Hybridization Chain Reaction Research Articles

Endogenous Glutathione-Activated Nucleic Acid Molecular Circuitry for Cell-Specific MicroRNA Imaging.

Sensitive and reliable microRNA imaging in living cells has significant implications for clinical diagnosis and monitoring. Catalytic DNA circuits have emerged as potent tools for tracking these intracellular biomarkers and probing the corresponding biochemical processes. However, their utility is hindered by the low resistance to external interference, leading to undesired off-site activation and consequent signal leakage. Therefore, achieving the endogenous control of the DNA circuit's activation is preferable to the reliable target analysis in living cells. In this study, we attempted to address this challenge by engineering a simple yet effective endogenous glutathione (GSH)-regulated hybridization chain reaction (HCR) circuit for acquiring high-contrast miRNA imaging. Initially, the HCR hairpin reactants were blocked by the engineered disulfide-integrated DNA duplex, thus effectively passivating their sensing function. And the precaged HCR hairpin was liberated by the cell-specific GSH molecule, thus initiating the HCR system for selectively amplified detection of microRNA-21 (miR-21). This approach prevented unwanted signal leakage before exposure into target cells, thus ensuring robust miR-21 imaging with high accuracy and reliability in specific tumor cells. Moreover, the endogenously responsive HCR circuit established a link between the small regulatory factors and miRNA, thereby enhancing the signal gain. In summary, the endogenously activatable DNA circuit represents a versatile toolbox for robust bioanalysis and exploration of potential signaling pathways in living cells.

Hook-Like DNAzyme-Activated Autocatalytic Biosensor for the Universal Detection of Pathogenic Bacteria.

DNAzyme-based assays have found extensive utility in pathogenic bacteria detection but often suffer from limited sensitivity and specificity. The integration of a signal amplification strategy could address this challenge, while the existing combination methods require extensive modification to accommodate various DNAzymes, limiting the wide-spectrum bacteria detection. We introduced a novel hook-like DNAzyme-activated autocatalytic nucleic acid circuit for universal pathogenic bacteria detection. The hook-like connector DNA was employed to seamlessly integrate the recognition element DNAzyme with the isothermal enzyme-free autocatalytic hybridization chain reaction and catalytic hairpin assembly for robust exponential signal amplification. This innovative autocatalytic circuit substantially amplifies the output signals from the DNAzyme recognition module, effectively overcoming DNAzyme's inherent sensitivity constraints in pathogen identification. The biosensor exhibits a strong linear response within a range of 1.5 × 103 to 3.7 × 107 CFU/mL, achieving a detection limit of 1.3 × 103 CFU/mL. Noted that the sensor's adaptability as a universal detection platform is established by simply modifying the hook-like connector module, enabling the detection of various pathogenic bacteria of considerable public health importance reported by the World Health Organization, including Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, and Salmonella typhimurium. Additionally, the specificity of DNAzyme in bacterial detection is markedly improved due to the signal amplification process of the autocatalytic circuit. This hook-like DNAzyme-activated autocatalytic platform presents a versatile, sensitive, and specific approach for pathogenic bacteria detection, promising to significantly expand the applications of DNAzyme in bacteria detection.

SERS biosensors based on catalytic hairpin self-assembly and hybridization chain reaction cascade signal amplification strategies for ultrasensitive microRNA-21 detection.

A highly sensitive surface-enhanced Raman scattering (SERS) biosensor has been developedfor the detection of microRNA-21 (miR-21) using an isothermal enzyme-free cascade amplification method involving catalytic hairpin assembly (CHA) and hybridization chain reaction (HCR). The CHA reaction is triggered by the target miR-21, which causes hairpin DNA (C1 and C2) to self-assemble into CHA products. After AgNPs@Capture captures the resulting CHA product, the HCR reaction is started, forming long-stranded DNA on the surface of AgNPs. A strong SERS signal is generated due to the presence of a large amount of the Raman reporter methylene blue (MB) in the vicinity of the SERS "hot spot" on the surface of AgNPs. The monitoring of the SERS signal changes of MB allows for the highly sensitive and specific detection of miR-21. In optimal conditions, the biosensor exhibits a satisfactory linear range and a low detection limit for miR-21 of 42.3 fM. Additionally, this SERS biosensor shows outstanding selectivity and reproducibility. The application of this methodology to clinical blood samples allows for the differentiation of cancer patients from healthy controls. As a result, the CHA-HCR amplification strategy used in this SERS biosensor could be a useful tool for miRNA detection and early cancer screening.

