Affinity biosensors play a crucial role in clinical diagnosis, pharmaceuticals, immunology, and other areas of human health. Affinity biosensors rely on the specific binding between target analytes and biological ligands such as antibodies, nucleic acids, aptamers, or other receptors to primarily generate electrochemical or optical signals. Considerable effort has been put into improving the performance of the affinity technologies to make them more sensitive, efficient and reproducible, of the many approaches electrokinetic phenomena are a viable option. In this perspective, studies that combine electrokinetic phenomena with affinity biosensor are discussed about their promise for achieving higher sensitivity and lower detection limit.

Methylation of the promoter region of cancer related genes plays a crucial role in the occurrence and development of cancer, and the degree of methylation has great potential for the early cancer diagnosis. At present, the technology used to quantify DNA methylation is mainly based on the DNA sequencing which are time-consuming and high-cost in the relating application. We have developed an ultrasensitive method of methylation specific enzyme-linked oligonucleotide assays (MS-ELONA) to detect and quantify the level of DNA methylation. We could detect as little as 2 pg of methylated DNA in the 100000-fold excess of unmethylated genes, and discriminate prostate cancer from benign prostatic hyperplasia (BPH) and control with serum samples. We also demonstrate the reversibility of DNA methylation modification by treatment with demethylation drugs. With 16-channel electrochemical work station, our research reveals a simple and inexpensive method to quantify the methylation level of specially appointed genes, and have the potential to be applied in the clinical research.

Circulating tumor DNA (ctDNA) was short and rare, making the detection performance of the current targeted sequencing methods unsatisfying. We developed the One-PrimER Amplification (OPERA) system and examined its performance in detecting mutations of low variant allelic frequency (VAF) in various samples with short-sized DNA fragments. In cell line-derived samples containing sonication-sheared DNA fragments with 50-150 bp, OPERA was capable of detecting mutations as low as 0.0025% VAF, while CAPP-Seq only detected mutations of >0.03% VAF. Both single nucleotide variant and insertion/deletion can be detected by OPERA. In synthetic fragments as short as 80 bp with low VAF (0.03%-0.1%), the detection sensitivity of OPERA was significantly higher compared to that of droplet digital polymerase chain reaction. The error rate was 5.9×10-5 errors per base after de-duplication in plasma samples collected from healthy volunteers. By suppressing "single-strand errors", the error rate can be further lowered by >5 folds in EGFR T790M hotspot. In plasma samples collected from lung cancer patients, OPERA detected mutations in 57.1% stage I patients with 100% specificity and achieved a sensitivity of 30.0% in patients with tumor volume of less than 1 cm3. OPERA can effectively detect mutations in rare and highly-fragmented DNA.

Quantification of microRNAs (miRNAs) at the single-molecule level is of great significance for clinical diagnostics and biomedical research. The challenges lie in the limits to transforming single-molecule measurements into quantitative signals. To address these limits, here, we report a new approach called a Single Microbead-based Space-confined Digital Quantification (SMSDQ) to measure individual miRNA molecules by counting gold nanoparticles (AuNPs) with localized surface plasmon resonance (LSPR) light-scattering imaging. One miRNA target hybridizes with the alkynyl-modified capture DNA probe immobilized on a microbead (60 μm) and the azide-modified report DNA probe anchored on AuNP (50 nm), respectively. Through the click reaction between the alkynyl and azide group, a single microbead can covalently link the AuNPs in the confined space within the view of the microscope. By digitally counting the light-scattering spots of AuNPs, we demonstrated the proposed approach with single-molecule detection sensitivity and high specificity of single-base discrimination. Taking the advantages of ultrahigh sensitivity, specificity, and the digital detection manner, the approach is suitable for evaluating cell heterogeneity and small variations of miRNA expression and has been successfully applied to direct quantification of miRNAs in one-tenth single-cell lysates and serum samples without RNA-isolated and nucleic acid amplification steps.

The use of phages-a natural predator of bacteria-has emerged as a therapeutic strategy for treating multidrug-resistant bacterial infections; thus, the isolation and detection of phages from the environment is crucial for advancing phage therapy. Herein, for the first time, we propose a nanoplasmonic-based biodetection platform for phages that utilizes bacterial outer membranes (OMs) as a biorecognition element. Conventional biosensors based on phage-bacteria interactions encounter multiple challenges due to the bacteriolytic phages and potentially toxic bacteria, resulting in instability and risk in the measurement. Therefore, instead of whole living bacteria, we employ a safe biochemical OMs fraction presenting phage-specific receptors, allowing the robust and reliable phage detection. In addition, the biochip is constructed on bimetallic nanoplasmonic islands through solid-state dewetting for synergy between Au and Ag, whereby sensitive detection of phage-OMs interactions is achieved by monitoring the absorption peak shift. For high detection performance, the nanoplasmonic chip is optimized by systematically investigating the morphological features, e.g., size and packing density of the nanoislands. Using our optimized device, phages are detected with high sensitivity (≥∼104 plaques), specificity (little cross-reactivity), and affinity (stronger binding to the host OMs than anti-bacterial antibodies), further exhibiting the cell-killing activities.

Norfloxacin (NOR) residues in water pose a serious threat to human health via the food chain, necessitating the development of a rapid on-site antibiotic detection technique. In this work, we utilize electrostatic spinning technology that combines polyacrylonitrile (PAN) fibers and adenosine triphosphate (ATP)-rare earth metal Tb3+ complexes (ATP/Tb) to construct a new ternary film-based sensor for sensitive, quick, and convenient field testing of NOR in water. The operating mechanism is that the ternary system produces gradually enhanced bright green fluorescence at increasing concentrations of NOR. The unique fluorescence property of the ternary systems is attributed to the use of ATP, rather than the commonly used adenosine monophosphate (AMP), to coordinate with Tb3+, which avoided the possible fluorescence quenching from competitive water binding. Benefiting from the PAN nanofiber's superior stability, acid, and alkali resistance, and flexibility as support, the ternary system exhibited a good linear response to NOR in a wide dynamic range of 0.04-30 μM at the detection limit of 16 nM. Additionally, the combination of a smartphone color recognition app allows for quick on-scene NOR detection. This film sensing strategy is instructive for the development of smart and portable sensing platforms for real-time detection of analytes such as antibiotics, pesticide residues, and hazardous materials in water bodies.

The origin and dissemination of silk have been hotly debated in the field of archaeology, and the key to resolving this controversy lies in the detection and species identification of ancient silk microtraces. Herein, a taxonomically specific anti-fibroin monoclonal antibody was successfully prepared and a layer-by-layer self assembly electrochemical immunosensor was innovatively proposed for detecting silk traces based on flexible carbon cloth. The immunosensor possessed a broad linear range of 10-2-103 ng mL-1 and a detection limit of 2.15 pg mL-1 for the ultrasensitive detection of Bombyx mori silk traces. In addition, the elaborate immunosensor exhibited satisfactory high specificity, storage stability and reproducibility. In particular, the qualitative and quantitative performance of the immunosensor was excellent in the analysis of archaeological samples. Therefore, this work demonstrates that the proposed method not only provides a reliable analytical tool for exploring the origin and spread of archaeological silk, but also improves our understanding of how to use emerging materials like two-dimensional titanium carbide to creat innovative biosensors.

The identification and detection of mesenchymal circulating tumor cells （mCTCs） is important for early warning of tumor metastasis. The majority of conventional detection methods for CTCs rely on the recognition of epithelial biomarkers, which is technically challenging for detecting CTCs with epithelial-mesenchymal transition (EMT)-induced phenotypic alteration. In this work, we have constructed a label-free biosensor for sensitive electrochemical assay of mCTCs. In our design, the capture probe can recognize the vimentin overexpressed on the surface of mCTCs with high specificity. Meantime, the network-like DNA nanoprobes with multiple G-quadruplex/hemin complexes and multiple cholesterol molecules can be grafted to the cell surface based on the high affinity between cholesterol molecules and cell membrane. Owing to the mimic horseradish peroxidase of G-quadruplex/hemin complexes, strong electrochemical responses will be obtained for sensitive quantification of mCTCs with a detection limit of 8 cell mL-1. Moreover, the biosensor can effectively overcome the interference of vimentin negative cells or secretory vimentin, and realize the recovery tests in whole blood with high accuracy, thereby may further promoting the diagnosis and personalized treatment of cancer in clinic.

Exosomal surface proteins are potentially useful for breast cancer diagnosis and awareness of risk. However, some detection techniques involving complex operations and expensive instrumentation are limited to advance to clinical applications. To solve this problem, we develop a dual-modal sensor combining naked-eye detection and electrochemical assay of exosomal surface proteins from breast cancer. Most of existing sensors rely on aptamers recognizing exosomes and generating amplified signals at the same time, which require well-designed aptamer probes to avoid difficulties in identifying exosomes. In our work, aptamers not bound by the exosomes can serve as complete templates to induce formation of G quadruplexes. The peroxidase activity of the G-quadruplex/hemin DNAzyme catalyze substrates can generate both color and electrochemical signals. The developed dual-modal sensor offers a remarkable capability to differentiate nonmetastatic, metastatic breast cancer patients, and healthy individuals through the analysis of exosomal surface proteins. The sensor's distinctive features, including its universality, simplicity, and cost-effectiveness, position it as a promising diagnostic tool in breast cancer research and clinical practice.

Nanomaterials with enzyme-like catalytic features (nanozymes) find wide use in analytical sensing. Apart from catalytic characteristics, some other interesting functions coexist in the materials. How to combine these properties to design multifunctional nanozymes for new sensing strategy development is challenging. Besides, in nanozymes it is still a challenge to conveniently control the catalytic process, which also hinders their further applications in advanced biochemical analysis. To remove the above barriers, here we design a light-controllable multifunctional nanozyme, namely manganese-inserted cadmium telluride (Mn-CdTe) particles, that integrates oxidase-like activity with luminescence together, to achieve the fluorometric/colorimetric dual-mode detection of toxic mercury ions (Hg2+) at ambient pH. The Mn-CdTe exhibits a light-triggered oxidase-mimicking catalytic behavior to induce chromogenic reactions, thus enabling one to start or stop the catalytic progress easily via applying or withdrawing light irradiation. Meanwhile, the quantum dot material can exhibit bright photoluminescence, which provides the fluorometric channel to sense targets. When Hg2+ is introduced, it rapidly leans toward Mn-CdTe through electrostatic interaction and Te-Hg bonding and induces the aggregation of the latter. As a result, the luminescence of Mn-CdTe is dynamically quenched, and the masking of active sites in aggregated Mn-CdTe leads to the decrease of light-initiated oxidase-mimetic activity. According to this principle, a new fluorometric/colorimetric bimodal method was established for Hg2+ determination with excellent performance. A 3D-printed portable platform combining paper-based test strips and an App-equipped smartphone was further fabricated, making it possible to achieve in-field sensing of the analyte in various matrices.

Herein, the methionine (Met)/N-acetyl-L-cysteine (NAC) templated copper nanoclusters (Met/NAC-Cu NCs) with tunable near-infrared region (NIR) electrochemiluminescence (ECL) emission wavelength was firstly synthesized as emitter for the ultrasensitive detection of matrix metalloproteinase-2 (MMP-2). Significantly, the NAC played the role of template and reductant of cupric to acquire Cu NCs, and the surface defect regulator Met was used to connect NAC through -S-S- bond, which could heighten the surface defect of Cu NCs to continuously regulate the maximum ECL emission by successively controlling the molar ratio of Met and NAC, leading to the ECL emission wavelength of Cu NCs ranged from 680 nm to 750 nm. In addition, a rapid target triggered catalyst hairpin assembly (CHA) recycling amplification strategy was constructed through orderly and equidistantly arranging hairpin to increase its local concentration, resulting in greatly accelerated signal amplification efficiency and reaction rate. As a proof of concept, based on Met/NAC-Cu NCs as NIR ECL emitter and effective signal amplification tactic, a super-sensitive ECL biosensor was fabricated to detect target MMP-2 with the detection limit (LOD) as low as 1.65 fg/mL and successfully utilized for detecting of MMP-2 that from Hela and MCF-7 cancer cells. This research provided a wonderful avenue for regulating the optical performance of metal nanoclusters-based ECL emitters, and the developed neoteric NIR ECL emitter with the merits of less photochemical damage and deeper tissue penetration exhibited great potential in ultrasensitive biosensing and high-definition ECL imaging.

Given the targeted binding ability and cleavage activity of the emerging CRISPR/Cas12a assay which transduces the target into its cleavage activity exhibited broadly prospective applications in integrated sensing and actuating system. Here, we elaborated a universal approach to quickly activate CRISPR/Cas12a for low-abundance biomarker detection based on the amplification strategy of a target-induced spherical nucleic acid enzyme (SNAzyme) network that could accelerate the output of activators. Specifically, multifunctional Y-shaped probes and hairpin probes (HPs, which contained the specific sequence of the activators of CRISPR/Cas12a and the substrate chain of DNAzyme) were rationally designed to construct SNAzyme. Target recognition induced disassembly of the Y-shaped probes, which released DNAzyme strands to active DNAzyme and accompanied by SNAzyme self-assembly into SNAzyme network. Interestingly, compared with randomly dispersed SNAzyme, the reaction kinetics of the SNAzyme network enhanced 1.6 times in response to Α-methyl acyl-CoA racemase (AMACR, a biomarker for prostate cancer), which was attributed to the promoted catalytic efficiency of DNAzyme by the confined SNAzyme network. Benefiting from these, the prepared biosensor based on electrochemiluminescence (ECL) platform by loading AuAg nanoclusters (AuAgNCs) into metal-organic framework-5 (MOF-5) exhibited satisfying detection performance for AMACR with a wide linear range (0.001 μg/mL to 100 μg/mL) and a low detection limit (1.0 × 10-4 μg/mL, which exhibited significant potential in clinical diagnoses.

Neonicotinoids such as thiamethoxam (TMX) were widely used in agricultural production and tended to accumulate in the environment, potentially harming human and ecosystem health. To enable widespread monitoring of TMX residues, it was essential to design a reliable and sensitive detection method. Here, we developed a novel smartphone-enablled molecularly imprinted ratiometric fluorescence sensing system for selective on-site detection of TMX. It was based on blue-emission carbon dots (CDs) wrapped with a molecularly imprinted layer (B-CDs@MIPs), which provided the response signal, while red-emission CDs (R-CDs) served as an internal reference. The fluorescence signal ratio of the sensor increased with the TMX concentration, resulting in an obvious fluorescence color change from red to blue. The sensor exhibited a satisfactory limit of detection (LOD) of 13.5 nM in fluorescence analysis while LOD of 70.1 nM in visual determination. In addition, the sensing system was validated using food and environment samples, exhibiting recoveries from 91.40% to 105.7%, indicating excellent reliability for TMX detection in actual samples. Thus, the sensing system developed in this study offered promising prospects for visual detection of pesticide residues in complex environmental samples.

A microneedle (MN) sensor coated with a sensing composite material was proposed for measuring glucose concentrations in interstitial fluid (ISF). The sensing composite material was prepared by blending a polymer containing glucose-responsive phenylboronic acid (PBA) moieties (i.e., polystyrene-block-poly(acrylic acid-co-acrylamidophenylboronic acid)) with conductive carbon nanotubes (CNTs). The polymer exhibited reversible swelling behavior in response to glucose concentrations, which influenced the distribution of the embedded CNTs, resulting in sensitive variations in electrical percolation, even when coated onto a confined surface of the MN in the sensor. We varied the ratio of PBA moieties and the loading amount of CNTs in the sensing composite material of the MN sensor and tested them in vitro using an ISF-mimicking gel with physiological glucose concentrations to determine the optimal sensitivity conditions. When tested in animal models with varying blood glucose concentrations, the MN sensor coated with the selected sensing material exhibited a strong correlation between the measured electrical currents and blood glucose concentrations, showing accuracy comparable to that of a glucometer in clinical use.

Real-time studies of biomarkers for neurological disorders present significant opportunities for diagnosing and treating related diseases, and fluorescent probes offer a promising approach to brain imaging. However, intracerebral fluorescence imaging is often limited by blood-brain barrier permeability and penetration depth. Moreover, only very few probes have rapid intracerebral metabolic properties, which are critical for in vivo imaging. Here, we developed a novel class of fluorescent dyes with two-photon excitation and near-infrared (NIR) emission (920/705 nm). The representative WAPP-4 probe exhibits a large Stokes shift (Δλ = 324 nm in ethanol) and excellent blood-brain barrier permeability. Notably, using WAPP-4, we achieved in vivo 3D dynamic imaging of Aβ plaques in the brains of living mice with Alzheimer's disease (AD). In addition, super-resolution imaging showed that WAPP-4 could effectively characterize the distribution and shape of Aβ plaques in isolated brain slices. This study demonstrates that newly developed fluorescent dyes with large Stokes shifts and blood-brain barrier permeability enable real-time imaging of amyloid plaques, which will contribute to the development of other valuable tools for near-infrared imaging and super-resolution imaging in the brain.

Compared with single signal readout, dual-signal readout commendably corrects the impact of systematic or background error, achieving more accurate results for the diagnosis of many diseases. This work aimed to design and prepare dual-emissive fluorescent probes for the construction of ratiometric fluorescence biosensors to detect liver disease biomarkers. Sodium alginate (SA) with numerous potential sub-fluorophores and active sites and 4,4',4'',4'''-(porphine-5,10,15,20-tetrayl) tetrakis (benzoic acid) (TCPP) with macrocyclic conjugated structures were introduced to prepare the carbonized polymer dots (CPDs) with red/blue dual emission based on the cross-linking enhanced emission (CEE) effect and the luminescence of macrocyclic conjugated structures. The ratiometric fluorescence sensing systems were constructed by integrating the specific response of CPDs to Cu2+ and the affinity difference of Cu2+ to substrates or products of enzymes. The sensing systems, CPDs/Cu2+/PPi and CPDs/Cu2+/BTCh, were designed to detect liver disease biomarkers, alkaline phosphatase (ALP) and butyrylcholinesterase (BChE), respectively. The limit of detection for ALP and BChE was 0.35 U/L and 0.19 U/L, respectively. The proposed sensors were successfully applied to human serum samples from different health stages with satisfactory recoveries. These results demonstrate the successful design of a novel dual-emissive fluorescent probe and provide a feasible strategy for clinical detection.

In view of the importance of quantification of neuron-specific enolase (NSE), an electrochemical NSE immunosensor was developed. The sandwich voltammetric immunosensor utilized vinyl-functionalized crystalline covalent organic framework (COFTAPT-Dva) modified electrode to load lots of Ab1 via thiol-ene "click" reaction as matrix. A crystalline cationic EB-COF:Br was used to load Au nanoparticles (AuNPs) and H3[PMo12O40] (PMo12) as immunoprobe. The AuNPs with the size of about 30 nm were firstly grown on EB-COF:Br and then a large number of electroactive PMo12 were uniformly assembled on AuNPs/EB-COF:Br via ion exchanging reaction. The AuNPs not only facilitated the bonding of Ab2 based on Au-S bond, but also improved performance of Ab2/AuNPs/EB-COF:PMo12 immunoprobe. The sensitivity of sandwich electrochemical immunosensor could be primarily amplified based on loaded abundant PMo12. Secondary sensitivity amplification of immunosensor could be achieved by using PMo12 to catalyze ascorbic acid. The linear range of sandwich voltammetric immunosensor based on current change of differential pulse voltammetry is 500 ± 36 fg mL-1 - 100 ± 8 ng mL-1. Thanks to the dual sensitivity amplification strategy, the sensitivity is as high as 54.06 ± 3.2 μA cm-2/lg(cNSE/ng mL-1), and the detection limit is as low as 166 ± 10.8 fg mL-1. It proves that it is completely feasible to amplify sensitivity of sandwich voltammetric immunosensors using polyoxometalate-COF and its catalytic substrate.

