analyzed in this study. A flow resistance mathematical model is established based on the face-centered cubic structure hypothesis. The energy conservation rate and efficiency of the isothermal piston are defined. The effect of rotational speed on resistance is discussed, and a comprehensive energy conservation performance assessment of the isothermal piston is analyzed. The results show that the increasing rate of the resistance work increases significantly proportional to the rotational speed, and the proportion of resistance work in the total work increases gradually and sharply. The total work including compression and resistance cannot be larger than the compression work under adiabatic conditions. The maximum rotational speed is 650 rpm.

Metal-Organic Framework-Based Artificial Organelle Corrects Microenvironment Interference for Accurate Intratumoral Glucose Analysis.

Enzymatic catalysis with high efficiency allows them a great prospect in metabolite monitoring in living cells. However, complex tumor microenvironments, such as acidity, H2 O2 , and hypoxia, are bound to disturb catalytic reactions for misleading results. Here, we report a spatially compartmentalized artificial organelle to correct intratumoral glucose analysis, where the zeolitic imidazolate framework-8 immobilized glucose oxidase-horseradish peroxidase cascade core and catalase-directed shell act as signal transduction and guarding rooms respectively. The acid-digested core and stable shell provide appropriate spaces to boost biocatalytic efficiency with good tolerability. Notably, the endogenous H2 O2 is in situ decomposed to O2 by catalase, which not only overcomes the interference in signal output but also alleviates the hypoxic states to maximize glucose oxidation. The marked protective effect and biocompatibility render artificial organelles to correct the signal transduction for dynamic monitoring glucose in vitro and in vivo, achieving our goal of accurate intratumoral metabolite analysis.

Water Activation for Boosting Electrochemiluminescence.

In conventional luminol electrochemiluminescence (ECL) systems, hydrogen peroxide and dissolved oxygen are employed as typical co-reactants to produce reactive oxygen species (ROS) for efficient ECL emission. However, the self-decomposition of hydrogen peroxide and limited solubility of oxygen in water inevitably restrict the detection accuracy and luminous efficiency of luminol ECL system. Inspired by ROS-mediated ECL mechanism, for the first time, we used cobalt-iron layered double hydroxide as co-reaction accelerator to efficiently activate water to generate ROS for enhancing luminol emission. Experimental investigations verify the formation of hydroxyl and superoxide radicals in the process of electrochemical water oxidation, which subsequently react with luminol anion radicals to trigger strong ECL signals. Finally, the detection of alkaline phosphatase has been successfully achieved with impressive sensitivity and reproducibility for practical sample analysis.

Bioinspired Single-Atom Sites Enable Efficient Oxygen Activation for Switching Anodic/Cathodic Electrochemiluminescence.

Exploring advanced co-reaction accelerators with superior oxygen reduction activity that generate rich reactive oxygen species (ROS) has attracted great attention in boosting luminol-O2 electrochemiluminescence (ECL). However, tuning accelerators for efficient and selective catalytic O2 activation to switch anodic/cathodic ECL is very challenging. Herein, we report that enzyme-inspired Fe-based single-atom catalysts with axial N/C coordination structures (FeN5 , FeN4 © SACs) can generate specific ROS for cathodic/anodic ECL conversion. Mechanistic studies reveal that FeN5 sites prefer to produce highly active hydroxyl radicals and afford direct cathodic luminescence by promoting the cleavage of O-O bonds through N-induced electron redistribution. In contrast, FeN4 © sites tend to produce superoxide radicals, resulting in inefficient anodic ECL. Benefiting from the enhanced cathodic ECL, FeN5 SAC-based immunosensor was constructed for the sensitive detection of cancer biomarkers.

Single-Cell VEGF Analysis by Fluorescence Imaging-Microfluidic Droplet Platform: An Immunosandwich Strategy on the Cell Surface.

Despite recent advances in single-cell analysis techniques, the ability of single-cell analysis platforms to track specific cells that secreted cytokines remains limited. Here, we report a microfluidic droplet-based fluorescence imaging platform that can analyze single cell-secreted vascular endothelial growth factor (VEGF), an important regulator of physiological and pathological angiogenesis, to explore cellular physiological clues at the single-cell level. Two kinds of silica nanoparticle (NP)-based immunoprobes were developed, and they were bioconjugated to the membrane proteins of the probed cell surface via the bridging of secreted VEGF. Thus, an immunosandwich assay was built above the probed cell via fluorescence imaging analysis of each cell in isolated droplets. This analytical platform was used to compare the single-cell VEGF secretion ability of three cell lines (MCF-7, HeLa, and H8), which experimentally demonstrates the cellular heterogeneity of cells in secreting cytokines. The uniqueness of this method is that the single-cell assay is carried out above the cell of interest, and no additional carriers (beads or reporter cells) for capturing analytes are needed, which dramatically improves the availability of microdroplets. This single-cell analytical platform can be applied for determining other secreted cytokines at the single-cell level by changing other immune pairs, which will be an available tool for exploring single-cell metabonomics.