Selective Activation of Cascade Assembly Amplification for DNA Methyltransferase Detection Using a Double-Loop Interlocked DNA Circuit.

Precise and reliable monitoring of DNA adenine methyltransferase (Dam) activity is essential for disease diagnosis and biological analysis. However, existing techniques for detecting Dam activity often rely on specific DNA recognition probes that are susceptible to DNA degradation and exhibit limited target sensitivity and specificity. In this study, we designed and engineered a stable and dynamic DNA nanodevice called the double-loop interlocked DNA circuit (DOOR) that enables the sensitive and selective monitoring of Dam activity in complex biological environments. The DOOR incorporates two interlocked specialized sequences: a palindromic sequence for Dam identification and an initiator sequence for signal amplification. In the presence of Dam, the DOOR is cleaved by double-stranded DNA phosphodiesterase I endonuclease, generating massive double-stranded DNA (dsDNA) units. These units can self-assemble into a long dsDNA scaffold, thereby enhancing the subsequent reaction kinetics. The dsDNA scaffold further triggers a hyperbranched hybrid chain reaction to produce a fluorescent 3D DNA nanonet, enabling more precise monitoring of the Dam activity. The DOOR device exhibits excellent sensitivity, specificity, and stability, rendering it a powerful tool for studying DNA methylation in various biological processes and diseases.

A Visible-Light-Driven Self-Powered Nodularin-R Biosensing Platform Controlled by an Integrated Portable Photoelectrochemical Detection Device.

The presence of nodularin-R (NOD-R) in water has gained considerable attention because of its widespread distribution and high toxicity. In this study, an accurate and rapid visible-light-driven self-powered photoelectrochemical (PEC) biosensor was developed by integrating a portable paper-based electrode with a custom-built miniaturized PEC detection device. The newly designed system successfully achieved on-site detection of NOD-R in real water samples based on PEC technology. First, target recognition triggers the initiation of the hybridization chain reaction to generate double-stranded DNA. The thus-formed double-stranded DNA entrapped methylene blue (MB), and the dye molecules were irradiated with visible light for conversion to leuco-MB in the presence of ascorbic acid. The resulting leuco-MB species significantly amplified the PEC signal output of TiO2-MXene, enabling NOD-R detection. Under optimal conditions, the proposed PEC assay strategy demonstrated NOD-R detection within a concentration range from 20 fg mL-1 to 10 ng mL-1 with a detection limit of 19.6 fg mL-1. In addition, a custom-built miniaturized PEC detection device conveniently integrates the detection component with the light source, enabling the real-time collection of results via a wireless module. This innovative self-powered PEC platform provides significant advancements in smooth and intelligent detection compared to traditional PEC detection devices.

Orthogonal DNA Self-Assembly-Based Expansion Microscopy Platform for Amplified, Multiplexed Biomarker Imaging.

Expansion microscopy (ExM) facilitates nanoscale imaging under conventional microscopes, but it frequently encounters challenges such as fluorescence losses, low signal-to-noise ratio (SNR), and limited detection throughput. To address these issues, a method of orthogonal DNA self-assembly-based ExM (o-DAExM) platform is developed, which employs hybridization chain reaction instead of conventional fluorescence labeling units, showcasing signal amplification efficacy, enhancement of SNR, and expandable multiplexing capability at any stage of the ExM process. In this work, o-DAExM has been applied to compare with immunofluorescence-based ExM for cellular cytoskeleton imaging, and the resolved nanoscale spatial distributions of cytoskeleton show outstanding performance and reliability of o-DAExM. Furthermore, the study demonstrates the utility of o-DAExM in accurately revealing exosome heterogeneous information and multiplexed analysis of protein targets in single cells, which provides infinite possibilities in super-resolution imaging of cells and other samples. Therefore, o-DAExM offers a straightforward expansion and signal labeling method, highlighting future prospects to study nanoscale structures and functional networks in biological systems.

