Assembly Reaction Research Articles

A ratiometric fluorescent biosensor based on catalytic hairpin assembly reaction for ultrasensitive detection of aflatoxin-producing genes

Aflatoxins as the world’s most toxic contamination is metabolized by Aspergillus flavus and its production is closely related to aflatoxin-producing genes. It is significant to monitor the aflatoxin-producing genes as early as possible to prevent the contamination and spread of aflatoxins. Therefore, a label-free and free separation fluorescent biosensor using the selective cation exchange reaction in conjunction with the signal amplification of catalytic hairpin assembly (CHA) was developed for the detection of aflatoxin-producing gene utilizing CdTe quantum dots (CdTe QDs) and Nitrogen-doped carbon quantum dots (NCQDs) as the beacon molecule. There were two types of hairpin structures in this sensing system, hairpin probe 1 (HP1) was generated by Ag+ and the recognition probe with the complex C-Ag+-C structure and hairpin probe 2 (HP2) was the DNA fragments showing a hairpin arrangement. Target nor-1 gene may activate the CHA, which created a significant quantity of HP1-HP2 double-stranded DNAand recycled the target nor-1 gene. The released amounts of Ag+ suppressed the fluorescence signal of CdTe QDs, while it wasn’t observed the discernible effect on NCQDs. This fluorescent biosensor displayed the excellent sensitivity for aflatoxin-producing nor-1 gene in a dynamic range of 5 pM to 50 nM with a detection limit of 1.66 pM. This fabricated sensor also showed the visible distinguish for different concentrations of nor-1 gene. It exhibited the potential to offer vital enhancements for food quality control through a simple, fast and sensitive monitoring test system.

A Multifunctional Peptide Nucleic Acid/Peptide Copolymer-Based Dual-Mode Biosensor with Macrophage-Hitchhiking for Enhanced Tumor Imaging and Urinalysis.

Biosensors are capable of diagnosing tumors through imaging in vivoor liquid biopsy, but they face the challenges of inefficient delivery into tumor sites and the lack of reliable tumor-associated biomarkers. Herein, we constructed a dual-mode biosensor based on a multifunctional peptide nucleic acid (PNA)/peptide copolymer and DNA tetrahedron for tumor imaging and urinalysis. The biosensor could enter the cancer cells to initiate a microRNA-21-specific catalytic hairpin assembly reaction after cleavage by matrix-metalloprotease (MMP) in the tumor microenvironment, and the MMP cleavage product was released into the bloodstream and then was filtered out by the kidney. As PNA was a synthetic DNA analogue that could not be degraded by nucleases and proteases, it could serve as a reliable synthetic biomarker and be easily detected by high-performance liquid chromatography in urine. Importantly, the biosensor was hitchhiked on the macrophage membrane to realize efficient delivery in the depth of tumor utilizing the macrophage ability of actively homing to the tumor site and infiltrating into the tumor. The results indicated that the signal output of the biosensor was improved remarkably and mice with a tumor volume as little as 30-40 mm3 could be reliably discriminated through urine assay. This innovative macrophage-hitchhiking dual-mode biosensor holds a great potential as a non-invasive and convenient tool for tumor diagnosis and tumor progression evaluation.

Mixed Carboxylate Ligands Bridging Tetra-Pr3+-Encapsulated Antimonotungstate: Syntheses, Structure, and Catalytic Activity for Imidazoles Synthesis.