The generation of guaiacol by Alicyclobacillus acidoterrestris (A. acidoterrestris) in fruit juices negatively affects public health and causes severe environmental pollution. Therefore, the sensitive detection and efficient degradation of guaiacol in real samples are crucial. Here, we develop an electrochemical sensor utilizing a copper single-atom nanozyme (CuN4-G) to detect and degrade guaiacol at the picomolar level. Density functional theory (DFT) calculations verify that the bonding electron coupling effect in the CuN4-G facilitates rapid electron transfer, enhances electrical conductivity, and provides abundant active sites, thereby leading to exceptional catalytic performance. Moreover, CuN4-G demonstrates a Km value similar to that of natural laccase but a higher Vmax, highlighting its potential as a highly efficient biocatalyst. The CuN4-G-based electrochemical sensor achieves a detection from 5 to 50,000 pM for guaiacol, with a 1.2 pM (S/N = 3) detection limit. Additionally, CuN4-G-modified electrodes display high selectivity and excellent stability. CuN4-G nanozyme can keep its activity in conditions of pH (3-9), temperature (30-90 °C), ionic strength (0-400 mM), and organic solvent (0-50% (v/v)), overcoming the deficiencies of natural enzymes. Furthermore, our electrochemical sensor can not only accurately detect guaiacol, but also degrade it in actual fruit juice samples infected by A. acidoterrestris, demonstrating its potential applications in food and environmental monitoring.

Acetaldehyde, which is an intermediate product of alcohol metabolism, is known to induce symptoms, including alcohol flushing, vomiting, and headaches in humans. Therefore, real-time monitoring of acetaldehyde levels is crucial to mitigating these health issues. However, current methods for detecting low-concentration gases necessitate the use of complex measurement equipment. In this study, we developed a low-cost, low-detection-limit, enzyme-based electrochemical biosensor for acetaldehyde gas detection that does not require sophisticated equipment. The sensor was constructed by screen-printing electrodes onto a porous polyimide film, using grafted MgO-templated carbon (GMgOC) as working electrode material, carbon for the counter electrode, and silver/silver chloride for the reference electrode. Pyrroloquinoline-quinone-dependent aldehyde dehydrogenase was immobilized on the working electrode, and a chamber was attached to the electrode chip and filled with 1-methoxy-5-methylphenazinium methyl sulfate solution. The sensor can be used to measure acetaldehyde gas concentrations from 0.02 to 0.1 ppm, making it suitable for monitoring human skin gas. This low detection limit was achieved by delivering the analyte through the porous polyimide film on which the electrodes were printed and accumulating acetaldehyde in the mesoporous GMgOC of the working electrode. This mechanism suggests that this sensor design can be adapted to develop other low-detection limit gas sensors, such as those for screening skin gas biomarkers.

The development of innovative and efficient strategy is of paramount importance for near-infrared (NIR) electrochemiluminescence (ECL) sensing, which can substantially promote ECL detection in a wide range of situations. Herein, the inner filter effect (IFE) strategy was designed to construct an ultrasensitive NIR ECL biosensor based on the well-matched AgBr nanocrystals (NCs) decorated nitrogen-doped Ti3C2 MXene nanocomposites (AgBr-N-Ti3C2) and hydrated defective tungsten oxide nanosheets (dWO3•H2O). Specifically, the AgBr-N-Ti3C2 nanocomposites displayed extremely effective NIR ECL emission because N-doping could accelerate electron transfer and boost the red-shift of the ECL spectrum. The nonmetallic plasmon dWO3•H2O was used as an absorber due to its facile tuning of the spectra overlap and higher molar extinction coefficients. Time-resolved emission decay curves proved that the decreased ECL intensity was ascribed to the IFE-based steady quenching mechanism. With the support of tetracycline (TC) aptamer and the complementary DNA chain, the fabricated NIR ECL-IFE biosensor performed a wide linear range of 100 nM ∼ 10 fM with a low detection limit of 2.2 fM (S/N = 3), and it exhibited excellent stability, sensitivity, and reproducibility, so as to be applied to real samples. This strategy opens a new avenue to constructing an efficient NIR ECL-IFE system and shows excellent practical potential in actual sample analysis.

The sensitive and accurate detection of glucose and lactate is essential for early diagnosis and effective management of diabetes complications. Herein, a 3D Printed ECL imaging system integrated with a Smartphone has been demonstrated to advance the traditional ECL to make a portable, affordable, and turnkey point-of-care solution to detect various human metabolites. A universal cross-platform application was introduced for analyzing ECL emitted signals to automate the whole detection process for real-time monitoring and rapid diagnostics. The developed ECL system was successfully applied and validated for detecting glucose and lactate using a single-electrode ECL biosensing platform. For glucose and lactate detection, the device showed a linear range from 0.1 mM to 1 mM and 0.1 mM-4 mM with a detection limit (LoD) of 0.04 mM and 0.1 mM, and a quantification limit (LoQ) of 0.142 mM and 0.342 mM, respectively. The developed method was evaluated for device stability, accuracy, interference, and real sample analysis. Furthermore, to assist in selecting the accurate and economic ECL sensing platform, SE-ECL devices fabricated via different fabrication approaches such as Laser-Induced Graphene, Screen Printing, and 3D Printing are studied for the conductivity of electrode and its significance on ECL signal. It was observed that emitted ECL signal is independent of the electrical conductivity for the same concentration of analytes. The findings suggested that the developed miniaturized point-of-care ECL platform would be a comprehensive and integrated solution for detecting other human metabolites and have the potential to be used in clinical applications.

Since microRNAs (miRNAs) are predictors of tumorigenesis, accurate identification and quantification of miRNAs with highly similar sequences are expected to reflect tumor diagnosis and treatment. In this study, a highly selective and sensitive electrochemiluminescence (ECL) biosensor was constructed for miRNAs determination based on Y-shaped junction structure equipped with locked nucleic acids (LNA), graphene oxide-based nanocomposite to enrich luminophores, and conductive matrix. Specifically, two LNA-modified probes were designed for specific miRNA recognition, that is, a dual-amine functionalized hairpin capture probe and a signal probe. A Y-shaped DNA junction structure was generated on the electrode surface upon miRNA hybridizing across the two branches, so as to enhance the selectivity. Carbon quantum dots-polyethylene imine-graphene oxide (CQDs-PEI-GO) nanocomposites were developed to enrich luminophores CQDs, and thus enhancing the ECL intensity. For indirect signal amplification, an electrochemically activated poly(2-aminoterephthalic acid) (ATA) film decorated with gold nanoparticles was prepared on electrode as an effective matrix to accelerate the electron transfer. The fabricated ECL biosensor achieved sensitive determination of miRNA-222 with a limit-of-detection (LOD) as low as 1.95 fM (S/N = 3). Notably, Y-shaped junction structures equipped with LNA probes endowed ECL biosensor with salient single-base discrimination ability and anti-interference capacity. Overall, the proposed Y-shaped ECL biosensor has considerable promise for clinical biomarker determination.

Heparan sulfate (HS) plays a critical role in various biological processes as a vital component of the extracellular matrix. In this study, we synthesized three fluorescent probes (1-3) comprising Arg-rich peptides as HS receptors and a fluorophore capable of exhibiting red-shifted emissions upon aggregation. All three probes demonstrated ratiometric responses to HS and heparin in aqueous solutions. Remarkably, probe 3 exhibited a unique ratiometric response to HS in both aqueous solutions at physiological pH and HS proteoglycans on live cells. Probe 3 displayed exceptional sensing properties, including high biocompatibility, water solubility, visible light excitation, a large Stokes shift for ratiometric detection and remarkable selectivity and sensitivity for HS (with a low limit of detection: 720 pM). Binding mode studies unveiled the crucial role of charge interactions between probe 3 and negatively charged HS sugar units. Upon binding, the fluorophore segments of the probes overlapped, inducing green and red emission changes through restricted intramolecular rotation of the fluorophore moiety. Importantly, probe 3 was effectively employed to quantify the reduction of HS proteoglycan levels in live cells by inhibiting HS sulfation using siRNA and an inhibitor. It successfully detected decreased HS levels in cells treated with doxorubicin and irradiation, consistent with results obtained from western blot and immunofluorescence assays. This study presents the first ratiometric fluorescent probe capable of quantitatively detecting HS levels in aqueous solutions and live cells. The unique properties of peptide-based probe 3 make it a valuable tool for studying HS biology and potentially for diagnostic applications in various biological systems.

This manuscript presents a novel approach to address the challenges of electrode fouling and highly complex electrode nanoarchitecture, which are primary concerns for biosensors operating in real environments. The proposed approach utilizes multiparametric impedance discriminant analysis (MIDA) to obtain a fingerprint of the macromolecular interactions on flat glassy carbon surfaces, achieved through self-organized, drop-cast, receptor-functionalized Au nanocube (AuNC) patterns. Real-time monitoring is combined with singular value decomposition and partial least squares discriminant analysis, which enables selective identification of the analyte from raw impedance data, without the use of electric equivalent circuits. As a proof-of-concept, the authors demonstrate the ability to detect Escherichia coli in real human urine using an aptamer-based biosensor that targets RNA polymerase. This is significant, as uropathogenic E. coli is a difficult-to-treat pathogen that is responsible for the majority of hospital-acquired urinary tract infection cases. The proposed approach offers a limit of detection of 11.3 CFU/mL for the uropathogenic E. coli strain No. 57, an analytical range in all studied concentrations (up to 105 CFU/mL), without the use of antifouling strategies, yet not being specific vs other E.coli strain studied (BL21(DE3)). The MIDA approach allowed to identify negative overpotentials (-0.35 to -0.10 V vs Ag/AgCl) as most suitable for the analysis, offering over 80% sensitivity and accuracy, and the measurement was carried out in just 2 min. Moreover, this approach is scalable and can be applied to other biosensor platforms.

Herein, the short peptide N-fluorenemethoxycarbonyl diphenylalanine (Fmoc-FF) was used to immobilize both diallyl viologen (DAV) and the enzyme formate dehydrogenase (FDH) to form Fmoc-FF/DAV/FDH supramolecular hydrogel films on an electrode surface by a simple solvent-controlled self-assembly method. The DAV component in the films exhibited multiple properties, such as electrochromism and electrofluorochromism, and acted as an electrochemical mediator. A high efficiency of bioelectrocatalytic reduction of CO2 to formate (HCOO-) was obtained by the natural FDH enzyme and the artificial coenzyme factor DAV both immobilized in the same films. The supramolecular hydrogel films with CO2, voltage and light as stimulating factors and current, fluorescence and UV-vis extinction as responsive signals, were further applied for the construction of complex biomolecular logic systems and information encryption. A 3-input/7-output biomolecular logic gate and several logic devices, including an encoder/decoder, a parity checker, and a keypad lock, were constructed. Especially, the biomolecular keypad lock with 3 types of signals as outputs significantly enhanced the security level of information encryption. In this work, a supramolecular self-assembly interface was simply fabricated with complex biomolecular computational functions using immobilized molecules as the computational core, greatly broadening the application range of supramolecular hydrogel films and providing an idea for new designs of bioinformation encryption through the use of a simple film system.

Nanomaterials with intrinsic enzyme activity, referred to as nanozymes, have attracted substantial attention in recent years. Among them, phosphatase-mimicking nanozymes have become an increasingly important focus for future research, considering that phosphatase is not only one of key enzymes for phosphorous metabolism, which is essential for many biological processes (e.g., cellular regulation and signaling), but also one of extensively used biocatalytic labels in the enzyme-linked assays as well as a powerful tool enzyme in molecular biology laboratories. Nevertheless, compared with extensive oxidoreductase-mimicking nanozymes, there are a very limited number of nanozymes with phosphatase-like activity have been explored at present. The increasing demand of complex and individualized phosphatase-involved catalytic behaviors is pushing the development of more advanced phosphatase-mimicking nanozymes. Thus, we present an overview on recently reported phosphatase-like nanozymes, providing guidelines and new insights for designing more advanced phosphatase-mimicking nanozyme with superior properties.

The escalating number of patients affected by various diseases, such as gout, attributed to abnormal uric acid (UA) concentrations in body fluids, has underscored the need for rapid, efficient, highly sensitive, and stable UA detection methods and sensors. Optical sensors have garnered significant attention due to their simplicity, cost-effectiveness, and resistance to electromagnetic interference. Notably, research efforts have been directed towards UA on-site detection, enabling daily monitoring at home and facilitating rapid disease screening in the community. This review aims to systematically categorize and provide detailed descriptions of the notable achievements and emerging technologies in UA optical sensors over the past five years. The review highlights the advantages of each sensor while also identifying their limitations in on-site applications. Furthermore, recent progress in instrumentation and the application of UA on-site detection in body fluids is discussed, along with the existing challenges and prospects for future development. The review serves as an informative resource, offering technical insights and promising directions for future research in the design and application of on-site optical sensors for UA detection.

Monitoring astronauts' health during space missions poses many challenges, including rapid assessment of crew health conditions. Sensitive genetic diagnostics are crucial for examining crew members and the spacecraft environment. CRISPR-Cas12a, coupled with isothermal amplification, has proven to be a promising biosensing system for rapid, on-site detection of genomic targets. However, the efficiency and sensitivity of CRISPR-based diagnostics have never been tested in microgravity. We tested the use of recombinase polymerase amplification (RPA) coupled with the collateral cleavage activity of Cas12a for genetic diagnostics onboard the International Space Station. We explored the detection sensitivity of amplified and unamplified target DNA. By coupling RPA with Cas12a, we identified targets in attomolar concentrations. We further assessed the reactions' stability following long-term storage. Our results demonstrate that CRISPR-based detection is a powerful tool for on-site genetic diagnostics in microgravity, and can be further utilized for long-term space endeavors to improve astronauts' health and well-being.

Polymerase chain reaction (PCR) in small fluidic systems not only improves speed and sensitivity of deoxyribonucleic acid (DNA) amplification but also achieves high-throughput quantitative analyses. However, air bubble trapping and growth during PCR has been considered as a critical problem since it causes the failure of DNA amplification. Here we report bubble-free diatom PCR by exploiting a hierarchically porous silica structure of single-celled algae. We show that femtoliters of PCR solution can be spontaneously loaded into the diatom interior without air bubble trapping due to the surface hydrophilicity and pore structure of the diatom. We discover that a large pressure gradient between air bubbles and nanopores rapidly removes residual air bubbles through the periodically arrayed nanopores during thermal cycling. We demonstrate the DNA amplification by diatom PCR without air bubble trapping and growth. Finally, we successfully detect DNA fragments of SARS-CoV-2 with as low as 10 copies/μl by devising a microfluidic device integrated with diatoms assembly. We believe that our work can be applied to many PCR applications for innovative molecular diagnostics and provides new opportunities for naturally abundant diatoms to create innovative biomaterials in real-world applications.

With extensive and widespread uses of miniaturized and intelligent wearable devices, continuously monitoring subtle spatial and temporal changes in human physiological states becomes crucial for daily healthcare and professional medical diagnosis. Wearable acoustical sensors and related monitoring systems can be comfortably applied onto human body with a distinctive function of non-invasive detection. This paper reviews recent advances in wearable acoustical sensors for medical applications. Structural designs and characteristics of the structural components of wearable electronics, including piezoelectric and capacitive micromachined ultrasonic transducer (i.e., pMUT and cMUT), surface acoustic wave sensors (SAW) and triboelectric nanogenerators (TENGs) are discussed, along with their fabrication techniques and manufacturing processes. Diagnostic applications of these wearable sensors for detection of biomarkers or bioreceptors and diagnostic imaging have further been discussed. Finally, main challenges and future research directions in these fields are highlighted.

Electrochemical nucleic acid biosensor has demonstrated great promise in clinical diagnostic tests, mainly because of its flexibility, high efficiency, low cost, and easy integration for analytical applications. Numerous nucleic acid hybridization-based strategies have been developed for the design and construction of novel electrochemical biosensors for diagnosing genetic-related diseases. This review describes the advances, challenges, and prospects of electrochemical nucleic acid biosensors for mobile molecular diagnosis. Specifically, the basic principles, sensing elements, applications in diagnosis of cancer and infectious diseases, integration with microfluidic technology and commercialization are mainly included in this review, aiming to provide new insights and directions for the future development of electrochemical nucleic acid biosensors.

Deoxyribonucleic acid (DNA) is a carrier of genetic information. DNA hybridization is characterized by predictability, diversity, and specificity owing to the strict complementary base-pairing assembly mode, which stimulates the use of DNA to build a variety of nanomachines, including DNA tweezers, motors, walkers, and robots. DNA nanomachines have become prevalent for signal amplification and transformation in the field of biosensing, providing a new method for constructing highly sensitive sensing analysis strategies. DNA tweezers have exhibited unique advantages in biosensing applications owing to their simple structures and fast responses. The two-state conformation of DNA tweezers, the open and closed states, enable them to open and close autonomously after stimulation, thus facilitating the quick detection of corresponding signal changes of different targets. This review discusses the recent progress in the application of DNA nanotweezers in the field of biosensing, and the trends in their development for application in the field of biosensing are summarized.

Following the recent pandemic and with the emergence of cell-free nucleic acids in liquid biopsies as promising biomarkers for a broad range of pathologies, there is an increasing demand for a new generation of nucleic acid tests, with a particular focus on cost-effective, highly sensitive and specific biosensors. Easily miniaturized electrochemical sensors show the greatest promise and most typically rely on the chemical functionalization of conductive materials or electrodes with sequence-specific hybridization probes made of standard oligonucleotides (DNA or RNA) or synthetic analogues (e.g. Peptide Nucleic Acids or PNAs). The robustness of such sensors is mostly influenced by the ability to control the density and orientation of the probe at the surface of the electrode, making the chemistry used for this immobilization a key parameter. This exhaustive review will cover the various strategies to immobilize nucleic acid probes onto different solid electrode materials. Both physical and chemical immobilization techniques will be presented. Their applicability to specific electrode materials and surfaces will also be discussed as well as strategies for passivation of the electrode surface as a way of preventing electrode fouling and reducing nonspecific binding.

Miniaturization is the trend to manufacture ever smaller devices and this process requires knowledge, experience, understanding of materials, manufacturing techniques and scaling laws. The fabrication techniques used in semiconductor industry deliver an exceptionally high yield of devices and provide a well-established platform. Today, these miniaturized devices are manufactured with high reproducibility, design flexibility, scalability and multiplexed features to be used in several applications including micro-, nano-fluidics, implantable chips, diagnostics/biosensors and neural probes. We here provide a review on the microfabricated devices used for biology driven science. We will describe the ubiquity of the use of micro-nanofabrication techniques in biology and biotechnology through the fabrication of high-aspect-ratio devices for cell sensing applications, intracellular devices, probes developed for neuroscience-neurotechnology and biosensing of the certain biomarkers. Recently, the research on micro and nanodevices for biology has been progressing rapidly. While the understanding of the unknown biological fields -such as human brain- has been requiring more research with advanced materials and devices, the development protocols of desired devices has been advancing in parallel, which finally meets with some of the requirements of biological sciences. This is a very exciting field and we aim to highlight the impact of micro-nanotechnologies that can shed light on complex biological questions and needs.