Conductivity-Regulated Bipolar Electrochemiluminescence Sensing Platform for Indicator-Free Homogeneous Bioassay.

Electrochemiluminescence (ECL) sensors have been widely developed because of their high sensitivity and low background. However, most of them suffered from tedious probe modification on the electrode and cross-interferences within the sensing and reporting reactions. The bipolar electrode based ECL (BPE-ECL) can effectively eliminate interference by physically separating the sensing and reporting cells, but there is still a need for exogenous electroactive indicators to transduce the variations between two poles of a BPE. Herein, based on the discovery that conductivity can be regulated in aqueous medium by homogeneous bioreaction, we showed a novel BPE-ECL sensing platform that combined the conductivity-based biosensing technology with ECL reporting system for the first time. Compared to many short nucleic acids, the target induced a hybridization chain reaction to produce the long nucleic acid aggregates, resulting in a conductivity decrease of the sensing cell and finally reducing the ECL response in the reporting cell. The BPE-ECL platform has already been applied to detect microRNA-21 for a demonstration. This innovative system not only separates the target sensing and reporting reactions but also avoids the use of electrochemical indicators for measurement. The BPE-ECL biosensing platform can be developed to detect different targets by changing the probe used.

Synergistic powering of DNA walker movement by endogenous dual enzymes for constructing dual-mode biosensors

To achieve highly sensitive and reliable detection of apurinic/apyrimidinic endonuclease 1 (APE1), a critical cancer diagnostic biomarker, we designed a DNA walker-based dual-mode biosensor, utilizing cellular endogenous dual enzymes (APE 1 and Flap endonuclease 1 (FEN 1)) to collaborate in activating and propelling DNA walker motion on DNA-functionalized Au nanoparticles. Incorporating both fluorescence and electrochemical detection modes, this system leverages signal amplification from DNA walker movement and cascade amplification through tandem hybridization chain reactions (HCR), achieving highly sensitive detection of APE 1. In the fluorescence mode, continuous DNA walker movement, initiated by APE1 and driven by FEN1, generates a robust signal response within a concentration range of 0.01–500 U mL−1, presenting a good linearity in the concentration range of 0.01–10 U mL−1, with a detection limit of 0.01 U mL−1. In the electrochemical detection module, the cascade upstream DNA walker and downstream HCR dual signal amplification strategy further enhances the sensitivity of APE1 detection, extending the linear range to 0.01–50 U mL−1 and reducing the detection limit to 0.002 U mL−1. Rigorous validation demonstrates the biosensor's specificity and anti-interference capability against multiple enzymes. Moreover, it effectively distinguishes cancer cells from normal cell lysates, exhibiting excellent stability and consistency in the dual-modes. Overall, our findings underscore the efficacy of the developed dual-mode biosensor for detecting APE1 in serum and cell lysates samples, indicating its potential for clinical applications in disease diagnosis.

Dual-modal biosensor for mercuric ion detection based on Cu2O@Cu2S/D-TA COF heterojunction with excellent catalase-like, electrochemical and photoelectrochemical properties

In this study, a dual-mode biosensor based on the heterojunction of Cu2O@Cu2S/D-TA COF was constructed for ultra-sensitive detection of Hg2+ using both photoelectrochemical and electrochemical approaches. Briefly, a 2D ultra-thin covalent organic framework film (D-TA COF film) with excellent photoelectrochemical signals was prepared on ITO surfaces through an in situ growth method. Subsequently, the probe H1 was immobilized onto the biosensor via Au–S bonds. In the presence of Hg2+, the formation of T-Hg2+-T complexes triggered hybridization chain reactions (HCR), leading to the attachment of abundant Cu2O@Cu2S probes onto the biosensor. As a p-type semiconductor, Cu2O@Cu2S could form a heterojunction with the underlying D-TA COF films. Meanwhile, it exhibited catalase-like activity, and the O2 produced by its catalytic decomposition of H2O2 can interact with the D-TA COF films, thus achieving double amplification of the photocurrent signal. Benefiting from the excellent and inherent Cu2+/Cu+ redox pairs of Cu2O@Cu2S, satisfactory differential pulse voltammetry (DPV) signals were obtained. As expected, the dual-mode biosensor was realized with wider linear ranges and low detection limits. Additionally, the analytical performance for Hg2+ in real water samples was excellent. Briefly, this suggested approach offers a facile and highly efficient modality for monitoring heavy metal ions in aquatic environments.