Multinuclear Pr-containing antimonotungstate [Pr4(H2O)10W6O13(mal)2(OAc)(B-α-SbW9O33)4]21- (Pr-1, mal = malate anion, OAc = acetate anion), bridged by organic carboxylic acid, was synthesized through a one-pot assembly reaction and structurally characterized. Pr-1 is composed of four [B-α-SbW9O33]9- fragments fused together by an organic-inorganic hybrid central [Pr4(H2O)10W6O13(mal)2(OAc)]15+ cluster core through 24 μ2-O atoms. Notably, the central cluster comprises unprecedented decanuclear Pr4(H2O)10W6O13 jointly decorated by two types of carboxylic acid ligands. This integration of rare earth-containing antimonotungstate with mixed organic carboxylate ligands is very rare in POMs chemistry. Pr-1 exhibits excellent catalytic activity in the cyclo-condensation reaction involving benzil, aldehyde, and NH4OAc. A series of 2,4,5-trisubstituted imidazoles were synthesized in remarkable yields using iPrOH as a green solvent.

Live-Cell Imaging of miRNAs with the Combination of CHA and HCR Techniques.

The abnormal expression of miRNA-21 is closely related to many cancers, and its sensitive detection plays an important role in the diagnosis and treatment of cancers. In the report, we designed a non-enzymatic amplification sensing system based on the catalytic hairpin assembly (CHA) and palindrome-based hybridization chain reaction (PHCR) technique for the detection and imaging of miRNA in living cells. Based on PHCR technology, two ingeniously designed palindrome-contained fluorescently labeled hairpin probes are sequentially opened upon the stimulation of short oligonucleotide triggers, promoting the autonomous assembly of cross-linked network-like structures (CNSs) and generating fluorescence resonance energy transfer (FRET) signal. Moreover, the PHCR technique can be combined with CHA technology to sufficiently improve amplification efficiency and signal output. By utilizing the CHA-PHCR sensing system, one miRNA-21 activates many triggers through CHA process and in turn initiates the assembly of CNSs by PHCR reaction, efficiently amplifying the FRET signal output. As such, the sensing system can detect target miRNAs down to 10pm with high detection specificity. Moreover, the CNS exhibits the enhanced intracellular stability, enabling the living cell imaging of miRNA-21. The CHA-PHCR sensing system can also screen the expression level of miRNA-21 in different living cells and differentiate cancer cells from healthy cells, which is consistent with the gold-standard PCR method.

Sensitive detection of K-ras gene by a dual-mode “on-off-on” sensor based on bipyridine ruthenium-MOF and bis-enzymatic cleavage technology

This study developed a dual-mode “on-off-on” sensor based on a bipyridine ruthenium metal–organic framework (Ru-MOF) and dual enzyme cleavage technology for the sensitive detection of the K-ras gene. The sensor combines electrogenerated chemiluminescence (ECL) and fluorescence (FL) detection modes, achieving high sensitivity and specificity in detecting the K-ras gene through catalytic hairpin assembly (CHA) and dual enzyme cleavage reactions. Experimental results showed that the detection limits for the K-ras gene were 0.044 fM (ECL) and 0.16 fM (FL), demonstrating excellent selectivity and stability during detection. Through testing actual samples, the sensor has shown potential for application in complex biological environments. This method offers an efficient and reliable new tool for cancer diagnosis and treatment.

Membrane-dependent reactions of blood coagulation: classical view and state-of-the-art concepts

The complex mechanism called hemostasis evolved in living organisms to prevent blood loss when a blood vessel is damaged. In this process, two closely interconnected systems are distinguished: platelet-vascular and plasmatic hemostasis. Plasmatic hemostasis is a system of proteolytic reactions, in which blood plasma proteins called coagulation factors are involved. A key feature of this system is the localization of enzymatic reactions on the surface of phospholipid membranes, which increases their rate by up to 5 orders of magnitude. This review describes the basic mechanisms of coagulation factors binding to phospholipid membranes, pathways for complex assembly and activation reactions, and discusses the role of membranes in this process, their composition and sources. The binding of coagulation factors to procoagulant membranes leads not only to the acceleration of coagulation reactions, but also to their selective localization in restricted areas and protection from being washed away by the flow. The efficiency of coagulation reactions is regulated by the composition of the outer layer of the membrane, primarily through a special mechanism of mitochondria-dependent necrotic platelet death.