Bioelectrical interfaces made of living electroactive bacteria (EAB) provide a unique opportunity to bridge biotic and abiotic systems, enabling the reprogramming of electrochemical biosensing. To develop these biosensors, principles from synthetic biology and electrode materials are being combined to engineer EAB as dynamic and responsive transducers with emerging, programmable functionalities. This review discusses the bioengineering of EAB to design active sensing parts and electrically connective interfaces on electrodes, which can be applied to construct smart electrochemical biosensors. In detail, by revisiting the electron transfer mechanism of electroactive microorganisms, engineering strategies of EAB cells for biotargets recognition, sensing circuit construction, and electrical signal routing, engineered EAB have demonstrated impressive capabilities in designing active sensing elements and developing electrically conductive interfaces on electrodes. Thus, integration of engineered EAB into electrochemical biosensors presents a promising avenue for advancing bioelectronics research. These hybridized systems equipped with engineered EAB can promote the field of electrochemical biosensing, with applications in environmental monitoring, health monitoring, green manufacturing, and other analytical fields. Finally, this review considers the prospects and challenges of the development of EAB-based electrochemical biosensors, identifying potential future applications.

Epidermal microfluidic devices are continuously being developed for efficient sweat collection and sweat rate detection. However, most microfluidic designs ignore the use of airtight/adhesive substrate will block the natural perspiration of the covered sweat pores, which will seriously affect normal sweat production and long-term wearable comfort. Herein, we present a Janus textile-embedded microfluidic sensor platform with high breathability and directional sweat permeability for synchronous sweat rate and total electrolyte concentration detection. The device consists of a hollowed-out serpentine microchannel with interdigital electrodes and Janus textile. The dual-mode signal of the sweat rate (0.2-4.0 μL min-1) and total ionic charge concentration (10-200 mmol L-1) can be obtained synchronously by decoupling conductance step signals generated when sweat flows through alternating interdigitated spokes at equal intervals in the microchannel. Meanwhile, the hollowed-out microchannel structure significantly reduces the coverage area of the sensor on the skin, and the Janus textile-embedded device ensures a comfortable skin/device interface (fewer sweat pores are blocked) and improves breathability (503.15 g m-2 d-1) and sweat permeability (directional liquid transportation) during long-term monitoring. This device is washable and reusable, which shows the potential to integrate with clothing and smart textile, and thus facilitate the practicality of wearable sweat sensors for personalized healthcare.

Switchable fluorescent proteins, for which fluorescence can be switched ON and OFF, are widely used for molecule tracking and super resolution imaging. However, the robust use of the switchable fluorescent proteins is still limited as either the switching is not repeatable, or such switching requires irradiation with coupled lasers of different wavelengths. Herein, we report an electrochemical approach to reversible fluorescence switching for enhanced green fluorescent proteins (EGFP) on indium tin oxide coated glass. Our results demonstrate that negative and positive electrochemical potentials can efficiently switch the fluorescent proteins between the dim (OFF) and bright (ON) states at the single molecule level. The electrochemical fluorescence switching is fast, reversible, and may be performed up to hundreds of cycles before photobleaching occurs. These findings highlight that this method of electrochemical fluorescence switching can be incorporated into advanced fluorescence microscopy.

Biotic and abiotic stresses are well known to increase the emission of volatile organic compounds (VOCs) from plants. The analysis of VOCs emissions from plants enables timely diagnostic of plant diseases, which is critical for prompting sustainable agriculture. Previous studies have predominantly focused on the utilization of commercially available devices, such as electronic noses, for diagnosing plant diseases. However, recent advancements in nanomaterials research have significantly contributed to the development of novel VOCs sensors featuring exceptional sensitivity and selectivity. This comprehensive review presents a systematic analysis of VOCs monitoring technologies for plant diseases diagnosis, providing insights into their distinct advantages and limitations. Special emphasis is placed on custom-made VOCs sensors, with detailed discussions on their design, working principles, and detection performance. It is noteworthy that the application of VOCs monitoring technologies in the diagnostic process of plant diseases is still in its emerging stage, and several critical challenges demand attention and improvement. Specifically, the identification of specific stress factors using a single VOC sensor remains a formidable task, while environmental factors like humidity can potentially interfere with sensor readings, leading to inaccuracies. Future advancements should primarily focus on addressing these challenges to enhance the overall efficacy and reliability of VOCs monitoring technologies in the field of plant disease diagnosis.

The growing interest in flexible electronics for physiological monitoring, particularly using flexible pressure sensors for cardiovascular pulse waveforms monitoring, has potential applications in cuffless blood pressure measurement and early diagnosis of cardiovascular disease. High sensitivity, fast response time, good pressure resolution and a high signal-to-noise ratio are essential for effective pulse waveform detection. This review focuses on flexible capacitive and piezoresistive pressure sensors, which have seen significant enhancements due to their simple operation, superior performance, wide range of materials, and easy fabrication. The comparison of sensing methods for acquiring pulse waveforms from the wrist artery, device integration configurations, high-quality pulse waveforms collection, and performance analysis of capacitive and piezoresistive sensors are discussed. The review also covers the use of machine learning for analyzing pulse waveforms for cardiovascular disease diagnosis and cuff-less blood pressure monitoring. Lastly, it provides perspectives on current challenges and further advancements in the field.

Glucose is the primary energy source of human cells. Therefore, monitoring glucose inside microphysiological systems (MPS) provides valuable information on the viability and metabolic state of the cultured cells. However, continuous glucose monitoring inside MPS is challenging due to a lack of suitable miniaturized sensors. Here we present an enzymatic, optical glucose sensor element for measurement inside microfluidic systems. The miniaturized glucose sensor (Ø 1 mm) is fabricated together with a reference oxygen sensor onto biocompatible, pressure-sensitive adhesive tape for easy integration inside microfluidic systems. Furthermore, the proposed microfluidic system can be used as plug and play sensor system with existing MPS. It was characterized under cell culture conditions (37 °C and pH 7.4) for five days, exhibiting minor drift (3% day-1). The influence of further cell culture parameters like oxygen concentration, pH, flow rate, and sterilization methods was investigated. The plug-and-play system was used for at-line measurements of glucose levels in (static) cell culture and achieved good agreement with a commercially available glucose sensor. In conclusion, we developed an optical glucose sensor element that can be easily integrated in microfluidic systems and is able to perform stable glucose measurements under cell culture conditions.

The detection of SARS-CoV-2 infection is crucial for effective prevention and surveillance of COVID-19. In this study, we report the development of a novel detection assay named CENSOR that enables sensitive and specific detection of SARS-CoV-2 RNA using a plasmonic chiral biosensor in combination with CRISPR-Cas13a. The chiral biosensor was designed by assembling gold nanorods (AuNR) into three-dimensional plasmonic architectures of controllable chirality on a DNA origami template. This modular assembly mode enhances the flexibility and adaptability of the sensor, thereby improving its universality as a sensing platform. In the presence of SARS-CoV-2 RNA, the CRISPR-Cas13a enzyme triggers collateral cleavage of RNA molecules, resulting in a differential chiral signal readout by the biosensor compared to when there are no RNA targets present. Notably, even subtle variations in the concentration of SARS-CoV-2 RNA can provoke significant changes in chiral signals after preamplification of RNA targets (calculated LOD: 0.133 aM), which establishes the foundation for quantitative detection. Furthermore, CENSOR demonstrated high sensitivity and accuracy in detecting SARS-CoV-2 RNA from clinical samples, suggesting its potential application in clinical settings for viral detection beyond SARS-CoV-2.

Exfoliated tumor cells are integral to malignant tumors diagnosis. The process of clinical cytology of exfoliation involves several complex steps that require at least two days of preparation. Here, we develop a balanced-etching visual kit based on concentration differences of Glutathione/Glucose (GSH/Glu) to distinguish normal from exfoliated tumor cells rapidly and accurately. The balanced-etching visualization kit can be used to obtain color cards and screen exfoliated tumor cells initially (within 10 min). Furthermore, by utilizing logic gates and machine learning algorithms for RGB extraction of the color card obtained from the kit, accurate screening of exfoliated tumor cells is achieved. Finally, a series of clinical tumor samples, such as urine, pleural fluids, ascites, and gastric fluids, have been validated. With effective experimental methods, accurate disease information, and appropriate therapeutic programs, the novel diagnostic strategy is expected to promote precision medicine.

Incorporating non-invasive biosensing features in organ-on-chip models is of paramount importance for a wider implementation of these advanced in vitro microfluidic platforms. Optical biosensors, based on Bioluminescence Imaging (BLI), enable continuous, non-invasive, and in-situ imaging of cells, tissues or miniaturized organs without the drawbacks of conventional fluorescence imaging. Here, we report the first-of-its-kind integration and optimization of BLI in microfluidic chips, for non-invasive imaging of multiple biological readouts. The cell line HEK293T-GFP was engineered to express NanoLuc® luciferase under the control of a constitutive promoter and were cultured on-chip in 3D, in standard ECM-like hydrogels, to assess optimal cell detection conditions. Using real-time in-vitro dual-color microscopy, Bioluminescence (BL) and fluorescence (FL) were detectable using distinct imaging setups. Detection of the bioluminescent signals were observed at single cell resolution on-chip 20 min post-addition of Furimazine substrate and under perfusion. All hydrogels enabled BLI with higher signal-to-noise ratios as compared to fluorescence. For instance, agarose gels showed a ∼5-fold greater BL signal over background after injection of the substrate as compared to the FL signal. The use of BLI with microfluidic chip technologies opens up the potential for simultaneous in situ detection with continuous monitoring of multicolor cell reporters. Moreover, this can be achieved in a non-invasive manner. BL has great promise as a highly desirable biosensor for studying organ-on-chip platforms.

Modification-free biosensing with high specificity and sensitivity is essential for miniaturized, online, integrated, and rapid, or even real-time molecular analyses. However, most optical biosensors are based on surface pre-modification or fluorescent labeling, and have either low sensitivity or low quality factor (Q). To address these difficulties, in this study, an optical sensor prototype was developed with a microbubble optofluidic channel integrated inside a Fabry-Pérot cavity to three-dimensionally tailor the intra-cavity light field via the intra-cavity lensing (microbubble) configuration. A high Q-factor (∼105), small mode volume, and high light energy density were experimentally achieved with this "stereo-sensor" while maintaining an ultrahigh refractive index (RI) sensitivity (679 nm/RIU) and ultra-small RI resolution (∼10-7 RIU at 950 nm). Moreover, specific detection of very low concentration of biomolecules (5 fg/mL for human IgG and 0.5 pg/mL for human serum albumin (HSA)) and wide range of protein concentrations (e.g., fg/mL-ng/mL for human IgG and pg/mL-ng/mL for HSA) without probe pre-modification were achieved owing to the RI change specifically associated with the probe-target binding and the corresponding bio-macromolecular conformation change. This modification-free stereosensing scenario is applicable to continuous, real-time, and multiplexed operations, thus showing potential for online, integrated, dynamic, biomolecular analyses in vitro or in vivo, such as the dynamic metabolic analysis of single cells or organoids and point-of-care tests.

High entropy (HE) compounds with chemically disordered multi-cation structures have become a hot research topic because of their fascinating "cocktail effect". However, high entropy effect with the efficient photoelectric response has not been reported for photoelectrochemical (PEC) immunoassays. Herein, an innovative PEC immunoassay for the sensitive detection of prostate-specific antigen (PSA) was ingeniously constructed using hollow nanocubic (ZnCdFeMnCu)xS photoactive matrices with high entropic effect via the cation exchange. Initially, a sandwich-type immunoreaction has behaved using dopamine-loaded liposome labeled with anti-PSA secondary antibodies. In the presence of PSA, addition of Triton X-100 caused the liposomal cleavage to release dopamine, which was then detected as a reduced photocurrent on (ZnCdFeMnCu)xS-based photoelectrode. Under optimal condition, the PEC immunoassay showed good photocurrent responses toward target PSA with the dynamic linear range of 0.1-50 ng mL-1 with a limit of detection of 34.1 pg mL-1. Significantly, this system can provide a new platform for the development of PEC immunoassays by coupling with high-entropy photoactive materials.

The search for reliable protein biomarker candidates is critical for early disease detection and treatment. However, current immunoassay technologies are failing to meet increasing demands for sensitivity and multiplexing. Here, the authors have created a highly sensitive protein microarray using the principle of single-molecule counting for signal amplification, capable of simultaneously detecting a panel of cancer biomarkers at sub-pg/mL levels. To enable this amplification strategy, the authors introduce a novel method of protein patterning using photolithography to subdivide addressable arrays of capture antibody spots into hundreds of thousands of individual microwells. This allows for the total sensor area to be miniaturized, increasing the total possible multiplex capacity. With the immunoassay realized on a standard 75x25 mm form factor glass substrate, sample volume consumption is minimized to <10 μL, making the technology highly efficient and cost-effective. Additionally, the authors demonstrate the power of their technology by measuring six secretory factors related to glioma tumor progression in a cohort of mice. This highly sensitive, sample-sparing multiplex immunoassay paves the way for researchers to track changes in protein profiles over time, leading to earlier disease detection and discovery of more effective treatment using animal models.

Herein, we report a scalable benchtop electrode fabrication method to produce highly sensitive and flexible third-generation fructose dehydrogenase amperometric biosensors based on water-dispersed 0D-nanomaterials. The electrochemical platform was fabricated via Stencil-Printing (StPE) and insulated via xurography. Carbon black (CB) and mesoporous carbon (MS) were employed as 0D-nanomaterials promoting an efficient direct electron transfer (DET) between fructose dehydrogenase (FDH) and the transducer. Both nanomaterials were prepared in water-phase via a sonochemical approach. The nano-StPE exhibited enhanced electrocatalytic currents compared to conventional commercial electrodes. The enzymatic sensors were exploited for the determination of D-fructose in model solutions and various food and biological samples. StPE-CB and StPE-MS integrated biosensors showed appreciable sensitivity (∼150 μA cm-2 mM-1) with μmolar limit of detection (0.35 and 0.16 μM, respectively) and extended linear range (2-500 and 1-250 μM, respectively); the selectivity of the biosensors, ensured by the low working overpotential (+0.15 V), has been also demonstrated. Good accuracy (recoveries between 95 and 116%) and reproducibility (RSD ≤8.6%) were achieved for food and urine samples. The proposed approach because of manufacturing versatility and the electro-catalytic features of the water-nanostructured 0D-NMs opens new paths for affordable and customizable FDH-based bioelectronics.

Diabetic retinopathy (DR) is a debilitating organ manifestation of diabetes. Absent of early diagnosis and intervention, vision tends to drastically and irreversibly decline. Previously, we showed higher vascular endothelial growth factor receptor 2 (VEGFR-2) expression in diabetic microvessels, and the suitability of this molecule as a biomarker for early DR diagnosis. However, a hurdle to translation remained generation of biodegradable nanoprobes that are sufficiently bright for in vivo detection. Here, an adhesive fluorescent nanoprobe with high brightness was developed using biodegradable materials. To achieve that, a fluorophore with bulky hydrophobic groups was encapsulated in the nanoparticles to minimize fluorophore π-π stacking, which diminishes brightness at higher loading contents. The nanoprobe selectively targeted the VEGFR-2 under dynamic flow conditions. Upon systemic injection, the nanoprobes adhered in the retinal microvessels of diabetic mice and were visualized as bright spots in live retinal microscopy. Histology validated the in vivo results and showed binding of the nanoprobes to the microvascular endothelium and firmly adhering leukocytes. Leukocytes were found laden with nanoprobes, indicating the potential for payload transport across the blood-retinal barrier. Our results establish the translational potential of these newly generated nanoprobes in early diagnosis of DR.

Lung cancer remains the leading cause of cancer-related death. In addition to chest X-rays and computerised tomography, the detection of cancer biomarkers serves as an emerging diagnostic tool for lung cancer. This review explores biomarkers including the rat sarcoma gene, the tumour protein 53 gene, the epidermal growth factor receptor, the neuron-specific enolase, the cytokeratin-19 fragment 21-1 and carcinoembryonic antigen as potential indicators of lung cancer. Biosensors, which utilise various transduction techniques, present a promising solution for the detection of lung cancer biomarkers. Therefore, this review also explores the working principles and recent implementations of transducers in the detection of lung cancer biomarkers. The transducing techniques explored include optical techniques, electrochemical techniques and mass-based techniques for detecting biomarkers and cancer-related volatile organic compounds. Graphene has outstanding properties in terms of charge transfer, surface area, thermal conductivity and optical characteristics, on top of allowing easy incorporation of other nanomaterials. Exploiting the collective merits of both graphene and biosensor is an emerging trend, as evidenced by the growing number of studies on graphene-based biosensors for the detection of lung cancer biomarkers. This work provides a comprehensive review of these studies, including information on modification schemes, nanomaterials, amplification strategies, real sample applications, and sensor performance. The paper concludes with a discussion of the challenges and future outlook of lung cancer biosensors, including scalable graphene synthesis, multi-biomarker detection, portability, miniaturisation, financial support, and commercialisation.

Therapeutic monoclonal antibodies (TmAb) have emerged as effective treatments for a number of cancers and autoimmune diseases. However, large interpatient disparities in the pharmacokinetics of TmAb treatment requires close therapeutic drug monitoring (TDM) to optimise dosage for individual patients. Here we demonstrate an approach for achieving rapid, sensitive quantification of two monoclonal antibody therapies using a previously described enzyme switch sensor platform. The enzyme switch sensor consists of a β-lactamase - β-lactamase inhibitor protein (BLA-BLIP) complex with two anti-idiotype binding proteins (Affimer proteins) as recognition elements. The BLA-BLIP sensor was engineered to detect two TmAbs (trastuzumab and ipilimumab) by developing constructs incorporating novel synthetic binding reagents to each of these mAbs. Trastuzumab and ipilimumab were successfully monitored with sub nM sensitivity in up to 1% serum, thus covering the relevant therapeutic range. Despite the modular design, the BLA-BLIP sensor was unsuccessful in detecting two further TmAbs (rituximab and adalimumab), an explanation for which was explored. In conclusion, the BLA-BLIP sensors provide a rapid biosensor for TDM of trastuzumab and ipilimumab with the potential to improve therapy. The sensitivity of this platform alongside its rapid action would be suitable for bedside monitoring in a point-of-care (PoC) setting.

Long-term wearing comfort is essential for future advanced electronic textiles (e-textiles). Herein, we fabricate a skin-comfortable e-textile for long-term wearing experience on human epidermis. Such e-textile was simply fabricated through two different dip coating methods and single-side air plasma treatment, which couples radiative thermal and moisture management for biofluid monitoring. The silk-based substrate with improved optical properties and anisotropic wettability can provide a temperature drop of 1.4 °C under strong sunlight. Moreover, the anisotropic wettability of the e-textile can provide a dryer skin microenvironment by comparing with traditional fabric. The fiber electrodes weaving into the inner side of the substrate can noninvasively monitor multiple sweat biomarkers (i.e., pH, uric acid, and Na+). Such a synergistic strategy may pave a new path to design next-generation e-textiles with significantly improved comfort.

Dual-mode biosensors have gained great attention due to their excellent detection accuracy provided by the mutual verification of output signals. However, current strategies for dual-mode sensing mainly rely on a signal probe exhibiting dual properties that may encounter unreliability. Herein, we report a dual-mode biosensing strategy by modulating the plasmonic and catalytic activities of nanoparticles through a surface growing approach. Ascorbic acid enables the growing of Au shell on Au@Pt NPs to tune both their peroxidase-like activity and plasmonic signal. Enzyme-catalyzed reactions can generate ascorbic acid to modulate the plasmonic and catalytic activities of nanoparticles. Combined with enzyme-linked immunosorbent assay, it enables dual-mode immunoassays of carbofuran with both a colorimetric readout by a spectrometer down to 0.1 ppb and a naked-eye readout of 5 ppb. This dual-mode biosensing technique advantages in tunable sensitivity and robustness, holding promise as an analytical platform for biomedical diagnosis and food safety.

of rare circulating extracellular vesicles (EV) from early cancers or different types of host cells requires extremely sensitive EV sensing technologies. Nanoplasmonic EV sensing technologies have demonstrated good analytical performances, but their sensitivity is often limited by EVs' diffusion to the active sensor surface for specific target EV capture. Here, we developed an advanced plasmonic EV platform with electrokinetically enhanced yields (KeyPLEX). The KeyPLEX system effectively overcomes diffusion-limited reactions with applied electroosmosis and dielectrophoresis forces. These forces bring EVs toward the sensor surface and concentrate them in specific areas. Using the keyPLEX, we showed significant improvements in detection sensitivity by ∼100-fold, leading to the sensitive detection of rare cancer EVs from human plasma samples in 10 min. The keyPLEX system could become a valuable tool for point-of-care rapid EV analysis.