Expression of novel androgen receptors in three GnRH neuron subtypes in the cichlid brain.

In teleosts, GnRH1 neurons stand at the apex of the Hypothalamo-Pituitary-Gonadal (HPG) axis, which is responsible for the production of sex steroids by the gonads (notably, androgens). To exert their actions, androgens need to bind to their specific receptors, called androgen receptors (ARs). Due to a teleost-specific whole genome duplication, A. burtoni possess two AR paralogs (ARα and ARβ) that are encoded by two different genes, ar1 and ar2, respectively. In A. burtoni, males stratify along dominance hierarchies, in which an individuals' social status determines its physiology and behavior. GnRH1 neurons have been strongly linked with dominance and circulating androgen levels. Similarly, GnRH3 neurons are implicated in the display of male specific behaviors. Some studies have shown that these GnRH neurons are responsive to fluctuations in circulating androgens levels, suggesting a link between GnRH neurons and ARs. While female A. burtoni do not naturally form a social hierarchy, their reproductive state is positively correlated to androgen levels and GnRH1 neuron size. Although there are reports related to the expression of ar genes in GnRH neurons in cichlid species, the expression of each ar gene remains inconclusive due to technical limitations. Here, we used immunohistochemistry, in situ hybridization chain reaction (HCR), and spatial transcriptomics to investigate ar1 and ar2 expression specifically in GnRH neurons. We find that all GnRH1 neurons intensely express ar1 but only a few of them express ar2, suggesting the presence of genetically-distinct GnRH1 subtypes. Very few ar1 and ar2 transcripts were found in GnRH2 neurons. GnRH3 neurons were found to express both ar genes. The presence of distinct ar genes within GnRH neuron subtypes, most clearly observed for GnRH1 neurons, suggests differential control of these neurons by androgenic signaling. These findings provide valuable insight for future studies aimed at disentangling the androgenic control of GnRH neuron plasticity and reproductive plasticity across teleosts.

An ultra-sensitive electrochemical biosensor for circulating tumor DNA utilizing dual enzyme-assisted target recycle and hybridization chain reaction amplification strategies

Circulating tumor DNA (ctDNA) has emerged as a pivotal biomarker for cancers, offering a non-invasive means of detection through liquid biopsies. However, the current methodologies for ctDNA detection often struggle with sensitivity, especially when confronted with ultra-low concentrations. To address this challenge, we have engineered a remarkably sensitive electrochemical biosensor that can detect low concentration of ctDNA. This breakthrough is achieved by integrating a dual enzyme-assisted amplification mechanism with hybridization chain reaction (HCR). In the initial phase, the presence of ctDNA triggers the unfolding of the H2 hairpin structure, resulting in a segment of double-stranded DNA. The subsequent introduction of Klenow (3′→5′ exo-) and nicking endonuclease (Nb.BbvCI) enzymes leads to the abundant production of extended DNA (EXTDNA). This step critically enhances the initial amplification of the target DNA, ensuring superior specificity and selectivity thanks to the synergistic enzymatic activity. EXTDNA efficiently unfolds the H1 hairpin structure immobilized on the gold electrode, which acts as the initiation point for the HCR. Further amplification of the trace target DNA is achieved through an additional H3 and H4 hybrid-mediated HCR. Finally, the differential pulse voltammetry signal of methylene blue is increased significantly. Our results reveal that the developed electrochemical biosensor displays an exceptional linear correlation with ctDNA across concentrations ranging from 10 fM to 20 pM, with an unprecedented detection limit of 2.3 fM (3 s/k). This innovative approach shows immense potential for the ultrasensitive detection of ctDNA, promising broad applications in early cancer detection and monitoring.