A simple and enzyme-free method for sensitive p53 analysis based on DNAzyme-mediated signal amplification

There is an urgent demand for a simple yet extremely accurate biosensor to analyze tumorigenesis. Herein, we present a novel fluorescent and enzyme-free approach for detecting p53 gene cascading proximity ligation-mediated catalytic hairpin assembly and DNAzyme-assisted signal reaction. When the target p53 gene is present, the interaction between p53 and L1 and L2 chains initiates catalytic hairpin assembly and subsequently exposes DNAzyme in the P3 probe. The exposed DNAzyme binds with the loop region of the P4 probe and generates a nicking site, resulting in the release of a significant amount of ATMND that is conjugated in the stem section of P4. This leads to an amplified fluorescence response, which serves as a fluorescence signal for the detection of the p53 gene. This method allows for the accurate and sensitive identification of the p53 gene, exhibiting a linear reaction range of 1 fM to 1 nM, with a limit of detection as low as 0.23 fM. Furthermore, this fluorescent method has been utilized for the examination of clinical samples with a favorable recovery rate. Crucially, this versatile platform may be expanded to analyze different targets by changing the corresponding recognition unit, showing great potential for point-of-care testing in tumorigenesis analysis.

Modeling compound lipid homeostasis using stable isotope tracing.

Lipids represent the most diverse pool of metabolites found in cells, facilitating compartmentation, signaling, and other functions. Dysregulation of lipid metabolism is linked to disease states such as cancer and neurodegeneration. However, limited tools are available for quantifying metabolic fluxes across the lipidome. To directly measure reaction fluxes encompassing compound lipid homeostasis, we applied stable isotope tracing, high-resolution mass spectrometry, and network-based isotopologue modeling to non-small cell lung cancer (NSCLC) models. Compound lipid metabolic flux analysis (CL-MFA) enables the concurrent quantitation of fatty acid synthesis, elongation, headgroup assembly, and salvage reactions within virtually any biological system. Here, we resolve liver kinase B1 (LKB1)-mediated regulation of sphingolipid recycling in NSCLC cells and precision-cut lung slice cultures. We also demonstrate that widely used tissue culture conditions drive cells to upregulate fatty acid synthase flux to supraphysiological levels. Finally, we identify previously uncharacterized isozyme specificity of ceramide synthase inhibitors, highlighting the molecular detail revealed by CL-MFA.

Highly sensitive β-amyloid1–42 oligomer aptamer-based assay via dual cyclic amplifications of three-way catalytic hairpin assembly and palindrome polymerase reaction

β-amyloid1–42 oligomer (AβO1–42), a neurotoxic protein, is considered the key factor in the progression of Alzheimer's disease (AD). And, high sensitivity detection of AβO1–42 is of considerable importance for the early diagnosis and deterioration prevention of AD. We present here an ultrasensitive fluorescent AβO1–42 detection approach based on aptamer recognition probes and a new dual cyclic signal amplification strategy by combining three-way catalytic hairpin assembly (3W-CHA) with palindromic sequence-based polymerase amplification (PBPA). Once the aptamers bind to target AβO1–42 molecules, ssDNA sequences are released to initiate 3W-CHA between three hairpins to unfold the signal hairpins and to form Y-shaped DNA structures (Y-DNAs). Subsequently, the assistance ssDNAs bind the Y-DNAs to expose the palindromic sequences to yield Y-DNA dimers, which lead to cyclic PBPA reaction with presence of polymerase to further unfold more signal hairpins for yielding considerably magnified fluorescence recovery, enabling sensitive assay of AβO1–42 down to 2.4 pM within a dynamic range of 5 pM∼50 nM. Furthermore, this strategy has been validated for interrogation of low AβO1–42 levels in artificial cerebrospinal fluid (CSF), suggesting its promising potential for application in early diagnosis and prevention of AD.