For high-performance dual-photoelectrode assay, developing a pair of photoactive materials with well-matched band structure and the design of a powerful sensing strategy are highly desirable. Herein, the Zn-TBAPy pyrene-based MOF and BiVO4/Ti3C2 Schottky junction were employed as photocathode and photoanode to form an efficient dual-photoelectrode system. The integration of the cascaded hybridization chain reaction (HCR)/DNAzyme-assisted feedback amplification with DNA walker-mediated cycle amplification strategy realizes femtomolar HPV16 dual-photoelectrode bioassay. Through the activation of the HCR cascaded with the DNAzyme system in the presence of HPV16, plentiful HPV16 analogs are generated that leads to exponential positive feedback signal amplification. Meanwhile on the Zn-TBAPy photocathode, the NDNA hybridizes with the bipedal DNA walker followed by circular cleavage by Nb.BbvCI NEase, producing a dramatically enhanced PEC readout. The achieved ultralow detection limit of 0.57 fM and a wide linear range of 10-6 nM-103 nM showcase the excellent performance of the developed dual-photoelectrode system.

In this work, we present the development of an immunosensor for the direct, selective, and sensitive determination of symmetric dimethylarginine (SDMA) in urine, in view of the emerging role of this molecule as a biomarker for renal disease. SDMA is almost completely excreted by the kidneys, hence in renal dysfunction, the excretion is decreased, resulting in accumulation in plasma. Reference values for plasma or serum have already been established in small animal practice. Values < 15 μg/dL are considered normal, 15-19 μg/dL are values of concern, and at values > 20 μg/dL kidney disease is likely. The proposed electrochemical paper-based sensing platform uses anti-SDMA antibodies for targeted detection of SDMA. Quantification is related to a decrease in the signal of a redox indicator due to the formation of an immunocomplex that interferes with electron transfer. Square wave voltammetry measurements showed a linear correlation of the peak decline for 50 nM - 1 μM SDMA with a detection limit of 15 nM. The influence of common physiological interferences caused no significant peak reduction, indicating excellent selectivity. The proposed immunosensor was successfully applied for the quantification of SDMA in human urine of healthy individuals. Surveillance of SDMA concentration in urine could prove to be very valuable in the diagnosis or monitoring of renal disease.

Microfluidics has shown great promise for point-of-care assays due to unique chemical and physical advantages that occur at the micron scale. Furthermore, integration of electrodes into microfluidic systems provides additional capabilities for assay operation and electronic readout. However, while these systems are abundant in biological and biomedical research settings, translation of microfluidic devices with embedded electrodes are limited. In part, this is due to the reliance on expensive, inaccessible, and laborious microfabrication techniques. Although innovative prior work has simplified microfluidic fabrication or inexpensively patterned electrodes, low-cost, accessible, and robust methods to incorporate all these elements are lacking. Here, we present MINX, a low-cost <1 USD and rapid (∼minutes) fabrication technique to manufacture microfluidic device with embedded electrodes. We characterize the structures created using MINX, and then demonstrate the utility of the approach by using MINX to implement an electrochemical bead-based biomarker detection assay. We show that the MINX technique enables the scalable, inexpensive fabrication of microfluidic devices with electronic sensors using widely accessible desktop machines and low-cost materials.

The photoconvertible fluorophore synthesis enables the light controlled imaging channels switch for accurate tracking the quantity and localization of intracellular biomolecules in chemical biology. Herein, we repurposed the photochemistry of Fischer's base and developed a sunlight-directed fluorophore-switch strategy for high-efficiency trimethine cyanine (Cy3.5/Cy3) synthesis. The unexpected sunlight-directed photoconversion of Fischer's base proceeds in conventional solvents and accelerates in chloroform via photo-oxidation and hydrogen atom transfer without using extra additives, and the heterogenous dimerization mechanism was proposed and confirmed by isolation of the reactive intermediates. The reliable strategy is employed in the photosynthesis of commercially available cytomembrane marker (DiI) and other cyanine based organelle markers with appreciable yields. Sunlight-controlled fluorophore-switch of subcellular organelle markers in living cells validated the feasibility of our strategy with cell-tolerant character. Moreover, remote control synthesis of Cy3.5 in vivo directed via sunlight further demonstrated the extended application of our strategy. Therefore, this sunlight-directed strategy will facilitate exploitation of cyanine-based probes with switched fluorescence imaging channels and further enable precise description of the dynamic variations in living cells with minimal autofluorescence and cellular disturbance.

Temperature is the most commonly collected vital sign in all of clinical medicine; it plays a critical role in care decisions related to topics ranging from infection to inflammation, sleep, and fertility. Most assessments of body temperature occur at isolated anatomical locations (e.g. axilla, rectum, temporal artery, or oral cavity). Even this relatively primitive mode for monitoring can be challenging with vulnerable patient populations due to physical encumbrances and artifacts associated with the sizes, weights, shapes and mechanical properties of the sensors and, for continuous monitoring, their hard-wired interfaces to data collection units. Here, we introduce a simple, miniaturized, lightweight sensor as a wireless alternative, designed to address demanding applications such as those related to the care of neonates in high ambient humidity environments with radiant heating found in incubators in intensive care units. Such devices can be deployed onto specific anatomical locations of premature infants for homeostatic assessments. The estimated core body temperature aligns, to within 0.05 °C, with clinical grade, wired sensors, consistent with regulatory medical device requirements. Time-synchronized, multi-device operation across multiple body locations supports continuous, full-body measurements of spatio-temporal variations in temperature and additional modes of determining tissue health status in the context of sepsis detection and various environmental exposures. In addition to thermal sensing, these same devices support measurements of a range of other essential vital signs derived from thermo-mechanical coupling to the skin, for applications ranging from neonatal and infant care to sleep medicine and even pulmonary medicine.

Viruses have unique coat proteins that are genetically modifiable. Their surface can serve as a nano-template on which electroactive molecules are immobilized. In this study, we report filamentous bacteriophage as a backbone to which redox mediators are covalently and densely tethered, constructing redox nanowire, i.e. an electron conducting biomaterial. The highly ordered coat proteins of a filamentous bacteriophage provide flexible and biocompatible platform to constitute a biohybrid redox nanowire. Incorporating bacteriophage and redox molecules form an entangled assembly of nanowires enabling facile electron transfer. Electron transfer among the molecular mediators in the entangled assembly originates apparent electron diffusion of which the electron transfer rate is comparable to that observed in conventional redox polymers. Programming peptide terminals suggests further enhancement in electron mediation by increasing redox species mobility. In addition, the redox nanowire film functions as a favorable matrix for enzyme encapsulation. The stability of the enzymes entrapped in this unique matrix is substantially improved.

Small molecule-binding aptamers often suffer from high cross reactivity to structure analogues in biological samples, limiting their value for clinical diagnosis. Herein, we present a method to overcome this issue, by performing binding-inhibited organic reaction-based regioselective selection of aptamers against homocysteine (Hcy), which is a marker for diagnosing many disorders including stroke and Alzheimer's. This approach has led to isolation of a DNA aptamer that binds to the alkane thiol chain of Hcy with exceptional specificity against cysteine. It also binds with oxidized Hcy at weaker affinity. Using this new aptamer, we produced a reusable fluorescent optical fiber aptasensor for direct and validated detection of both free and total Hcy in nondeproteinized patient plasma in the diagnostic concentration range. The binding site-specific aptamer selection and optical-fiber-sensing strategy can expand the practical utility of aptamers in clinical diagnosis.

Biosensors developed through a sandwich approach have demonstrated favorable detection performance for exosomal programmed cell death 1 ligand 1 (ExoPD-L1) detection. However, the reported PD-L1 antibodies, peptides, and aptamers utilized in these biosensors typically bind to the extracellular region, with overlapping binding sites that severely constrain the fabrication of biosensors. In this study, we present a simple approach to specifically identify and analyze ExoPD-L1 through the non-selective trapping effect of Ti3C2TX (X=-O, -F, -OH) MXene on exosomes via the formation of Ti-O-P complexation, and the selective capture of peptide-functionalized Au@MPBA (4-Mercaptophenylboronic acid) @SiO2 surface enhanced Raman scattering (SERS) tags on ExoPD-L1. The biosensor delivered a both hypersensitive and reliable performance in exosome detection with a low limit of detection (20.74 particles/mL) in the linear range of 102 to 5×106 particles/mL. Furthermore, the biosensor demonstrated excellent stability and interference resistance in detecting ExoPD-L1 in clinical serum samples, enabling the easy differentiation of breast cancer patients from healthy controls. This work provides new insights into the design of biosensors for exosome detection and can serve as a replicable template for sandwich immunoassay detection for other types of sensors, including but not limited to SERS.

Early diagnosis of chronic kidney disease (CKD) and constant monitoring to guide optimal intervention is critical to prevent renal failure and other critical diseases. However, the conventional blood tests in hospital are time-consuming and have poor patient compliance. Herein, we demonstrate a real-time, minimally invasive, and self-administrable approach to detect kidney biomarkers in the skin interstitial fluid (ISF) using a polymeric microneedle coupled electrochemical sensor array (MNESA). Microneedles can readily penetrate stratum corneum and quickly extract ISF onto the sensors. Four biomarkers are simultaneously detected to avoid false positive and provide an accurate assessment of kidney functions. Using an artificial skin model, it is shown that MNSEA gives specific and sensitive responses to these kidney biomarkers in physiologically relevant ranges (phosphate: 0.3-1.8 mM, 3.62 μA/mM; uric acid: 50-550 μM, 4.19 nA/μM; creatinine: 50-550 μM, 12.58 nA/μM; urea: 1-16 mM, 44.6 mV/decade). Using a mouse model, we demonstrate that this approach is as reliable as the commercial assays and is feasible to readily monitor the progression of CDK.

Umami, a taste sensation known for its savory and delicious properties, has garnered considerable attention from both consumers and the food industry. However, current understanding and evaluation of umami characteristics remain limited, presenting a long-standing issue. To address this challenge, we have developed a self-assembled biosensor based on matured taste receptor cells (TRCs), obtained through isolation and culture of taste stem cells. TRCs, as the recognition element, were mounted onto the surface of a glassy carbon electrode (GCE) treated with gold nanoparticles (AuNPs) and poly-L-lysine (PLL). Key parameters including the cell incubation time and concentration were optimized to ensure the optimal performance of the TRCs-based biosensor. AuNPs were deposited onto the GCE surface via 90 s electrochemical reduction. TRCs concentration of 106 cells/mL and incubation time of 12 h were chosen by electrochemical characterization. Using this novel, rapid, and sensitive TRCs-based biosensor, we successfully detected L-monosodium glutamate (MSG) and other umami substances, demonstrating a good linear relationship within the range of 10-9 - 10-5 M between response signals and concentration of MSG stimuli. Our results provide insights into taste signal transduction mechanisms and suggest the potential for biomimetic sensors in intelligent perception applications.

Conventional pathogen detection strategies based on the molecular structure or chemical characteristics of biomarkers can only provide the "physical abundance" of microorganisms, but cannot reflect the "biological effect abundance" in the true sense. To address this issue, we report an erythrocyte membrane-encapsulated biomimetic sensor cascaded with CRISPR-Cas12a (EMSCC). Taking hemolytic pathogens as the target model, we first constructed an erythrocyte membrane-encapsulated biomimetic sensor (EMS). Only hemolytic pathogens with biological effects can disrupt the erythrocyte membrane (EM), resulting in signal generation. Then the signal was amplified by cascading CRISPR-Cas12a, and more than 6.67 × 104-fold improvement in detection sensitivity compared to traditional erythrocyte hemolysis assay was achieved. Notably, compared with polymerase chain reaction (PCR) or enzyme linked immunosorbent assay (ELISA)-based quantification methods, EMSCC can sensitively respond to the pathogenicity change of pathogens. For the detection of simulated clinical samples based on EMSCC, we obtained an accuracy of 95% in 40 samples, demonstrating its potential clinical value.

The coronavirus disease (COVID-19) pandemic has highlighted the importance of rapid and accurate diagnosis, and loop-mediated isothermal amplification (LAMP) has become a popular method because of its powerful amplification ability using a simple instrument such as a heater or water bath. However, LAMP has limitations such as the complexity of primer design and the difficulty of designing sequence-specific probes, leading to non-specific amplicons and false-positive results. To overcome these limitations, we developed a novel isothermal amplification system called the Extension-mediated self-folding Isothermal amplification Technology (ExIT). ExIT uses a newly designed, self-folding primer (SP) with two key functions. Hairpin structures are formed when the extended strand of the SP hybridizes, exposing the priming site for continuous binding of the new SP. This results in exponential amplification with only two primers, unlike conventional LAMP primer systems. Additionally, an unnatural base was introduced into the SP, which terminated the extension of polymerase and generated a ssDNA amplicon. This makes it easier to design and apply probes, reducing the possibility of false-positive results even if non-specific amplicons are produced. Through this strategy, we confirmed a sensitivity of 90 copies (3.6 copies/μL) and verified the specificity by testing for the presence or absence of non-complementary targets. Therefore, the validation of the ExIT was completed. In conclusion, ExIT will be key to solving the complexity of conventional LAMP design and offers great potential for successfully introducing sequence-specific probes to improve false positives.

Color-switch electrochemiluminescence (ECL) sensing platform based on a dual-bipolar electrode (D-BPE) is reported in this work. The D-BPE was composed of a cathode filled with buffer and two anodes filled with [Ru(bpy)3]2+-TPrA and luminol-H2O2 solutions, respectively. Both anodes were modified with capture DNA and served as ECL reporting platforms. After introducing ferrocene-labeled aptamer (Fc-aptamer) on both anodes, the ECL emission signal of the [Ru(bpy)3]2+ was difficult to be observed (anode 1), while luminol emitted a strong and visible ECL signal (anode 2). Ferrocene (Fc) did not only prevent the oxidation of [Ru(bpy)3]2+ due to its lower oxidation potential, its oxidation product Fc+ also quenched the [Ru(bpy)3]2+ ECL through efficient energy transfer. For luminol, Fc+ catalyzes the accelerated formation of the excited-state of the luminol anion radical, which leads to the enhancement of the luminol ECL. In the presence of food-borne pathogens, the aptamer was assembled with them, leading to the leaving of Fc from the surface of the D-BPE anodes. The ECL intensity of [Ru(bpy)3]2+ was enlarged, meanwhile, the blue emission signal of luminol became weakened. By self-calibrating the ratio of the two signals, 1-106 CFU mL-1 food-borne pathogenic bacteria can be sensitively detected with a detection limit of 1 CFU mL-1. Ingeniously, the color-switch biosensor can be used to detect S. aureus, E. coli and S. typhimurium by assembling the corresponding aptamers onto the D-BPE anodes.

The global pandemic resulting from the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its emerging variants highlights the need for convenient and accurate detection protocols to facilitate timely prevention and management of the disease. Herein, we propose a new self-priming hairpin-mediated isothermal amplification (SIAM) protocol enabling one-pot and ultrasensitive identification of SARS-CoV-2 in a multiplexed way. This approach works by targeting a specific RNA sequence with a self-priming hairpin (SP) probe and promoting continuously repeated extension and nicking reactions to produce numerous trigger molecules, which could specifically bind to molecular beacons (MBs) and produce fluorescent signals. Under an isothermal condition of 37 °C, this technique allowed for the simultaneous identification of the spike (S) and nucleocapsid (N) genes of SARS-CoV-2 down to single copy/μL levels. We further validated the practical diagnostic capabilities of the SIAM method by accurately testing 20 clinical samples with 100% sensitivity and specificity. The SIAM method has a lot of potential to be a reliable nucleic acid testing protocol to identify infections caused by a wide range of pathogens.

Self-assembled functional nanomaterials with electromagnetic hot spots are crucial and highly desirable in surface-enhanced Raman scattering (SERS). Due to its versatile biological scaffold, the M13 phage has been employed to produce novel nano-building blocks and devices. In this study, we propose a novel M13 phage-based SERS nanocarrier, that utilizes the pVIII capsid in M13 to conjugate Au@Ag core-shell nanorod (Au@AgNR) with linker carboxy-PEG-thiol (M13-Au@AgNR) and the pIII capsid to specifically target Escherichia coli (E. coli). The M13-Au@AgNR@DTTC (3,3'- diethylthiocarbocyanine iodide) SERS probe was used to detect E. coli in a concentration range of 6 to 6 × 105 cfu/mL, achieving a limit of detection (LOD) of 0.5 cfu/mL. The proposed SERS platform was also tested in real samples, showing good recoveries (92%-114.3%) and a relative standard deviation (RSD) of 1.2%-4.7%. Furthermore, the system demonstrated high antibacterial efficiency against E. coli, approximately 90%, as measured by the standard plate-count method. The investigation provides an effective strategy for in vitro bacteria detection and inactivation.

Light sources are crucial for photoelectrochemical (PEC) self-powered sensing, where visible light is widely used. However, due to its high energy, it has some downsides as an irradiation source for overall system, so it is urgent to achieve effective near-infrared (NIR) light absorption because it makes up a significant portion of the solar spectrum. Herein, up-conversion nanoparticles (UCNPs) that could increase the energy of low-energy radiation were combined with semiconductor CdS as the photoactive material (UCNPs/CdS), which broadens the response range of solar spectrum. The NIR light-excited self-powered sensor could be produced via oxidizing H2O at photoanode and lowering dissolved oxygen at cathode under the NIR light without external voltage. Meanwhile, molecularly imprinted polymer (MIP) was added to photoanode as a recognition element to increase the sensor's selectivity. The open-circuit voltage of the self-powered sensor grew linearly as chlorpyrifos concentration climbed from 0.01 to 100 ng mL-1, showing good selectivity as well as reproducibility. This work provides valuable basis for the preparation of efficient and practical PEC sensor with NIR light response.

The controllable modulation of the response mode is highly attractive for the construction of photoelectrochemical (PEC) sensors with improved sensitivity and anti-interference ability in complex real samples matrix. Here, we present a charming proof-of-concept ratiometric PEC aptasensor of enrofloxacin (ENR) analysis via the controllable signal transduction. Different with the traditional sensing mechanism, this ratiometric PEC aptasensor integrates the anodic PEC signal induced by PtCuCo nanozyme-catalyzed precipitation reaction and the polarity-switching cathodic PEC response mediated by Cu2O nanocubes on S-scheme FeCdS@FeIn2S4 heterostructure. Taking advantages of the photocurrent-polarity-switching signal response model and the superior performance of the photoactive substrate material, the proposed ratiometric PEC aptasensor displays a good detection linear range for ENR analysis from 0.01 pg mL-1 to 10 ng mL-1, with a detection limit of 3.3 fg mL-1. This study provides a general platform for detecting interested trace analytes in real samples and expands the diversity of sensing strategy design.