A portability self-powered sensor facilitates sensitive Cd2+ detection: Dual mechanism and three quantitative mode

As a common heavy metal contaminant, Cd2+ has adverse effects on food safety and consumer health. It is very important for human health to realize highly sensitive Cd2+ detection methods. The self-powered sensing system based on enzyme biofuel cells (EBFCs) does not need an external power supply, which can simplify the experimental equipment and has great application value in portable detection. Thus, the biosensor is innovatively integrated into the screen-printed electrode to construct a new type of portable sensor suitable for on-site and real-time Cd2+ detection. Hybridization chain reaction (HCR) combined with the Cd2+-dependent deoxyribose (DNAzyme) signal amplification strategy is used to enhance the detection sensitivity while specifically recognizing the Cd2+. Moreover, the self-powered sensor combines with smartphones to realize quantitative Cd2+ detection without other instruments and has the characteristic of Effectively improving the hazard detection technology is essential to ensure food safety. Portability, simplicity, and speed are suitable for real-time Cd2+ detection in the field. The dual mechanism and three quantitative modes combining colorimetric and two electrical signals output modes are adopted to realize the visualization and accurate detection. A series of research results confirm that this strategy is of great significance to strengthen the development of intelligent Cd2+ technology, expand the application of self-powered sensing technology, and improve the safety detection system.

The sensing of circRNA with tetrahedral DNA nanostructure modified microfluidic chip

BackgroundCircular ribonucleic acids (circRNAs) are a type of covalently closed noncoding RNA with disease-relevant expressions, making them promising biomarkers for diagnosis and prognosis. Accurate quantification of circRNA in biological samples is a necessity for their clinical application. So far, methods developed for detecting circRNAs include northern blotting, reverse transcription quantitative polymerase chain reaction (RT-qPCR), microarray analysis, and RNA sequencing. These methods generally suffer from disadvantages such as large sample consumption, cumbersome process, low selectivity, leading to inaccurate quantification of circRNA. It was thought that the above drawbacks could be eliminated by the construction of a microfluidic sensor. ResultsHerein, for the first time, a microfluidic sensor was constructed for circRNA analysis by using tetrahedral DNA nanostructure (TDN) as the skeleton for recognition probes and target-initiated hybridization chain reaction (HCR) as the signal amplification strategy. In the presence of circRNA, the recognition probe targets the circRNA-specific backsplice junction (BSJ). The captured circRNA then triggers the HCR by reacting with two hairpin species whose ends were labeled with 6-FAM, producing long DNA strands with abundant fluorescent labels. By using circ_0061276 as a model circRNA, this method has proven to be able to detect circRNA of attomolar concentration. It also eliminated the interference of linear RNA counterpart, showing high selectivity towards circRNA. The detection process can be implemented isothermally and does not require expensive complicated instruments. Moreover, this biosensor exhibited good performance in analyzing circRNA targets in total RNA extracted from cancer cells. SignificanceThis represents the first microfluidic system for detection of circRNA. The biosensor showed merits such as ease of use, low-cost, small sample consumption, high sensitivity and specificity, and good reliability in complex biological matrix, providing a facile tool for circRNA analysis and related disease diagnosis in point-of care application scenes.

Multiplexed in situ protein imaging using DNA-barcoded antibodies with extended hybridization chain reactions.

Antibodies have long served as vital tools in biological and clinical laboratories for the specific detection of proteins. Conventional methods employ fluorophore or horseradish peroxidase-conjugated antibodies to detect signals. More recently, DNA-conjugated antibodies have emerged as a promising technology, capitalizing on the programmability and amplification capabilities of DNA to enable highly multiplexed and ultrasensitive protein detection. However, the nonspecific binding of DNA-conjugated antibodies has impeded the widespread adoption of this approach. Here, we present a novel DNA-conjugated antibody staining protocol that addresses these challenges and demonstrates superior performance in suppressing nonspecific signals compared to previously published protocols. We further extend the utility of DNA-conjugated antibodies for signal-amplified in situ protein imaging through the hybridization chain reaction (HCR) and design a novel HCR DNA pair to expand the HCR hairpin pool from the previously published 5 pairs to 13, allowing for flexible hairpin selection and higher multiplexing. Finally, we demonstrate highly multiplexed in situ protein imaging using these techniques in both cultured cells and tissue sections.