Highly sensitive detection of DNA methyltransferase1 triggered by methylation protection

Type 2 diabetes is on the rise year by year, and 80 % of patients with type 2 diabetes die from cardiovascular complications. Strict control of blood glucose can significantly reduce the occurrence and development of diabetic microvascular complications, but can not effectively prevent the occurrence of diabetic macrovascular lesions. DNA methyltransferase 1 (DNMT1) plays an important role in developing phenotypic switching and secretion of inflammatory factors in vascular smooth muscle cells(VSMC), leading to the deregulation of vascular function. In this work, a biosensor based on methylation protection was designed. In the presence of the target DNMT 1, the formed methylation sites protect the dsDNA from being digested by the HpaII restriction enzyme. At the same time, a large number of G-quadruplex are produced by the catalyzed hairpin assembly (CHA) reaction cycle, and the peroxidase-like activity of G-tetraplex and hemin is used to continuously catalyze the color change of substrate ABTS, thus realizing dual signal amplification. The binding of magnetic beads to dsDNA effectively reduces the background signal and enables a highly selective and sensitive analysis of the target DNMT 1 in complex biological samples, with a detection limit as low as 0.22 UmL−1, showing a good linear relationship in the range of 0.22–10 UmL−1. A new approach for the study of macrovascular damage is provided in type 2 diabetes.

CRISPR/Cas13a-mediated triple signal amplification strategy for viral RNA detection

CRISPR/Cas13a system has been combined with different isothermal amplification strategies for RNA analysis. Nevertheless, expensive enzymes are usually indispensable in these isothermal amplification methods, which increases the detection cost. In this study, a CRISPR/Cas13a-mediated triple signal amplification strategy has been designed for selective, sensitive and inexpensive detection of viral RNA. SARS-CoV-2 S RNA activated CRISPR/Cas13a to cut hairpin DNA, triggering catalytic hairpin assembly (CHA) and subsequent hybridization chain reaction (HCR) to form a long-stranded Y-type DNA. G-quadruplexes were distributed on the branch of the long-stranded DNA and bound with N-methyl mesoporphyrin IX (NMM) to enhance the fluorescence signal. SARS-CoV-2 S RNA was specifically detected with a limit of detection (LOD) of 285 fM. Notably, no additional expensive enzymes were required in the entire detection process. The CRISPR/Cas13a-mediated triple signal amplification strategy holds great potential for specific and sensitive viral RNA detection with low cost.

A narrow ratio of nucleic acid to SARS-CoV-2 N-protein enables phase separation

SARS-CoV-2 Nucleocapsid protein (N) is a viral structural protein that packages the 30kb genomic RNA inside virions and forms condensates within infected cells through liquid-liquid phase separation (LLPS). In both soluble and condensed forms, N has accessory roles in the viral life cycle including genome replication and immunosuppression. The ability to perform these tasks depends on phase separation and its reversibility. The conditions that stabilize and destabilize N condensates and the role of N-N interactions are poorly understood. We have investigated LLPS formation and dissolution in a minimalist system comprised of N protein and an ssDNA oligomer just long enough to support assembly. The short oligo allows us to focus on the role of N-N interaction. We have developed a sensitive FRET assay to interrogate LLPS assembly reactions from the perspective of the oligonucleotide. We find that N alone can form oligomers but that oligonucleotide enables their assembly into a three-dimensional phase. At a ∼1:1 ratio of N to oligonucleotide, LLPS formation is maximal. We find that a modest excess of N or of nucleic acid causes the LLPS to break down catastrophically. Under the conditions examined here, assembly has a critical concentration of about 1 μM. The responsiveness of N condensates to their environment may have biological consequences. A better understanding of how nucleic acid modulates N-N association will shed light on condensate activity and could inform antiviral strategies targeting LLPS.