Microelectrode Arrays (MEAs) are popular tools for in vitro extracellular recording. They are often optimized by surface engineering to improve affinity with neurons and guarantee higher recording quality and stability. Recently, PEDOT:PSS has been used to coat microelectrodes due to its good biocompatibility and low impedance, which enhances neural coupling. Herein, we investigate on electro-co-polymerization of EDOT with its triglymated derivative to control valence between monomer units and hydrophilic functions on a conducting polymer. Molecular packing, cation complexation, dopant stoichiometry are governed by the glycolation degree of the electro-active coating of the microelectrodes. Optimal monomer ratio allows fine-tuning the material hydrophilicity and biocompatibility without compromising the electrochemical impedance of microelectrodes nor their stability while interfaced with a neural cell culture. After incubation, sensing readout on the modified electrodes shows higher performances with respect to unmodified electropolymerized PEDOT, with higher signal-to-noise ratio (SNR) and higher spike counts on the same neural culture. Reported SNR values are superior to that of state-of-the-art PEDOT microelectrodes and close to that of state-of-the-art 3D microelectrodes, with a reduced fabrication complexity. Thanks to this versatile technique and its impact on the surface chemistry of the microelectrode, we show that electro-co-polymerization trades with many-compound properties to easily gather them into single macromolecular structures. Applied on sensor arrays, it holds great potential for the customization of neurosensors to adapt to environmental boundaries and to optimize extracted sensing features.

Transistor-based biosensors represent an emerging technology for inexpensive point-of-care testing (POCT) applications. However, the limited sensitivity of the current transistor technologies hinders their practical deployment. In this study, we developed tri-channel In2O3/ZnO heterojunction thin-film transistors (TFTs) featuring the surface-immobilized enzyme glucose oxidase to detect glucose in various biofluids. This unusual channel design facilitates strong coupling between the electrons transported along the buried In2O3/ZnO heterointerface and the electrostatic perturbations caused by the interactions between glucose and surface-immobilized glucose oxidase. The enzyme selectively binds to glucose, causing a change in charge density on the channel surface. By exploring this effect, the solid-state biosensing TFT (BioTFT) can selectively detect glucose in artificial and real saliva over a wide range of concentrations from 500 nM to 20 mM with limits of detection of ∼365 pM (artificial saliva) and ∼416 nM (real saliva) in less than 60 s. The specificity of the sensor towards glucose has been demonstrated against various interfering species in artificial saliva, further highlighting its unique capabilities. Moreover, the BioTFTs exhibited good operating stability upon storage for up to two weeks, with relative standard deviation (RSD) values ranging from 2.36% to 6.39% for 500 nM glucose concentration. Our BioTFTs are easy to manufacture with reliable operation, making them ideal for non-invasive POCT applications.

Experiential richness creates tissue-level changes and synaptic plasticity as patterns emerge from rhythmic spatiotemporal activity of large interconnected neuronal assemblies. Despite numerous experimental and computational approaches at different scales, the precise impact of experience on network-wide computational dynamics remains inaccessible due to the lack of applicable large-scale recording methodology. We here demonstrate a large-scale multi-site biohybrid brain circuity on-CMOS-based biosensor with an unprecedented spatiotemporal resolution of 4096 microelectrodes, which allows simultaneous electrophysiological assessment across the entire hippocampal-cortical subnetworks from mice living in an enriched environment (ENR) and standard-housed (SD) conditions. Our platform, empowered with various computational analyses, reveals environmental enrichment's impacts on local and global spatiotemporal neural dynamics, firing synchrony, topological network complexity, and large-scale connectome. Our results delineate the distinct role of prior experience in enhancing multiplexed dimensional coding formed by neuronal ensembles and error tolerance and resilience to random failures compared to standard conditions. The scope and depth of these effects highlight the critical role of high-density, large-scale biosensors to provide a new understanding of the computational dynamics and information processing in multimodal physiological and experience-dependent plasticity conditions and their role in higher brain functions. Knowledge of these large-scale dynamics can inspire the development of biologically plausible computational models and computational artificial intelligence networks and expand the reach of neuromorphic brain-inspired computing into new applications.

Constructing mono-luminophor-based electrochemiluminescence (ECL) ratio system is a great challenge due to the limitations of the luminescent species with dual-signal-output, luminescence efficiency and coreactant. This work developed carboxyl-functionalized poly[9,9-bis(3'-(N,N-dimethylamino) propyl)-2,7-fluorene]-alt-2,7-(9,9 dioctylfluorene)] nanoparticles(PFN NPs) as dual-emitting luminophors, which can synchronously output strong cathodic and anodic ECL signals without any exogenous coreactants. The inherent molecular structure enabled efficient intramolecular electron transfer between tertiary amine groups and backbone of PFN to generate strong cathodic and anodic ECL emission. Particularly, H+ in aqueous solution played an irreplaceable role for cathodic ECL emission. The silver nanoparticles (AgNPs) were developed as signal regulator because of their excellent hydrogen evolution reaction (HER) activity, which significantly quenched the cathodic signal while kept the anodic signal unchanged. The dual-emitting PFN NPs cleverly integrated signal regulator AgNPs and bicyclic strand displacement amplification (SDA) to construct a coreactant-free mono-luminophor-based ratiometric ECL sensing for SARS-CoV-2 RdRp gene assay. The strong dual-emitting of PFN NPs and excellent quenching effect of AgNPs on cathodic emission endowed the biosensor with a high detection sensitivity, and the detection limit was as low as 39 aM for RdRp gene. The unique dual-emitting properties of PFN NPs open up a new path to construct coreactant-free mono-luminophor-based ECL ratio platform, and excellent HER activity of AgNPs offers some new thoughts for realizing ECL signal changes.

High performance sorting of circulating tumor cells (CTCs) from peripheral blood is key to liquid biopsies. Size-based deterministic lateral displacement (DLD) technique is widely used in cell sorting. But conventional microcolumns have poor fluid regulation ability, which limits the sorting performance of DLD. When the size difference between CTCs and leukocytes is small (e.g., less than 3 μm), not only DLD, many size-based separation techniques fail due to low specificity. CTCs have been confirmed to be softer than leukocytes, which could serve as a basis for sorting. In this study, we presented a multistage microfluidic CTCs sorting method, first sorting CTCs using a size-based two-array DLD chip, then purifying CTCs mixed by leukocytes using a stiffness-based cone channel chip, and finally identifying cell types using Raman techniques. The entire CTCs sorting and analysis process was label free, highly pure, high-throughput and efficient. The two-array DLD chip employed a droplet-shaped microcolumn (DMC) developed by optimization design rather than empirical design. Attributed to the excellent fluid regulation capability of DMC, the CTCs sorter system developed by parallelizing four DMC two-array DLD chips was able to process a sample of 2.5 mL per minute with a recovery efficiency of 96.30 ± 2.10% and a purity of 98.25 ± 2.48%. To isolate CTCs mixed dimensionally by leukocytes, a cone channel sorting method and chip were developed based on solid and hydrodynamic coupled analysis. The cone channel chip allowed CTCs to pass through the channel and entrap leukocytes, improving the purity of CTCs mixed by leukocytes by 1.8-fold.

Surface-enhanced Raman spectroscopy (SERS) sandwich biosensors have received tremendous attention in early diagnosis of bacterial infections. However, efficiently engineering nanoscale plasmonic hots pots (HS) towards ultrasensitive SERS detection still remains challenging. Herein, we propose a bioinspired synergistic HS engineering strategy to construct ultrasensitive SERS sandwich bacterial sensor (named USSB), by coupling bioinspired signal module and plasmonic enrichment module to synergistically boost the number and intensity of HS. The bioinspired signal module is based on dendritic mesoporous silica nanocarrier (DMSN) loaded with plasmonic nanoparticles and SERS tag, while magnetic Fe3O4 nanoparticles coated with Au shell are employed in plasmonic enrichment module. We demonstrate that DMSN effectively shrank nanogaps between plasmonic nanoparticles to improve HS intensity. Meanwhile, plasmonic enrichment module contributed to plenty of additional HS inside and outside individual "sandwich". Ascribing to the boosted number and intensity of HS, the constructed USSB sensor exhibits ultrahigh detection sensitivity (7 CFU/mL) and selectivity towards model pathogenic bacteria of Staphylococcus aureus. Remarkably, the USSB sensor enables fast and accurate bacterial detection in real blood samples of septic mice, achieving early diagnosis of bacterial sepsis. The proposed bioinspired synergistic HS engineering strategy opens up a new direction for constructing ultrasensitive SERS sandwich biosensors, and may promote their advancing applications in the early diagnosis and prognosis of devastating diseases.

Microcystis aeruginosa (M. aeruginosa) cause the eutrophication of lakes and rivers. To effectively control the overgrowth of M. aeruginosa, a suitable measurement method should be required in the aquatic fields. To address this, we developed a field-ready cyanobacterial pretreatment device and an electrochemical clustered regularly interspaced short palindromic repeats (EC-CRISPR) biosensor. The cyanobacterial pretreatment device consists of a syringe, glass bead, and graphene oxide (GO) bead. Then, the M. aeruginosa dissolved in the freshwater sample was added to fabricated filter. After filtration, the purified gene was loaded onto a CRISPR-based electrochemical biosensor chip to detect M. aeruginosa gene fragments. The biosensor was composed of CRISPR/Cpf1 protein conjugated with MXene on an Au microgap electrode (AuMGE) integrated into a printed circuit board (PCB). This AuMGE/PCB system maximizes the signal-to-noise ratio, which controls the working and counter electrode areas requiring only 3 μL samples to obtain high reliability. Using the extracted M. aeruginosa gene with a pre-treatment filter, the CRISPR biosensor showed a limit of detection of 0.089 pg/μl in fresh water. Moreover, selectivity test and matrix condition test carried out using the EC-CRISPR biosensor. These handheld pre-treatment kit and biosensors can enable field-ready detection of CyanoHABs.

Developing substrates that combine sensitivity and signal stability is a major challenge in surface enhanced Raman scattering (SERS) research. Herein, we present a flexible triple-enhanced Raman Scattering MXene/Au nanocubes (AuNCs) sensor fabricated by selective filtration of Ti3C2Tx MXene/AuNCs hybrid on the Ti3C2Tx MXene membrane and subsequent treatment with 1H,1H,2H,2H-perfluoro-octyltriethoxysilane (FOTS). The resultant superhydrophobic MXene/AuNCs-FOTS membrane not only provides the SERS substrate with environmental stability, but also imparts analyte enrichment to enhance the sensitivity (LOD = 1 × 10-14 M) and reliability (RSD = 6.41%) for Rhodamine 6G (R6G) molecules owing to the attenuation of the coffee ring effect. Moreover, the triple enhancement mechanism of combining plasmonic coupling enhancement from plasmonic coupling (EM) of nearby AuNCs at lateral and longitudinal direction of MXene/AuNCs-FOTS membrane, charge transfer (CT) from Ti3C2Tx MXene and target molecules and analyte enrichment function provides the substrate with excellent SERS performance (EF = 3.19 × 109), and allows efficient quantification of biomarkers in urine. This work could provide new insights into MXenes as building blocks for high-performance substrates and fill existing gaps in SERS techniques.

Bisphenol A (BPA) is an endocrine disrupting chemical and broadly used in plastics. The leakage of BPA in food and water cycles poses a significant risk to the environment and human health. Thus, monitoring the concentration of BPA to avoid its potential risk is highly important. In this work, a simple and efficient oxygen deficient molecularly imprinted TiO2 electrochemical sensor was proposed for the detection of BPA. The introduction both oxygen vacancies and molecular imprinting evidently enhanced the electrochemical oxidation signal of BPA. The sensor had a good linear response ranging from 0.01 μM to 20 μM with a limit of detection of 3.6 nM. Additionally, the sensor showed remarkable stability, reproducibility and interference resistant ability. It also exhibits excellent recovery during the detection of real water. These findings suggested that the sensor has the potential to be developed as a simple, efficient and low-cost monitoring system for the monitoring of BPA in water.

MicroRNA-21 (miRNA-21) is currently the only known oncogenic miRNA that is upregulated in almost all malignant tumors and exhibits a broad spectrum of tumor recognition characteristics. It holds significant value in the early diagnosis, malignant degree assessment, and prognostic evaluation of tumors. In this study, a novel dual-mode self-powered sensing platform is developed using Au nanoparticles/graphdiyne as the electrode substrate and combined with DNA nanoring for highly sensitive and specific detection of miRNA-21. The DNA nanoring structure, which is easy to prepare and contains multiple recognition sites, induces significant electrochemical/colorimetric signal responses of the signaling molecule methylene blue. Under optimal conditions, the linear ranges of the electrochemical and colorimetric detection modes of this self-powered sensor are 0.1 fM-100 pM and 0.1 fM-10 nM, respectively, with the detection limits of 35.1 aM and 61.6 aM (S/N=3). This strategy provides a new reference for the sensitive detection of microRNA and has immense potential for application in the screening and detection of clinical nucleic acid diseases.

The conductivity and emission efficiency of metal-organic frameworks (MOFs) remain challenging factors that limit their electrogenerated chemiluminescence (ECL) sensing applications. Herein, we report a facile approach to address these challenges by integrating an electroactive linker (H2-TCPP) with an ECL active electrogenerated chemiluminescence linker (H4-TBAPy) to construct a highly photoelectrochemical active mixed-linker MOFs (ML-MOFs). ECL results revealed a remarkable 15.4-fold enhancement for the top-performing ML-MOFs (M6-MOFs), surpassing the single linker MOFs. In addition, M6-MOFs also exhibit a remarkable 73-fold enhancement in ECL efficiency compared to commercial Ru (bpy)32+. This improvement should be attributed to the synergistic effects resulting from the combination of two linkers. Furthermore, M6-MOFs are found to be served as a model ECLphore for sensitive and selective detection of α-glucosidase for the first time with good potential practicability in human serum samples. This work represents a promising direction to guide for designing good conductivity and high ECL efficiency MOFs in terms of linker functionalization and thus bandgap modulation for advancing their ECL sensing applications.

A double-cycle system has been developed for specifically detecting trace amounts of Pb2+ by significantly decreasing the background signal. The detection involves two types of RNA cleavage reactions: one using a Pb2+-specific GR5 DNAzyme (PbDz) and the other utilizing a newly constructed 10-23 DNAzyme with two hairpins embedded in its catalytic center (hpDz). The ring-structured hpDz (c-hpDz) exhibits significantly lower activity compared to the circular 10-23 DNAzyme without hairpin structures, which plays a crucial role in reducing the background signal. When Pb2+ is present, PbDz cleaves c-hpDz to its active form, which then disconnects the molecular beacon to emit the fluorescent signal. The method allows for rapid and sensitive Pb2+ detection within 40 min for 10 fM of Pb2+ and even as short as 10 min for 100 nM of Pb2+. Additionally, visual detection is possible through the non-crosslinking assembly of Au nanoparticles. The entire process can be performed in one pot and even one step, making it highly versatile and suitable for a wide range of applications, including food safety testing and environmental monitoring.

In recent years, the demand for rapid, sensitive, and simple methods for diagnosing deoxyribonucleic acid (DNA) has grown due to the increase in the variation of infectious diseases. This work aimed to develop a flash signal amplification method coupled with electrochemical detection for polymerase chain reaction (PCR)-free tuberculosis (TB) molecular diagnosis. We exploited the slightly miscible properties of butanol and water to instantly concentrate a capture probe DNA, a single-stranded mismatch DNA, and gold nanoparticles (AuNPs) to a small volume to reduce the diffusion and reaction time in the solution. In addition, the electrochemical signal was enhanced once two strands of DNA were hybridized and bound to the surface of the gold nanoparticle at an ultra-high density. To eliminate non-specific adsorption and identify mismatched DNA, the self-assembled monolayers (SAMs) and Muts proteins were sequentially modified on the working electrode. This sensitive and specific approach can detect as low as attomolar levels of DNA targets (18 aM) and is successfully applied to detecting tuberculosis-associated single nucleotide polymorphisms (SNPs) in synovial fluid. More importantly, as this biosensing strategy can amplify the signal in only a few seconds, it possesses a great potential for point-of-care and molecular diagnosis applications.

Surface plasmon polaritons in graphene can enhance the performance of mid-infrared spectroscopy, which is key for the study of both the composition and the conformation of organic molecules via their vibrational resonances. In this paper, a plasmonic biosensor using a graphene-based van der Waals heterostructure on a piezoelectric substrate is theoretically demonstrated, where far-field light is coupled to surface plasmon-phonon polaritons (SPPPs) through a surface acoustic wave (SAW). The SAW creates an electrically-controlled virtual diffraction grating, suppressing the need for patterning the 2D materials, that limits the polariton lifetime, and enabling differential measurement schemes, which increase the signal-to-noise ratio and allow a quick commutation between reference and sample signals. A transfer matrix method has been used for simulating the SPPPs propagating in the system, which are electrically tuned to interact with the vibrational resonances of the analytes. Furthermore, the analysis of the sensor response with a coupled oscillators model has proven its capability of fingerprinting ultrathin biolayers, even when the interaction is too weak to induce a Fano interference pattern, with a sensitivity down to the monolayer limit, as tested with a protein bilayer or a peptide monolayer. The proposed device paves the way for the development of advanced SAW-assisted lab-on-chip systems combining the existing SAW-mediated physical sensing and microfluidic functionalities with the chemical fingerprinting capability of this novel SAW-driven plasmonic approach.

Tauopathies are neurodegenerative diseases characterized by abnormal conformational changes in tau protein. Early hyperphosphorylation-induced conformational changes are considered a hallmark of tauopathy, but real-time tracking methods are lacking. Here, we present two novel fluorescence resonance energy transfer (FRET)-based tau biosensors that detect such changes with high spatiotemporal resolution at the single-cell level. The TAUCON biosensor measures instantaneous conformational changes in hyperphosphorylated tau within 20 min, while the TAUCOM biosensor detects changes in the paper-clip structure of microtubule-associated tau. Our biosensors provide faster and more precise detection than conventional methods and can serve as valuable tools for investigating the initial causes, mechanisms, progression, and treatment of tauopathies.

Sensitive and anti-interference detection of targeted signal(s) in body fluids is one of the paramount tasks in biosensing. Overcoming the complication and high cost of antibody/aptamer-modification, surface-enhanced Raman spectroscopy (SERS) based on antibody/aptamer-free (AAF) substrates has shown great promise, yet with rather limited detection sensitivity. Herein, we report ultrasensitive and anti-interference detection of SARS-CoV-2 spike protein in untreated saliva by an AAF SERS substrate, applying the evanescent field induced by the high-order waveguide modes of well-defined nanorods for SERS for the first time. A detection limit of 3.6 × 10-17 M and 1.6 × 10-16 M are obtained in phosphate buffered saline and untreated saliva, respectively; the detection limits are three orders of magnitude improved than the best records from AAF substrates. This work unlocks an exciting path to design AAF SERS substrates for ultrasensitive biosensing, not limited to detection of viral antigens.

Sensitive and accurate cell-free plasma Epstein-Barr virus (EBV) DNA measurement is essential in the routine diagnosis, monitoring and treatment of Nasopharyngeal Carcinoma (NPC). This measurement in commercial and in-house assay are commonly based on real-time quantitative PCR (qPCR) method, which requires reference materials for standardization and lack quantitative precision due to amplification bias or cross-contamination. To address these issues, we developed a CRISPR/Cas12a-mediated amplification-free digital DNA assay, which targets the repetitive sequences of EBV DNA and utilizes the cis-cleavage activity of CRISPR-Cas12a prior to droplet generation. By this mean, more activated Cas12a-crRNA duplexes could be produced for subsequent target detection and counting, thus improving the performance in detecting low EBV DNA load. We demonstrated that it was more robust than conventional qPCR for detecting plasma EBV DNA in a case-control study of 208 participants, especially when the target concentrations were around the diagnostic cut-off value for NPC. More importantly, this assay allowed a more accurate diagnosis of early-stage NPC, with an area under the curve (AUC) of 0.9883 (versus 0.7682 for qPCR). Furthermore, its absolute quantification capability enabled dynamic monitoring of EBV load in NPC patients during initial diagnosis, treatment, and recurrence, thereby potentially improving disease management and prognosis. Taken together, our results demonstrate that this amplification-free digital assay has the potential to be a robust tool to improve the diagnosis and surveillance of NPC.