Label-free and ultrasensitive detection of environmental lead ions based on spatially localized DNA nanomachines driven by hyperbranched hybridization chain reaction

A label-free fluorescent sensing strategy for the rapid and highly sensitive detection of Pb2+ was developed by integrating Pb2+ DNAzyme-specific cleavage activity and a tetrahedral DNA nanostructure (TDN)-enhanced hyperbranched hybridization chain reaction (hHCR). This strategy provides accelerated reaction rates because of the highly effective collision probability and enriched local concentrations from the spatial confinement of the TDN, thus showing a higher detection sensitivity and a more rapid detection process. Moreover, a hairpin probe based on a G-triplex instead of a G-quadruplex or chemical modification makes hybridization chain reaction more controlled and flexible, greatly improving signal amplification capacities and eliminating labeled DNA probes. The enhanced reaction rates and improved signal amplification efficiency endowed the biosensors with high sensitivity and a rapid response. The label-free detection of Pb2+ based on G-triplex combined with thioflavin T can be achieved with a detection limit as low as 1.8 pM in 25 min. The proposed Pb2+-sensing platform was also demonstrated to be applicable for Pb2+ detection in tap water, river water, shrimp, rice, and soil samples, thus showing great potential for food safety and environmental monitoring.

HCR-Assisted RTF-EXPAR-Based Lateral Flow Analysis for Sensitive Detection of H1N1 Influenza Virus.

The H1N1 influenza virus is a significant pathogen responsible for seasonal influenza, and its frequent outbreaks pose substantial challenges to global public health. The present study successfully developed a lateral flow analysis platform that integrates reverse transcription-free exponential amplification reaction (RTF-EXPAR) and hybridization chain reaction (HCR) processes with functionalized quantum dots for the direct detection of H1N1 influenza virus RNA, eliminating the need for reverse transcription. The fluorescence signal on the band recorded with a smartphone can be utilized for the quantitative determination of the target. Interestingly, the dual signal amplification strategy exhibits high sensitivity with a remarkably low detection limit of 10 aM. Moreover, this platform exhibits excellent flexibility and universality, where the various pathogens can be determined by replacing the specific nucleic acid fragments in RTF-EXPAR. The aforementioned advantages reveal its huge potential in the early diagnosis of H1N1 influenza virus infection and developing point-of-care testing (POCT) equipment for nucleic acid analysis.

Catalytic hairpin-assembled and DNAzyme-driven DNA walker integrated hybridization chain reaction to construct a label-free electrochemical aptasensor for the detection of acetamiprid

As a new type of nicotine pesticide, the excessive residue of acetamiprid (ACE) in food and the environment seriously poses human health. Hence, this article developed an electrochemical aptasensor for detecting ACE based on a cascaded amplification circuit composed of catalytic hairpin-assembled and DNAzyme-driven DNA walker and hybridization chain reaction. Through the target triggered catalytic hairpin assembly (CHA), the free travelling strand of DNA walker and intact catalytic core of DNAzyme were formed, and with the assistance of Mg2+, DNAzyme was activated, driving the travelling strand to walk autonomously, repeatedly cutting hairpin DNA substrates on the electrode surface, and obtaining abundant DNA fragments. These fragments contained the initiators of hybridization chain reaction (HCR) could trigger HCR and produce the long double-strand DNA (dsDNA) for loading methylene blue (MB). Owing to the fact that Ce(Ⅲ, Ⅳ)-MOF material with electrocatalytic activity, the electrochemical reduction of MB could be effectively catalyzed, obtain significantly enhanced current signals, and ultimately achieve label-free and sensitive detection of ACE, with a detection limit of 16.38 fM (S/N = 3). Moreover, the aptasensor demonstrated excellent selectivity, stability, and reproducibility, and exhibited good reliability for detecting ACE in the vegetable soil and tomato samples, providing a new idea for the detection of pesticide residues in the fields of food and environment.