Wearable point-of-care testing devices are essential for personalized and decentralized healthcare. They can collect biofluid samples from the human body and use an analyzer to detect biomolecules. However, creating an integrated system is challenging due to the difficulty of achieving conformality to the human body, regulating the collection and transport of biofluids, developing a biosensor patch capable of precise biomolecule detection, and establishing a simple operation protocol that requires minimal wearer attention. In this study, we propose using a hollow microneedle (HMN) based on soft hollow microfibers and a microneedle-integrated microfluidic biosensor patch (MIMBP) capable of integrated blood sampling and electrochemical biosensing of biomolecules. The soft MIMBP includes a stretchable microfluidic device, a flexible electrochemical biosensor, and a HMN array made from flexible hollow microfibers. The HMNs are fabricated by electroplating flexible and mechanically durable hollow microfibers made from a nanocomposite matrix of polyimide, a poly (vinylidene fluoride-co-trifluoroethylene) copolymer, and single-walled carbon nanotubes. The MIMBP uses the negative pressure generated by a single button push to collect blood and deliver it to a flexible electrochemical biosensor modified with a gold nanostructure and Pt nanoparticles. We have demonstrated that glucose can be accurately measured up to the molar range in whole human blood collected through the microneedle. The MIMBP platform with HMNs has great potential as a foundation for the future development of simple, wearable, self-testing systems for minimally invasive biomolecule detection. This platform capable of sequential blood collection and high sensitivity glucose detection, which are ideal for personalized and decentralized healthcare.

The detection of severe acute respiratory syndrome coronavirus (SARS-CoV-1) was demonstrated using screened Fv-antibodies for SPR biosensor and impedance spectrometry. The Fv-antibody library was first prepared on the outer membrane of E. coli using autodisplay technology and the Fv-variants (clones) with a specific affinity toward the SARS-CoV-1 spike protein (SP) were screened using magnetic beads immobilized with the SP. Upon screening the Fv-antibody library, two target Fv-variants (clones) with a specific binding affinity toward the SARS-CoV-1 SP were determined and the Fv-antibodies on two clones were named "Anti-SP1" (with CDR3 amino acid sequence: 1GRTTG5NDRPD11Y) and "Anti-SP2" (with CDR3 amino acid sequence: 1CLRQA5GTADD11V). The binding affinities of the two screened Fv-variants (clones) were analyzed using flow cytometry and the binding constants (KD) were estimated to be 80.5 ± 3.6 nM for Anti-SP1 and 45.6 ± 8.9 nM for Anti-SP2 (n = 3). In addition, the Fv-antibody including three CDR regions (CDR1, CDR2, and CDR3) and frame regions (FRs) between the CDR regions was expressed as a fusion protein (Mw. 40.6 kDa) with a green fluorescent protein (GFP) and the KD values of the expressed Fv-antibodies toward the SP estimated to be 15.3 ± 1.5 nM for Anti-SP1 (n = 3) and 16.3 ± 1.7 nM for Anti-SP2 (n = 3). Finally, the expressed Fv-antibodies screened against SARS-CoV-1 SP (Anti-SP1 and Anti-SP2) were applied for the detection of SARS-CoV-1. Consequently, the detection of SARS-CoV-1 was demonstrated to be feasible using the SPR biosensor and impedance spectrometry utilizing the immobilized Fv-antibodies against the SARS-CoV-1 SP.

Conventional rapid detection methods are difficult to identify or distinguish various pesticide residues at the same time. And sensor arrays are also limited by the complexity of preparing multiple receptors and high cost. To address this challenge, a single material with multiple properties is considered. Herein, we first found that different categories of pesticides have diverse regulatory behaviors on the multiple catalytic activities of Asp-Cu nanozyme. Thus, a three-channel sensor array based on the laccase-like, peroxidase-like, and superoxide dismutase-like activities of Asp-Cu nanozyme was constructed and successfully used for the discrimination of eight kinds of pesticides (glyphosate, phosmet, isocarbophos, carbaryl, pentachloronitrobenzene, metsulfuron-methyl, etoxazole, and 2-methyl-4-chlorophenoxyacetic acid). In addition, a concentration-independent model for qualitative identification of pesticides has been established, and 100% correctness was achieved in the recognition of unknown samples. Then, the sensor array also exhibited excellent interference immunity and was reliable for real sample analysis. It provided a reference for pesticide efficient detection and food quality supervision.

As one of the important means for eukaryotic cells to maintain homeostasis, autophagy allows for transporting deformed biomacromolecules and damaged organelles to lysosome for digestion and degradation. The process of autophagy entails the merging of autophagosomes and lysosomes, culminating in the breakdown of biomacromolecules. This, in turn, leads to a change in lysosomal polarity. Therefore, fully understanding the changes of lysosomal polarity during autophagy is of significance to the study of membrane fluidity and enzymatic reaction. However, the shorter emission wavelength has greatly damaged the imaging depth, thus seriously limiting its biological application. Therefore, in this work, a near infrared in and out lysosome-targeted polarity-sensitive probe NCIC-Pola was developed. The fluorescence intensity of NCIC-Pola showed an approximate 1160-fold increase when the polarity decreased under two-photon excitation (TPE). In addition, the excellent fluorescence emission wavelength (692 nm) enabled the deep imaging analysis of scrap leather induced autophagy in vivo.

In this work, we have developed a nanocomposite consisting of horseradish peroxidase (HRP)-encapsulated 2D Zn-Co zeolite imidazole framework (ZIF) nanosheets strung on a ZnCoO nanowire array on a Ti support (denoted as 2D-Zn/Co-ZIF(HRP)|ZnCoO|Ti). This nanocomposite was then applied to constructing an electrochemical biosensor for detecting H2O2 derived from O2∙- released by mitochondria in living cells. This sensing platform shows excellent catalytic performance towards H2O2, attributable to the enzyme/metal-catalytic effect of HRP and Zn/Co-ZIF. The unique nano-string structure alleviates the aggregation of Zn/Co-ZIF nanosheets, readily exposes the catalytic active sites, protects the bioactivity of HRP, and reduces the charge/mass transfer pathway within Zn/Co-ZIF. The 2D-Zn/Co-ZIF(HRP)|ZnCoO|Ti biosensor offers two linear ranges of 0.2-10 μ M and 10-1100 μ M, a limit of detection of 0.082 μ M, a sensitivity of 3.3 mA mM-1 cm-2, good selectivity and stability over 40 days for H2O2 detection. After treating with specific mitochondrial complex inhibitors, the chronoamperometric results at the 2D-Zn/Co-ZIF(HRP)|ZnCoO|Ti confirmed complex I and III within the mitochondria electron transfer chain as the main electron leakage sites. This biosensor may contribute to the development of diagnostic health-care devices that shed light on the precaution and even treatment of oxidative stress diseases.

Heavy metal contamination has become a severe threat to dairy products through contaminated feed and the environment water. Among them, Pb(II) is highly toxic to the human body even under minimal exposure. Therefore, establishing a fast and sensitive Pb2+ detection technology is significant for rapid screening of vast number of dairy products. Hererin, we report the development of a sensitive and selective Pb(II) biosensor based on a solution-gated graphene transistor (SGGT) with the gate modified by Pb2+-dependent DNAzyme probes. It has also been explored that the DNAzymes working in simple binding mode integrate better with the SGGT than those working in normal catalytic mode, showing significantly stronger channel current responses and lower detection limit down to 0.39 μg/L (or 1.9 nM). Finally, the biosensor was practicably applied to the detection of lead ions in pure milk samples with a high recovery rate. We believe that this work reveals the best strategy for integrating metal ion dependent DNAzyme probes with SGGT sensing platforms to selectively and sensitively detect many metal ions.

Interstitial fluid glucose sensors have promising prospects in noninvasive glucose monitoring. However, the commonly used method of extracting interstitial fluid, reverse iontophoresis (RI), still remains to be optimized to solve problems such as insufficient extraction flux and skin irritation. To find the optimal RI conditions, in this study we explored the effects of multiple factors such as current frequency, duration, duty cycle and their interactions on extraction with the design of experiments (DOE) method. A multifunctional extraction and detection device was designed to control extraction conditions and measure the surface water content of the extraction electrode in situ and real time. A micro glucose monitoring device (MicroTED) combined with a cheap and flexible paper-based electrode was developed under the determined optimal extraction conditions. In on-body continuous glucose monitoring tests carried out to verify the performance of the device, the optimized conditions can facilitate stable extraction of up to 1.0 mg without any skin discomfort. The mean Pearson correlation coefficient between the measurement results of MicroTED and commercial glucometer is above 0.9. In the Clarke error grid analysis, all data points fell within Clarke error grid areas A and B, demonstrating the feasibility of further clinical application of the device.

Nanozyme-based colorimetric assays have shown great potential in the rapid and sensitive determination of pesticide residue in environment. However, the non-specific enzyme inhibition makes the assays generally lack of selectivity. In this study, we proposed a colorimetric sensing platform for the specific detection of the agricultural fungicide thiophanate-methyl (TM) based on its distinctive inhibitory effect on the nanozyme activity. Since TM contains the symmetric ethylenediamine- and bisthiourea-like groups, it displays strong affinity to the metal site, leading to a loss of the catalytic activity. Accordingly, a Cu-doped carbon nanozyme with excellent oxidase-like properties was designed, and the oxidation process of chromogenic substrate is promoted by Cu-induced generation of reactive oxygen species. Interestingly, the nanozyme activity can be directly and strongly restrained by TM, rather than other probably coexistent pesticides. Consequently, the as-proposed analytical method exhibits specific response toward TM and good linear relationship in the range of 0.2-15 μg mL-1 with a low limit of detection of 0.04 μg mL-1 (S/N = 3). Besides, a smartphone-assisted rapid detection was achieved through identifying the RGB value of the chromogenic system. This work provides a new nanozyme inhibition strategy for the specific detection of TM in environmental sample.

Interleukin-6 (IL-6) is a proinflammatory cytokine with a critical role in immune regulation and treatment of many diseases, including breast cancer. Herein, we developed a novel V2CTx MXene-based immunosensor for rapid and accurate IL-6 detection. The chosen substrate was V2CTx, a 2-dimensional (2D) MXene nanomaterial with excellent electronic properties. Prussian blue (Fe4[Fe(CN)6]3), used for its electrochemical properties, and spindle-shaped gold nanoparticles (Au SSNPs), used to combine with antibodies, were in-situ synthesized on the surface of the MXene. The in-situ synthesis ensures a firm chemical connection compared to other tags formed by a less stable physical absorption. Inspired by a sandwich ELISA test, the modified V2CTx tag was captured by the electrode surface with cysteamine to detect the analyte, IL-6, after being attached with a capture antibody (cAb). Benefiting from an increased surface area, an enhanced charge transfer rate, and a firm connection of the tag, this biosensor exhibited excellent analytical performance. The high sensitivity, high selectivity, and wide detection range covering the IL-6 level of both healthy individuals and breast cancer patients were obtained to meet clinical demands. Herein, this V2CTx MXene-based immunosensor is a potential therapeutic and diagnostic point-of-care alternative to routine ELISA IL-6 detection methods.

Respiratory viruses have long been a major cause of a global pandemic, emphasizing the urgent need for high-sensitivity diagnostic tools. Typical PCR technology can only determine the type of virus in the sample, which is unable to detect different variants of the same virus without costly and time-consuming gene sequencing. Here, we introduce a simple, fully enclosed, and highly integrated microfluidic system based on CRISPR/Cas12a and isothermal amplification techniques (LOC-CRISPR) that can specifically identify multiple common respiratory viruses and their variants. The LOC-CRISPR chip integrates viral nucleic acid extraction, recombinant polymerase amplification, and CRISPR/Cas12a cleavage reaction-based detection, contamination-free detection. In addition, the LOC-CRISPR chip was designed for multiplexed detection (two-sample input and ten-result outputs), which can not only detect the presence of SARS-CoV-2, H1N1, H3N2, IVB and HRSV but also differentiate the BA.1, BA.2, and BA.5 variants of SARS-COV-2. For clinical validation, the LOC-CRISPR chip was used to analyze 50 nasopharyngeal swab samples (44 positive and 6 negative) and achieved excellent sensitivity (97.8%) and specificity (100%). This innovative LOC-CRISPR system has the ability to quickly, sensitively, and accurately detect multiple target nucleic acid sequences with single-base mutations, which will further improve the rapid identification and traceability of respiratory viruses infectious diseases.

Desalting of biosamples is crucial for analytical techniques intolerant to abundant salts. However, there is no simple tool to monitor the desalting of low-volume biosamples so far. Here we developed a handheld capacitively coupled contactless conductivity detector (hC4D) as a miniaturized device to measure the conductivity of 75 μL biosamples. Polyether-ether-ketone (PEEK) tubing was selected as the sample reservoir for sample loading via a pipette. Another pipetting of air pushed the sample solution out of the tubing to recollect the sample. Owing to the low sample consumption and easy sample recollection, hC4D is advantageous for testing expensive biosamples, such as viruses and cells. In addition, the whole process of sample injection, conductivity measurement, recollection, and calibration of conductivity can be completed within 1 min. To verify the feasibility of hC4D, we monitored the desalting progress of gel filtration (GF) of 200 μL blood samples, ultrafiltration (UF) of 300 μL virus samples, and dialysis of 7 mL cell samples. Three rounds of GF and UF completely removed the salts but led to poor sample recovery. In contrast, low concentrations of residual salts remained and better recovery was achieved after two rounds of GF and UF. We further utilized the hC4D to monitor the dialysis and tuned the salt concentration in the cell sample, such that we maintained the viability of cells in a low conductivity environment. These results indicated that hC4D is a promising tool for optimizing the desalting procedure of low-volume biosamples.

This paper describes a novel signal processing method to characterize the activity of ion channels on a lipid bilayer system in a real-time and quantitative manner. Lipid bilayer systems, which enable single-channel level recordings of ion channel activities against physiological stimuli in vitro, are gaining attention in various research fields. However, the characterization of ion channel activities has heavily relied on time-consuming analyses after recording, and the inability to return the quantitative results in real time has long been a bottleneck to incorporating the system into practical products. Herein, we report a lipid bilayer system that integrates real-time characterization of ion channel activities and real-time response based on the characterization result. Unlike conventional batch processing, an ion channel signal is divided into short segments and processed during the recording. After optimizing the system to maintain the same characterization accuracy as conventional operation, we demonstrated the usability of the system with two applications. One is quantitative control of a robot based on ion channel signals. The velocity of the robot was controlled every second, which was around tens of times faster than the conventional operation, in proportion to the stimulus intensity estimated from changes in ion channel activities. The other is the automation of data collection and characterization of ion channels. By constantly monitoring and maintaining the functionality of a lipid bilayer, our system enabled continuous recording of ion channels over 2 h without human intervention, and the time of manual labor has been reduced from conventional 3 h to 1 min at a minimum. We believe the accelerated characterization and response in the lipid bilayer systems presented in this work will facilitate the transformation of lipid bilayer technology toward a practical level, finally leading to its industrialization.

Recombinase polymerase amplification (RPA) is one of the most promising diagnostic methods for pathogen detection, owing to the simplified isothermal amplification technique. Using one-step digital reverse transcription RPA (dRT-RPA) to detect viral RNA provides a fast diagnosis and absolute quantification. Here, we present a chip that purifies, digitalizes, and detects viral RNA of SARS-CoV-2 in a fully automated and sensitive manner. The chip purifies the RNA using the surface charge concept of magnet bead-RNA binding, then mixes the RNA with the amplification reagents, digitalizes the amplification mixture, and performs dRT-RPA. RNA-bead complex is transported among purification buffers that are separated by an oil phase. For reagent manipulation and mixing, a magnetic valve system is integrated on the chip, where an external magnet controls the reagent direction and time of addition. Besides, a novel vacuum system is suggested to drive and regulate the reagents into two fluid systems simultaneously in ∼2 min. We also developed a cost-effective way to perform fluorescent detection for dRT-RPA on chip by using EvaGreen® dye. With integrated heating and optical detection system, the on-chip dRT-RPA presents a sample-to-answer detection platform for absolute viral RNA quantitation in 37 min and a sensitivity as low as 10 RNA copies/μL. Hence, this platform is expected to be a useful tool for accurate and automated diagnosis of infectious diseases.

Scientific interest in the investigation and application of multifunctional nanomaterials in medical diagnostics has been increasing. The employment of magnetocatalytic immunoconjugates as both analyte-capturing agents and enzyme-like catalytic labels may enable rapid preconcentration and determination of relevant antigens. In this work, we synthesized and comprehensively characterized two types of noble metal-decorated magnetic nanocubes (MDMCs) which were subsequently applied in the one-step, sandwich nanozyme-linked immunosorbent assay (NLISA). Magnetic cores allow for rapid separation from complex samples of biological origin. The catalytically active shell composed of Au-decorated Pt or Ru can effectively mimic the activity of horseradish peroxididase, retaining at the same time the ability to form stable bioconstructs through self-assembly of thiolated ligands. As a result, hybrid multifunctional nanoparticles were synthesized and used to detect C-reactive protein (CRP) in serum samples. We have also paid considerable attention to the mechanistic studies of the formation of sandwich immunocomplexes with nanoparticle labels by means of immunoenzymatic methods and surface plasmon resonance. Analytical parameters of the Pt-MDMCs-labeled NLISA (detection limit LOD = 0.336 ng mL-1, recovery = 98.0%, linear response window covering two logarithmic units) turned out to be superior to the classical, one-step ELISA based on a horseradish peroxidase. In addition, our method offers further possibility of sensitivity adjustment by changing the parameters of magnetic preconcentration, together with good long-term stability of MDMCs conjugates and their good resistance to common interferences. We believe that the proposed simple synthetic protocol will guide a new approach to applying metal-decorated magnetic nanozymes as versatile and multifunctional labels for application in subsequent pre-analytical analyte concentration and immunoassays towards clinical applications.

Matrix metalloproteinase-9 (MMP-9) has been implicated in various tumor cell invasions and metastases. In light of the limitations of traditional methods for MMP-9 detection, we have constructed a novel biosensor depending on cucurbit[8]uril (CB[8]) -mediated host-guest interactions and a sacrificial iron metal-organic framework (FeMOF). Herein, MMP9-specific peptides modified on the gold bare electrode are bonded to the FeMOF@AuNPs@peptide complex through CB[8] addition. The connection between MMP9-specific peptides and signal peptides via CB[8] provides stability as well as enables the immobilization of FeMOF on the electrode surface. When Fe3+ from the FeMOF interacts with electrochemical buffer K4Fe(CN)6, Prussian blue will be generated on the gold electrode surface, and a significantly enlarged current response can be detected. However, in the presence of MMP-9, their peptide substrates are specifically cleaved at the site between serine (S) and Leucine (L), which causes an abrupt decrease in the electrochemical signal. The change of signal can reflect MMP-9 concentration. This sensor can reach an ultrahigh sensitivity with a wide detection range of 0.5 pg⋅mL-1 to 500 ng⋅mL-1 and a low detection limit of 1.30 pg⋅mL-1. Importantly, this sensor is very simple, relying solely on self-sacrificial label of FeMOF, rather than complex functional materials. Additionally, it has been well used in serum samples, showing attractive potential for practical applications.

Point-of-care testing of pathogens is vital for prevention of food poisoning. Herein, a colorimetric biosensor was elaborately developed to rapidly and automatically detect Salmonella in a sealed microfluidic chip with one central chamber for housing immunomagnetic nanoparticles (IMNPs), bacterial sample and immune manganese dioxide nanoclusters (IMONCs), four functional chambers for housing absorbent pad, deionized water and H2O2-TMB substrate, and four symmetric peripheral chambers for achieving fluidic control. Four electromagnets were placed under peripheral chambers and synergistically controlled to manipulate their respective iron cylinders at the top of these chambers for deforming these chambers, resulting in precise fluidic control with designated flowrate, volume, direction and time. First, the electromagnets were automatically controlled to mix IMNPs, target bacteria and IMONCs, resulting in the formation of IMNP-bacteria-IMONC conjugates. Then, these conjugates were magnetically separated by a central electromagnet and the supernatant was directionally transferred to the absorbent pad. After these conjugates were washed by deionized water, the H2O2-TMB substrate was directionally transferred to resuspend the conjugates and catalyzed by the IMONCs with peroxidase-mimic activity. Finally, the catalysate was directionally transferred back to its initial chamber, and its color was analyzed by the smartphone APP to determinate bacterial concentration. This biosensor could detect Salmonella quantitatively and automatically in 30 min with a low detection limit of 101 CFU/mL. More importantly, the whole bacterial detection procedure from bacterial separation to result analysis was achieved on a sealed microfluidic chip through multiple electromagnet synergistic control, and this biosensor has great potential for point-of-care testing of pathogens without cross contaminations.

Laser-scribed graphene (LSG), a promising electrode material has attracted special research interest in recent years. Here, the fabricating process-electrochemical property correlation of laser-scribed graphene (LSG) devices was discussed emphatically and a pertinent optimization was performed to achieve better electroanalytical performance. Experiment results indicated that the laser scribing technique possessed great process latitude and reducing laser power and scribing speed facilitated fabricating high-quality graphene electrodes. Benefiting from its binder-free 3D porous network structure and high active/geometric area ratio, the optimized LSG electrode was superior to the screen-printed carbon electrode (SPCE) on electrochemical performance in the [Fe(CN)6]3-/4- redox system. Integrating the LSG electrode with a homemade hand-held detector, a portable electrochemical sensing platform with smartphone readout was developed. It realized a specific detection of H2O2 (linear range: 0.02-3.4 mM, sensitivity: 24.56 μA mM-1 cm-2), glucose (linear range: 0.04-4.0 mM, sensitivity: 16.35 μA mM-1 cm-2) by directly decorating biological enzymes without artificial redox mediator and featured a satisfactory comprehensive performance. The constructed immunosensor for tumor necrosis factor-α exhibited a wide linear range (2-500 pg mL-1) and a 4.3-fold enhancement in sensitivity compared with that of SPCE. With satisfactory selectivity, reproducibility, and sensitivity, the developed smartphone-based electrochemical sensing platform held great promise in accurate detection on the spot. This work also provided a significant reference for tailoring binder-free carbonaceous electrode materials toward the desired application.

DNA methylation, a kind of epigenetic alteration, plays a vital role in tumorigenesis and offers a new class of targets for cancer treatment. DNA hypermethylation at the E-Box site (CACGTG, -288 bp) in the SLC22A2 promoter was related to multidrug resistance of renal cell carcinoma (RCC), which can provide the target for both treatment and monitoring. Herein, we developed a novel phosphorothioated primer based loop-mediated isothermal amplification (PS-LAMP) assay to detect circulating tumor DNA (ctDNA) methylation levels in E-Box sites in tumor tissue, urine, and plasma samples from patients with RCC. Bisulfite treatment converted methylated/unmethylated discrepancy to a single base discrepancy (C/U). PS-LAMP amplified the templates to a tremendous amount. One-step strand displacement (OSD) probe provided single base resolution in amplified products and finally realized the specific site methylation detection. Our proposed method provided a linear range from 0% to 100% for methylation levels and was available in samples at low concentrations (102 copies/μL). Visually observable colorimetric detection can be achieved by incorporating the OSD probe with gold nanoparticles (AuNP). Our assay performed better than traditional methods in biological samples with low ctDNA concentration. Further, we found a potential consistency of methylation levels between tumor tissue and plasma sample from the same patient (Spearman's ρ = 0.886, P = 0.019, n = 6). In general, this work provides a PS-LAMP assay combining OSD probes for site-specific methylation detection in various biological samples, offering a method for noninvasive detection.

Efficient detection of related markers is significant for the early screening of COVID-19. Near infrared (NIR) light excited up-conversion fluorescence probes are ideal for biosensing but limited by the low luminescence efficiency. In this work, a novel highly stable opal photonic crystal (OPC) structure was designed to provide an OPC effect for up-conversion fluorescence enhancement, and sensitive Novel Coronavirus IgG up-conversion FRET-based sensor was further constructed. For the problems of water stability and mechanical stability of polymer OPC which cannot be solved for a long time, polymer spray combined with a flipped OPC film strategy is presented. Fragmented size OPC film was firmly fixed by polymer modification layer, which gave large size OPC film great water stability, mechanical stability and bending performance without affecting the fluorescence enhancement property. On this basis, the up-conversion emission intensity was enhanced significantly, and fluorescence resonant energy transfer (FRET) based Novel Coronavirus IgG antibody sensor was constructed. Monolayer up-conversion nanoparticles (UCNPs) on the surface of the polydopamine (PDA)/OPC film can make the fluorescent signal more sensitive, and effectively reduce the detection limit. The test device integrating NIR excitation and mobile phone realized the visual fast detection, showing remarkable sensing performance for COVID-19 antibodies with the limit of detection (LOD) of 0.1 ng mL-1. This detection platform will provide a more effective tool for early detection of the novel coronavirus.

The rapid and accurate assessment of glucose concentration has been demonstrated to play a significant role in human health, such as the diagnosis and treatment of diabetes, pharmaceutical research and quality monitoring in the food industry, necessitating further development of the performance for glucose sensor especially at low concentrations. However, glucose oxidase-based sensors suffer from crucial restriction in bioactivity because of their poor environmental tolerance. Recently, catalytic nanomaterials with enzyme-mimicking activity, known as nanozymes, have gained considerable interest to overcome the drawback. In this scenario, we report an inspiring surface plasmon resonance (SPR) sensor for non-enzymatic glucose detection employing ZnO nanoparticles and MoSe2 nanosheets composite (MoSe2/ZnO) as sensing film, featuring desirable advantages of high sensitivity and selectivity, lab-free and low cost. The ZnO was used to specifically recognize and bind glucose, and further signal amplification was realized by incorporating of MoSe2 owing to its larger specific surface area and favorable bio-compatibility, as well as high electron mobility. These unique features of MoSe2/ZnO composite film result in an obvious improvement of sensitivity for glucose detection. Experimental results show that the measurement sensitivity of the proposed sensor could reach 72.17 nm/(mg/mL) and a detection limit of 4.16 μg/mL by appropriately optimizing the componential constitutions of MoSe2/ZnO composite. In addition, the favorable selectivity, repeatability and stability are demonstrated as well. This facile and cost-effective work provides a novel strategy for constructing high-performance SPR sensor for glucose detection and a prospective application in biomedicine and human health monitoring.

Parkinson's disease (PD) is a neurodegenerative disorder caused by various factors such as neuroinflammation, oxidative stress, mitochondrial dysfunction, and neuronal apoptosis. Recent studies have shown that H2S supplementation reverses neuronal loss and mitigates motor deficits in PD patients through anti-inflammatory, antioxidant, improved mitochondrial function and proautophagic. Therefore, the discovery and use of H2S donors may be an exciting and intriguing strategy for the treatment of PD. Herein, we report a red emission mitochondria-targetable fluorescent probe, Rho-H2S, which can specifically and sensitively detect H2S with a limit of detection of 62.5 nM. Bioimaging experiments have shown that the probe has excellent mitochondrial targeting and good imaging capabilities for the detection of exogenous and endogenous H2S in cells. More importantly, based on the Rho-H2S probe, we first confirmed the sulforaphane (SFN) among 15 glucosinolate and isothiocyanate compounds from cruciferous vegetables with an outstanding ability to release H2S and we further proved that SFN could alleviate the symptoms of PD in vivo. All results demonstrate that Rho-H2S could be an effective tool for screening H2S donors and can contribute to the development of new therapeutic strategies for PD.

A surface plasmon resonance (SPR) optical fiber sensor with multimode-coreless-multimode (MNM) structure was developed, which modified by L-glutamine-binding protein (QBP) for detection of L-glutamine (Gln). The QBP was immobilized on the surface of gold films by chemical cross-linking and exhibited a binding affinity for L-glutamine. The conformation of QBP can be changed from the "open" to the "closed", which led to a red-shift of the SPR peak when QBP bounded to L-glutamine. There was a good linear correlation between is a dependence of the SPR peak on and the concentration of L-glutamine concentration in the range 10-100 μM, with a sensitivity of 10.797nm/log10[Gln] for L-glutamine in the in vitro embryo culture (IVC) medium environment, and the limit of detection (LOD) is 1.187 μM. This QBP-modified MNM structure optical fiber SPR sensor provides a new idea for the developmental potential assessment of embryos in the process of in vitro embryo culture.

Detection of cardiac troponin I (cTnI) plays a critical role in diagnosing acute myocardial infarction (AMI). In this report, a new kind of spherical AuPt@FexOy core@shell nanoparticles (termed as AuPt@FexOy NPs) were one-pot synthesized by a redox interaction-engaged strategy (RIES) without the addition of any surfactants or reducing agents. The as-synthesized AuPt@FexOy NPs not only retain the plasmonic activity of gold nanoparticles (AuNPs), but also possess excellent catalytic activities of platinum nanoparticles (PtNPs) and FexOy nanoclusters. The features of AuPt@FexOy NPs enable greatly enhance the colorimetric detection sensitivity of lateral flow immunoassay (LFIA) through integrating AuPt@FexOy NPs labeling procedure and catalyzing oxidation of chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) signal amplification strategy. The as-developed colorimetric LFIA (termed as AuPt@FexOy-LFIA) exhibits the limit of detection (LOD) as 26.0 pg mL-1 cTnI under the TMB signal amplification mode. In particular, the detection results of cTnI in 40 clinical seral samples by AuPt@FexOy-LFIA are correlated well with those of cTnI in the same samples by commercial enzyme-linked immunosorbent assay (ELISA) detection kit (R2 = 0.97, slope = 1), demonstrating the highly reliable analytical performance and good application prospect of AuPt@FexOy-LFIA.

A biosensor was engineered to enable the study of the novel quorum sensing molecule (QSM), 3,5-dimethylpyrazin-2-ol (DPO), employed by Vibrio cholerae to regulate biofilm formation and virulence factor production. Investigations into bacterial quorum sensing (QS), a form of communication based on the production and detection of QSMs to coordinate gene expression in a population dependent manner, offer a unique window to study the molecular underpinnings of microbial behavior and host interactions. Herein, we report the construction of an engineered microbial whole-cell bioluminescent biosensing system that incorporates the recognition of the VqmA regulatory protein of Vibrio cholerae with the bioluminescent reporting signal of luciferase for the selective, sensitive, stable, and reproducible detection of DPO in a variety of samples. Importantly, using our newly developed biosensor our studies demonstrate the detection of DPO in rodent and human samples. Employing our developed biosensor should help enable elucidation of microbial behavior at the molecular level and its impact in health and disease.

The practical application of DNA biosensors is impeded by numerous limitations in complicated physiological environments, particularly the susceptibility of common DNA components to nuclease degradation, which has been recognized as a major barrier in DNA nanotechnology. In contrast, the present study presents an anti-interference and reinforced biosensing strategy based on a 3D DNA-rigidified nanodevice (3D RND) by converting a nuclease into a catalyst. 3D RND is a well-known tetrahedral DNA scaffold containing four faces, four vertices, and six double-stranded edges. The scaffold was rebuilt to serve as a biosensor by embedding a recognition region and two palindromic tails on one edge. In the absence of a target, the rigidified nanodevice exhibited enhanced nuclease resistance, resulting in a low false-positive signal. 3D RNDs have been proven to be compatible with 10% serum for at least 8 h. Once exposed to the target miRNA, the system can be unlocked and converted into common DNAs from a high-defense state, followed by polymerase- and nuclease-co-driven conformational downgrading to achieve amplified and reinforced biosensing. The signal response can be improved by approximately 700% within 2 h at room temperature, and the limit of detection (LOD) is approximately 10-fold lower under biomimetic conditions. The final application to serum miRNA-mediated clinical diagnosis of colorectal cancer (CRC) patients revealed that 3D RND is a reliable approach to collecting clinical information for differentiating patients from healthy individuals. This study provides novel insights into the development of anti-interference and reinforced DNA biosensors.

β-glucosyltransferase (β-GT) can specifically catalyze the conversion of 5-hydroxymethylcytosine (5-hmC) to 5-glucosylhydroxy methylcytosine (5-ghmC), and it is associated with the control of phage-specific gene expression by affecting transcription process in vivo and in vitro. The current strategies for β-GT assay usually involve expensive equipment, laborious treatment, radioactive hazard, and poor sensitivity. Here, we report a Spinach-based fluorescent light-up biosensor for label-free measurement of β-GT activity by utilizing 5-hmC glucosylation-initiated rolling circle transcription amplification (RCTA). We design a 5-hmC-modified multifunctional circular detection probe (5-hmC-MCDP) that integrates the functions of target-recognition, signal transduction, and transcription amplification in one probe. The introduction of β-GT catalyzes 5-hmC glucosylation of 5-hmC-MCDP probe, protecting the glucosylated 5-mC-MCDP probe from the cleavage by MspI. The remaining 5-hmC-MCDP probe can initiate RCTA reaction with the aid of T7 RNA polymerase, generating tandem Spinach RNA aptamers. The tandem Spinach RNA aptamers can be lightened up by fluorophore 3,5-difluoro-4-hydroxybenzylidene imidazolinone, facilitating label-free measurement of β-GT activity. Notably, the high specificity of MspI-catalyzed cleavage of nonglucosylated probe can efficiently inhibit nonspecific amplification, endowing this assay with a low background. Due to the higher efficiency of RCTA than the canonical promoter-initiated RNA synthesis, the signal-to-noise ratio of RCTA is 4.6-fold higher than that of linear template-based transcription amplification. This method is capable of sensitively detecting β-GT activity with a limit of detection of 2.03 × 10-5 U/mL, and it can be used for the screening of inhibitors and determination of kinetic parameters, with great potential in epigenetic research and drug discovery.

MicroRNAs are likely to be a next-generation clinical biomarker for many diseases. While gold-standard technologies, e.g., reverse transcription-quantitative polymerase chain reaction (RT-qPCR), exist for microRNA detection, there is a need for rapid and low-cost testing. Here, an emulsion loop-mediated isothermal amplification (eLAMP) assay was developed for miRNA that compartmentalizes a LAMP reaction and shortens the time-to-detection. The miRNA was a primer to facilitate the overall amplification rate of template DNA. Light scatter intensity decreased when the emulsion droplet got smaller during the ongoing amplification, which was utilized to moitor the amplification non-invasively. A custom low-cost device was designed and fabricated using a computer cooling fan, a Peltier heater, an LED, a photoresistor, and a temperature controller. It allowed more stable vortexing and accurate light scatter detection. Three miRNAs, miR-21, miR-16, and miR-192, were successfully detected using the custom device. Specifically, new template and primer sequences were developed for miR-16 and miR-192. Zeta potential measurements and microscopic observations confirmed emulsion size reduction and amplicon adsorption. The detection limit was 0.01 fM, corresponding to 2.4 copies per reaction, and the detection could be made in 5 min. Since the assays were rapid and both template and miRNA + template could eventually be amplified, we introduced the success rate (compared to the 95% confidence interval of the template result) as a new measure, which worked well with lower concentrations and inefficient amplifications. This assay brings us one step closer to allowing circulating miRNA biomarker detection to become commonplace in the clinical world.

The working efficiency of traditional 3D DNA nanomachines is extremely restricted due to the complex DNA components modified on nanoparticles in the same spatial height. Herein, an ultrafast dual-layer 3D DNA nanomachine (UDDNM) based on catalytic hairpin assembly (CHA) was developed by assembling two different lengths of hairpin DNA on the surface of gold nanoparticles, the long hairpin 1 (H1), to capture the trigger, and the short hairpin 2 (H2), as the signal probe, to recycle the trigger. Compared to the traditional single-layer 3D DNA nanomachine, the dual-layer 3D DNA nanostructure greatly enhances the effective collision between trigger and targeted DNA probe, H1, since the H1 located in outer layer would react with the trigger, inhibiting the invalid collision between the trigger and residual DNA component, H2, and remarkably decreasing the steric hindrance associated with the nucleic acids layer around the nanoparticles. Especially, when the distance of two layers was fixed at 3 nm, the corresponding UDDNM could accomplish the overall reaction only in 3 min with a dramatically high initial rate of up to 5.93 × 10-7 M s-1, which was at least 5-fold beyond that of the typical single-layer 3D DNA nanomachines. As a proof of concept, the described UDDNM was successfully applied in ultrasensitive fluorescence detection and sensitive intracellular imaging of miRNA-21. Consequently, our strategy, based on the creation of dual-layer 3D DNA nanostructure, may create a new approach to designing the next generation of DNA nanomachine and has enormous potential for applications in bio-analysis, logic gate operations, and clinical diagnoses.

The microfluidic technology provides an ideal platform for in situ screening of enzyme inhibitors and activators from natural products. This work described a surface-modified ITO glass-PDMS hybrid microfluidic chip for evaluating thrombin interaction with its potential inhibitors by fluorescence imaging and matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI). The fluorescence-labeled substrate was immobilized on a conductive ITO glass slide coated with gold nanoparticles/thiol-β-cyclodextrin modified TiO2 nanowires (Au-β-CD@TiO2 NWs) via Au-S bonds. A PDMS microchannel plate was placed on top of the modified ITO slide. The premixed solutions of thrombin and candidate thrombin inhibitors were infused into the microchannels to form a microreactor environment. The enzymatic reaction was rapidly monitored by fluorescence microscopy, and MALDI MS was used to validate and quantify the enzymatic hydrolysate of thrombin to determine the enzyme kinetic process and inhibitory activities of selected flavonoids. The fluorescence and MALDI MS results showed that luteolin, cynaroside, and baicalin have good thrombin inhibitory activity and their half-maximal inhibitory concentrations (IC50) were below 30 μM. The integration of fluorescence imaging and MALDI MSI for in situ monitoring and quantifying the enzymatic reaction in a microfluidic chip is capable of rapid and accurate screening of thrombin inhibitors from natural products.

Previous studies have identified that Geobacter sulfurreducens has three different electron transfer pathways for respiration, and it switches between these pathways to adapt to the redox potential of its electron acceptor. However, only a small fraction of the electron carriers from each pathway have been identified. In this study, we combined electrochemical and gene expression data to identify electron carriers in the inner membrane, periplasm, outer membrane, and exterior of the cell that may be induced by the use of the three different electron transfer pathways. Cyclic voltammetry was performed on thin biofilms grown on anodes poised at different redox potentials, providing a quantitative assessment of the relative use of three electron-transfer pathways in each condition (catalytic midpoint potentials (EKAs) of -0.227 V [Low], -0.15 V [Medium], -0.1 V [High] vs. SHE). Transcriptomic analyses as a function of electrochemical signals or fumarate utilization showed differential induction in inner membrane (Medium: cbcL), periplasmic (Low: ppcB/ppcE, Medium: ppcA), outer membrane (Low: extA/extC, Medium: extJ/extK, Fumarate: extF/extG), and extracellular (Medium: omcZ, High/Fumarate: omcS/omcT) cytochromes, suggesting the pathway signals are associated with complex transcriptomic responses in genes across the electron transfer pathway. Our method combining electrochemical modeling and transcriptomics could be adapted to better understand electron transport in other electroactive organisms with complex metabolisms.