Aptasensors and Advancement in Molecular Recognition Technology

AbstractSynthetic oligonucleic acids known as aptamers exhibit remarkable selectivity and affinity for target recognition and binding. Selected via an iterative process known as “selective evolution of ligands by exponential enrichment” (SELEX), aptamers fold into defined 3D conformations to interact with their targets. The incorporation of aptamers as recognition elements has driven notable progress in biosensors, giving rise to the development of aptasensors. Here, the process of aptamer discovery and the development of various types of aptasensors are summarized. The fundamental design principles of aptasensors are elaborated along with the superiority of aptamers compared to antibodies. The various modes employed by aptasensors, such as structure‐switching design, hybridization chain reaction amplification, enzyme‐assisted recycling, and split aptamer design are examined. Further light is shed on the diverse landscape of aptasensors, their adaptability to different analytes aptasensors as well as their potential to propel advancements in modern biosensor technology. As a nucleic acids‐based biosensor platform, aptasensors poise to become a next generation of sensitive and cost‐effective technology to shape the future of molecular recognition in biosensing.

Topographic Organization of Glutamatergic and GABAergic Parvalbumin-Positive Neurons in the Lateral Habenula.

Parvalbumin-expressing (PV) neurons, classified by their expression of the calcium-binding protein parvalbumin, play crucial roles in the function and plasticity of the lateral habenular nucleus (LHb). This study aimed to deepen our understanding of the LHb by collecting information about the heterogeneity of LHb PV neurons in mice. To achieve this, we investigated the proportions of the transmitter machinery in LHb PV neurons, including GABAergic, glutamatergic, serotonergic, cholinergic, and dopaminergic neurotransmitter markers, using transcriptome analysis, mRNA in situ hybridization chain reaction, and immunohistochemistry. LHb PV neurons comprise three subsets: glutamatergic, GABAergic, and double-positive for glutamatergic and GABAergic machinery. By comparing the percentages of the subsets, we found that the LHb was topographically organized anteroposteriorly; the GABAergic and glutamatergic PV neurons were preferentially distributed in the anterior and posterior LHb, respectively, uncovering the anteroposterior topography of the LHb. In addition, we confirmed the mediolateral topography of lateral GABAergic PV neurons. These findings suggest that PV neurons play distinct roles in different parts of the LHb along the anteroposterior and mediolateral axes, facilitating the topographic function of the LHb. It would be interesting to determine whether their topography is differentially involved in various cognitive and motivational processes associated with the LHb, particularly the involvement of posterior glutamatergic PV neurons.

Highly conductive AuNPs/Co-MOF nanocomposites synergistic hybridization chain reaction enzyme-free electrochemical aptasensor for ultrasensitive detection of Aflatoxin B1

Aflatoxin B1 (AFB1) is the most poisonous and carcinogenic pollutant among numerous mycotoxins, posing a serious risk to the health of humans. Herein, an electrochemical aptasensor was constructed for AFB1 detection using gold nanoparticles/cobalt-based metal–organic skeleton (AuNPs/Co-MOF) as a modifying material combined with an enzyme-free signal amplification strategy. AuNPs/Co-MOF have an extensive specific surface area and outstanding electrical conductivity, which allows for the loading of a large number of DNA strands and enhancement of the thionine (THi) redox signal, leading to a significant increase in signal response THi loaded bimetallic core–shell nanocomposites (Au@PtNPs) were synthesized as signal labels. Au@PtNPs have high catalytic activity towards THi while improving the efficiency of the loaded THi, which enables the electrochemical signal of THi to be amplified. In addition, the hybridization chain reaction (HCR) signal amplification strategies for enzyme-free amplification elicits a strong response in the electrochemical signal while ensuring sensor stability. The aptasensor has a detection range of 0.001–500 ng/mL and achieves the limit of detection (LOD) was 1.2 × 10-2 pg/mL and limit of quantification (LOQ) was 4.0 × 10-2 pg/mL under optimal experimental conditions. Therefore, applications for electrochemical aptasensors in the realm of food safety monitoring are quite promising.