On-site analytical techniques continue being developed with advances in modern technology. To demonstrate the applicability of four-dimensional printing (4DP) technologies in the direct fabrication of stimuli-responsive analytical devices for on-site determination of urea and glucose, we used digital light processing three-dimensional printing (3DP) and 2-carboxyethyl acrylate (CEA)-incorporated photocurable resins to fabricate all-in-one needle panel meters. When adding a sample having a value of pH above the pKa of CEA (ca. 4.6-5.0) into the fabricated needle panel meter, the [H+]-responsive layer of the needle, printed using the CEA-incorporated photocurable resins, swelled as a result of electrostatic repulsion among the dissociated carboxyl groups of the copolymer, leading to [H+]-dependent bending of the needle. When coupled with a derivatization reaction (urease-mediated hydrolysis of urea to decrease [H+]; glucose oxidase-mediated oxidization of glucose to increase [H+]), the bending of the needle allowed reliable quantification of urea or glucose when referencing pre-calibrated concentration scales. After method optimization, the method's detection limits for urea and glucose were 4.9 and 7.0 μM, respectively, within a working concentration range from 0.1 to 10 mM. We verified the reliability of this analytical method by determining the concentrations of urea and glucose in samples of human urine, fetal bovine serum, and rat plasma with spike analyses and comparing the results with those obtained using commercial assay kits. Our results confirm that 4DP technologies can allow the direct fabrication of stimuli-responsive devices for quantitative chemical analysis, and that they can advance the development and applicability of 3DP-enabling analytical methods.

The rapid and sensitive detection of pathogenic viruses is important for controlling pandemics. Herein, a rapid, ultrasensitive, optical biosensing scheme was developed to detect avian influenza virus H9N2 using a genetically engineered filamentous M13 phage probe. The M13 phage was genetically engineered to bear an H9N2-binding peptide (H9N2BP) at the tip and a gold nanoparticle (AuNP)-binding peptide (AuBP) on the sidewall to form an engineered phage nanofiber, M13@H9N2BP@AuBP. Simulated modelling showed that M13@H9N2BP@AuBP enabled a 40-fold enhancement of the electric field enhancement in surface plasmon resonance (SPR) compared to conventional AuNPs. Experimentally, this signal enhancement scheme was employed for detecting H9N2 particles with a sensitivity down to 6.3 copies/mL (1.04 × 10-5 fM). The phage-based SPR scheme can detect H9N2 viruses in real allantoic samples within 10 min, even at very low concentrations beyond the detection limit of quantitative polymerase chain reaction (qPCR). Moreover, after capturing the H9N2 viruses on the sensor chip, the H9N2-binding phage nanofibers can be quantitatively converted into plaques that are visible to the naked eye for further quantification, thereby allowing us to enumerate the H9N2 virus particles through a second mode to cross-validate the SPR results. This novel phage-based biosensing strategy can be employed to detect other pathogens because the H9N2-binding peptides can be easily switched with other pathogen-binding peptides using phage display technology.

Recombinase polymerase amplification (RPA) running at 37-42 °C is fast, efficient and less-implemented; however, the existing technologies of nucleic acid testing based on RPA have some limitations in specificity of single-base recognition and multiplexing capability. Herein, we report a highly specific and multiplex RPA-based nucleic acid detection platform by combining flap endonuclease 1 (FEN1)-catalysed invasive reactions with RPA, termed as FEN1-aided RPA (FARPA). The optimal conditions enable RPA and FEN1-based fluorescence detection to occur automatically and sequentially within a 25-min turnaround time and FARPA exhibits sensitivity to 5 target molecules. Due to the ability of invasive reactions in discriminating single-base variation, this one-pot FARPA is much more specific than the Exo probe-based or CRISPR-based RPA methods. Using a universal primer pair derived from tags in reverse transcription primers, multiplex FARPA was successfully demonstrated by the 3-plex assay for the detection of SARS-CoV-2 pathogen (the ORF1ab, the N gene, and the human RNase P gene as the internal control), the 2-plex assay for the discrimination of SARS-CoV-2 wild-type from variants (Alpha, Beta, Epsilon, Delta, or Omicrons), and the 4-plex assay for the screening of arboviruses (zika virus, tick-borne encephalitis virus, yellow fever virus, and chikungunya virus). We have validated multiplex FARPA with 103 nasopharyngeal swabs for SARS-CoV-2 detection. The results showed a 100% agreement with RT-qPCR assays. Moreover, a hand-held FARPA analyser was constructed for the visualized FARPA due to the switch-like endpoint read-out. This FARPA is very suitable for pathogen screening and discrimination of viral variants, greatly facilitating point-of-care diagnostics.

Enrofloxacin (ENR) and its metabolite Ciprofloxacin (CIP) are both a class of fluoroquinolone antibiotics effective against a broad-spectrum microbial infection. Recent surge in the consumption of CIP and ENR has been linked to increased cases of drug-resistant pathogens. This is due to the fact that the antibiotic residues remain in milk, meat, soil and environmental water for a prolonged duration. Although gold standard methods such as LC-MS are sensitive, they suffer from expensive operation and maintenance cost, and would need dedicated facilities and tedious sample preparation steps. Such limitations make on site detection impossible for regulatory bodies in developing countries. To address this issue, we developed a portable device that can detect the presence of CIP and ENR antibiotics in the range of parts per billion (ppb) concentrations accurately. It consists of a polyaniline (PAni) coated U-bent optical fiber with anti-ENR/CIP antibody immobilized on the polymer surface. The sensor relies on the principle of evanescent wave absorbance by antigen-antibody complex. The sensor showed limit of detection (LOD) of 1 ppb with a linear range of operation from 1 ppb to 500 ppb (R2 = 0.96-0.99) in lake water, waste water treatment plant effluent, urine, blood serum, milk and meat samples. The recovery of the sensor ranges from 88% to 120% indicating reasonable accuracy. The sensor has excellent selectivity towards CIP and ENR and showed stability for four weeks indicating its field deployability and robustness. The portable sensor is scalable and contract has been given to an industry partner to mass manufacture the device.

As a tumor biomarker, the accumulation of amyloid β oligomers (Aβo) in the brain has been suggested as a key feature in the pathogenesis and progression of Alzheimer's disease (AD). In this work, we designed a novel photoelectrochemical (PEC) and electrochemiluminescence resonance energy transfer (ECL-RET) dual-mode biosensor to achieve ultra-sensitive detection of Aβo. Specifically, the electrode surface modified Carbon Dots (C Dots) and the electrodeposited polyaniline (PANI) film formed a Z-scheme heterojunction reversing the photocurrent signal, and then the Aβo specific recognition peptide was attached to the surface via amide bonding between the amino group of PANI and carbonyl group of peptide. After that, in the presence of CdTe labeled specific recognition aptamer for Aβ (CdTe-Apt), Aβo was captured to construct a sandwich-type biosensor and exhibited a significantly enhanced cathodic photocurrent response because the formed dual Z-scheme heterojunction promoted charge separation efficiency. Interestingly, the proposed biosensor also caused a ratiometric change in the ECL intensity at 555 nm and 640 nm. Therefore, the developed biosensor achieved dual-mode detection of Aβo, where the PEC detection range of Aβo was from 10 fM to 0.1 μM (with a detection limit of 4.27 fM) and the ECL method provided a linear detection range of 10 fM to 10 nM (with a detection limit of 6.41 fM). The stability and reliability of the experimental results indicate that this has been a promising biosensing pattern and could be extended to the analysis of other biomarkers.

Recently, up-conversion luminescent (UCL) materials have caught extensive sight on account of their excellent biocompatibility and weak automatic fluorescence background, but the low optical signal makes researchers shy away. Organic dye-sensitized UCL materials can improve the low optical signal drawback of UCL and rejuvenate it with adjustable optical properties and unique antenna effects. In this work, an efficient, simple and selective electrochemiluminescence (ECL) sensing platform was developed for determination of enrofloxacin (ENR). 3,4,9,10-perylene tetracarboxylic acid (PTCA) was successfully used as an "antenna" to improve the ECL performance of the UCL nanoparticles (PEI-NaYF4: Yb, Er) due to its appropriate excitation spectrum position and superior electron transfer rate. The specific recognition function of the aptamer enabled the sensor to eliminate the interference from conspecific impurity. In the presence of ENR, the specific combination of ENR with aptamer made the aptamer fall from surface of the electrode, thus we could see a considerable enhancement of signal. Under the most favourable conditions, the aptasensor based on antenna effect displayed a wide detection range (1.0 × 10-14∼1.0 × 10-6 M), low limit of detection (LOD = 3.0 × 10-15 M) and receivable recoveries (96.0%-102.4%) during water samples analysis. At this point, antenna effect provides a powerful strategy to expand the application of UCL in the field of ECL biosensing.

The measurement of tumor biomarker levels is of great significance for early diagnosis of breast cancer. The combination diagnosis of multiple tumor biomarkers will significantly improve the accuracy of early diagnosis. Here, we successfully developed a dual-ratio fluorescent sensing platform for the detection of breast cancer biomarkers (PTK7, miRNA-21) using single excitation triple-signal detection. Introducing three types of fluorescence nanomaterials with narrow emission peaks and long Stokes shift as signal markers, the three peaks (430 nm, 530 nm and 640 nm) of which do not interfere with each other in fluorescence spectra under a single excitation (360 nm). The sensing platform linked aptamer (apt) modified green fluorescence quantum dots (gQDs-apt1) and aptamer modified red fluorescence quantum dots (rQDs-apt2) to Fe3O4-cDNA1 and Fe3O4-cDNA2, respectively, via base complementary pairing with aptamer molecules. When PTK7/miRNA-21 is present in the system, gQDs-apt1/rQDs-apt2 bound to the Fe3O4 MNPs surface will be released to recover fluorescence. Upon DNase I digestion of free apt1 and apt2, the target molecules will be released to bind to gQDs-apt1/rQDs-apt2 for signal amplification. After magnetic separation, PTK7 and miRNA-21 can be quantified using the fluorescence intensity ratio of gQDs with bCDs and rQDs with bCDs at a single excitation of 360 nm wavelength. This method has high sensitivity, good selectivity, and can quantify both PTK7 and miRNA-21 simultaneously with an LOD of 0.426 ng mL-1 and 0.072 nM, respectively. Additionally, the sensing platform was used for serum detection of health man and breast cancer patients with satisfactory results.

The efficiency of aggregation-induced electrochemiluminescence (AIECL) in tetraphenylethene (TPE) derivatives were significantly enhanced by combining the regulation of molecular structure and solvent. Firstly, the linear increase of the benzene ring resulted in enhanced molecular aggregation and promoted the electrochemical reaction of the anode, due to increased molecular conjugation and higher lowest unoccupied molecular orbital (LUMO) and highest occupied molecular orbital (HOMO). The ECL efficiency of 4,4,4,4-(Ethene-1,1,2,2-tetrayl) tetrakis (([1,1,4,1-terphenyl]-4-carbaldehyde)) (T3) nanoparticles (NPs) with more benzene rings were 5558 times that of 4,4,4,4-(ethene-1,1,2,2-tetrayl) tetrabenzaldehyde (T1) NPs, and its relative ECL efficiency of T3 NPs reached 55.58% compared to the [Ru (bpy)3]2+/tripropylamine (TPrA) system. Furthermore, solvents with different polarities played a crucial role in modulating the degree of molecular aggregation, which also effectively facilitated the AIE process and reduced the aggregation-caused quenching (ACQ) effect caused by excessively dense aggregation. This aspect had often been overlooked in previous AIECL studies. T3 NPs demonstrated optimal ECL performance at fw = 70% (fw was the H2O content in tetrahydrofuran (THF)/H2O), and its ECL efficiency was 232 times greater than fw = 100% and 1853 times greater than fw = 0%. Additionally, it was found that melatonin (MT), one of the hormones widely used to treat insomnia, exhibited antioxidant and free radical scavenging properties, which exerted a significant quenching effect on the ECL of the T3 NPs/TPrA system. Consequently, a sensitive sensing platform was developed for MT with a low detection limit of 8.78 × 10-10 mol L-1, which promoted the application of AIECL in efficient ultra-sensitive biosensing.

Programmed death ligand-1 (PD-L1) can enhance the immune tolerance of tumor cells by suppressing the activity of T-cells, and is one of the culprits that lead to the immune escape of tumor cells. Thus, the sensitive and portable detection of PD-L1 levels is essential for many types of tumor prognosis. Herein, a novel dual-mode analytical device for the ultrasensitive detection of PD-L1 has been developed. In this configuration, an advanced organic-inorganic hybrid material of poly(3,4-ethylenedioxythiophene) -BiOBr0.8I0.2 is designed as photocathode to enhance the photogenerated electron migration efficiency of the MWCNTs/SnS2-photoanode by external circuit, amplifying cathodic photocurrent without extra energy supply. The PD-L1 aptamer is loaded on the photocathode surface to ensure selectivity. The obtained sensing platform can achieve highly sensitive and specific detection of PD-L1 in complex environment, with a low detection limit of 0.29 pg mL-1. On the other hand, electrochromic material Prussian blue (PB) and MWCNTs/SnS2 are integrated to fabricate a portable sensing chip for PD-L1. Under illumination, photogenerated electrons of MWCNTs/SnS2 are injected into Prussian blue, and the blue PB is reduced to white product, indicating the concentration of PD-L1, without need of other instrument. This self-powered photoelectrochemical and visual analysis system has good practicability and is a promising clinical diagnosis tool.

The detection and comparison of the amount of superoxide anion (O2.-) released by different complexes in mitochondrial electron transport chain (ETC) can locate the main electron leakage sites in mitochondria. In order to realize this, we designed an ultrasensitive photoelectrochemical (PEC) sensor by in situ hydrothermal growth of MnO2 nanosheets on Co3O4 nanowires array modified Ti substrate (NWA|Ti). Due to the formation of a core-shell p-n heterojunction with high specific surface area, tight surface contact and plentiful oxygen vacancies (OVs), MnO2@Co3O4 NWA|Ti possesses a strong visible light absorption, high charges transfer and separation ability. The proposed PEC sensor exhibited a wide linear range of 0.1-50000 nM and a low detection limit of 0.025 nM towards H2O2. Due to the rapid conversion of O2.- to H2O2 inside mitochondria, the PEC sensor can indirectly monitor the electron leakage in the ETC. Specifically, four selected mitochondrial inhibitors specifically inhibited the corresponding complex in mitochondria extracted from living HepG2 cells (hepatocellular carcinoma cells), and the H2O2 levels converted from O2.- was measured by the PEC sensor. It is evident that IQ (ubiquinone binding) site of complex I and Qo (ubiquinol oxidation) site of complex III are the key sites at which electron leakage occurred. This study could provide meaningful information for the diagnosis and treatment of certain disease caused by oxidative stress due to the electron leakage.

Violet phosphorene (VP) have been proved to be more stable than black phosphorene, but few reports for its application in electrochemical sensors. In this study, a highly-stable VP decorated with phosphorus-doped hierarchically porous carbon microsphere (PCM) with multiple enzyme-like activities as a nanozyme sensing platform for portable intelligent analysis of mycophenolic acid (MPA) in silage with machine learning (ML) assistance is successfully fabricated. The pore size distribution on the PCM surface is discussed using N2 adsorption tests, and morphological characterization indicates that the PCM is embedded in the layers of lamellar VP. The affinity of the VP-PCM nanozyme obtained under the guidance of the ML model reaches Km = 12.4 μmol/L for MPA. The VP-PCM/SPCE for the efficient detection of MPA exhibits high sensitivity, a wide detection range of 2.49 μmol/L - 71.14 μmol/L with a low limit of detection of 18.7 nmol/L. The proposed ML model with high prediction accuracy (R2 = 0.9999, MAPEP = 0.0081) assists the nanozyme sensor for intelligent and rapid quantification of MPA residues in corn silage and wheat silage with satisfactory recoveries of 93.33%-102.33%. The excellent biomimetic sensing properties of the VP-PCM nanozyme are driving the development of a novel MPA analysis strategy assisted by ML in the context of production requirements of livestock safety.

In spite of organic-inorganic perovskite emerging as a novel efficient light-harvesting material owing to their superior optical properties, excitonic properties, and electrical conductivity, the related applications are severely limited for their poor stability and selectivity. Herein, we introduced hollow carbon spheres (HCSs) and 2-(perfluorohexyl) ethyl methacrylate (PFEM) based molecularly imprinted polymers (MIPs) to dual-functionalize CH3NH3PbI3. HCSs can provide perovskite load conditions, passivate perovskite defects, increase carrier transport and effectively improve its hydrophobicity. The perfluorinated organic compound based MIPs film can not only enhance the water and oxygen stability of perovskite, but also endow it specific selectivity. Moreover, it can reduce the photoexcited electron-hole pair recombination and prolong the electron lifetime. Benefiting from the synergistic sensitization of HCSs and MIPs, an ultrasensitive photoelectrochemical platform (MIPs@CH3NH3PbI3@HCSs/ITO) for cholesterol sensing was acquired with a very wide linear range of 5.0 × 10-14-5.0 × 10-8 mol/L and an extremely low detection limit of 2.39 × 10-15 mol/L. The designed PEC sensor exhibited good selectivity and stability, as well as practicality for real sample analysis. The present work extended the development of the high-performance perovskite and showed its broad application prospect for advanced PEC construction.

Most current redox-type nanozyme-based colorimetric sensing platforms are susceptible to interference from the reductant when using chromogenic probe, and the unstable H2O2 used in the peroxidase-like nanozyme-based systems is prone to difficulty in sensing signal reproducibility, while peroxidase-like nanozyme with oxidase-mimicking activity is easy to bring background interference by O2. Since the strong structural designability of covalent organic frameworks (COFs) endows them great application value in the sensing fields, therefore, we envision the construction a COF oxidase-like nanozyme-based controllable sensing system that integrates self-reporting, self-correcting and light-responsive functions to avoid these affects. Herein, 3-nitrotyrosine (3-NT) biomarker was selected as model analyte. 1,3,5-triformylphloroglucinol (Tp) and 3,6-diaminoacridine (DA) were acted as building monomers of the multifunctional COF nanozyme (termed as TpDA). Owing to the excellent light-responsive oxidase-mimicking property of TpDA, 3-NT can be efficiently oxidized, the inner filter effect (IFE) between TpDA and the 3-NT oxidation product greatly quenches the intrinsic fluorescence of TpDA, making it a controllable self-reporting system for fluorescence turn-off sensing 3-NT. Additionally, the excessive reactive oxygen species (ROS) that generated continuously during photocatalysis can resist the interference of endogenous reductants. This study not only provides new insights to avoid the interference of H2O2, background and reductants from conventional redox-type nanozyme-based colorimetric systems but also opens avenues to rational construct versatile COF nanozyme-based sensor.

DNA intercalation has increasingly been studied for various scenario implementations due to the diverse functions of DNA/intercalators. Nascent organic photoelectrochemical transistor (OPECT) biosensing taking place in organic electronics and photoelectrochemical bioanalysis represents a promising technological frontier in the arena. In this work, we first devise DNA intercalation-enabled OPECT for miRNA detection with a superior gain up to 17100. Intercalation of [Ru(bpy)2dppz]2+ within the miRNA-initiated hybrid chain reaction (HCR)-derived duplex DNA is realized for producing anodic photocurrent upon light stimulation, causing the corresponding target-dependent alternation in gate voltage (VG) and hence the modulated channel current (IDS) of poly (3,4-ethylenedioxythiophene) doped with poly (styrene sulfonate) (PEDOT:PSS) under specific drain voltage (VDS) for quantitative miRNA-21 analysis, which shows a wide linear relationship and a low detection limit of 5.5 × 10-15 mol L-1. This study features the DNA intercalation-enabled organic electronics with superior gain and is envisaged to attract more attention to explore DNA adducts for innovative bioelectronics and biosensing, given the diverse DNA binders with multiple functions.