Portable Monitoring System for Assessing Therapeutic Efficacy in HER2‐Overexpressing Breast Cancer: One‐Step Simultaneous Detection of Cancer‐Derived Exosomal Proteins and mRNA in Urine

AbstractPoint‐of‐care (POC) monitoring of patient condition is crucial for effective cancer treatment and prognosis. This can be achieved non‐invasively by analyzing exosomes in body fluids. However, the heterogeneity of exosomes and non‐standardized quantification methods may interfere with clearly determining the patient's condition. Therefore, there is a need for technology that can precisely analyze both tumor‐derived exosomes and normal exosomes. Herein, this study presents the exosome multiple‐separation for simultaneous detection (EXO‐MUSSID) platform, which simultaneously isolates different exosomes based on their magnetization properties and monitors therapeutic efficacy of drugs. Using immunoaffinity magnetophoresis technology, HER2‐overexpressing and normal exosomes are collected separately, enabling real‐time monitoring of HER2 (also known as ERBB2) expression by analyzing the mRNA of each exosome based on a catalytic hairpin assembly (CHA) reaction. A portable fluorescence reader customized for the EXO‐MUSSID platform is developed for POC monitoring of HER2‐overexpressing breast cancer (HER2+ BC). The performance of the EXO‐MUSSID platform is validated using urine samples from HER2+ BC mouse models, confirming the progression of HER2+ BC and the changes in HER2 expression due to trastuzumab treatment. It is expected to serve as a valuable tool for exosome‐based liquid biopsy in disease monitoring.

Intelligent Analysis of Avian Influenza Virus Biomarkers Based on Chemiluminescence Resonance Energy Transfer and DNA Logic Analysis.

A one-step logical analysis of multiple targets remains challenging. Herein, we report a one-pot and intelligent DNA logical analysis platform for the diagnosis of avian influenza virus (AIV) biomarkers based on chemiluminescence resonance energy transfer (CRET) and logic operations. On the surface of Lum/PEI/CaCO3 microparticles, the excited state of luminol underwent CRET with fluorescein isothiocyanate or rhodamine B isothiocyanate, producing three well-separated light emissions at 425, 530, and 590 nm, respectively. Taking advantage of the close distance between fluorophores aligned by the catalytic hairpin assembly reaction, the CRET efficiency was greatly enhanced (53.1%). H1N1, H7N9, and H5N1 were detected with limits of detection values as low as 15, 34, and 58 pM, respectively. Three-input logic circuits were simultaneously conducted on the surface of Lum/PEI/CaCO3 microparticles, enabling the rapid and accurate discrimination of multiple AIV biomarkers in one solution. In terms of peak positions and the normalized value of the total peak intensity, three biomarkers can be simultaneously discriminated without any other complex operations. In summary, the CRET-based multiple analytical assay was developed as an intelligent biosensor for identifying AIV biomarkers, having promising application prospects in the field of multiple analysis and precise disease diagnosis.

Synthesis pathways of (HfZrTiCe/La/Y)O2-x nanoparticles via benzyl alcohol route at critical temperature

The controllable synthesis of high-entropy fluorite oxide (HEO) having large ionic radius mismatch remains a challenging due to poor understanding on nucleation. The (HfZrTiLn)-5HEO nanoparticles with 15 % ionic radius mismatch were synthesized via benzyl alcohol route at 220 °C-5 min in presence of PtCl4 and Fe(acac)3, exhibiting novel optical, electrical and magnetic properties. Nucleation pathways of the 5HEO at the critical temperature were elucidated by using a comparison study of conventional heating and microwave irradiation heating. Consistency of XRD patterns and STEM-EDX observation indicate that the resultant Hf-OBn monomers acted as the nucleation center of the 5HEO, determined by diffusion kinetics. The nucleation rate depended on the metal monomers assembly and esterification reaction, which was accelerated by water vapor pressure produced in-situ by 0.5×10−4mol/l PtCl4 catalyst. The Fe-metal organic cages derived from 1.5×10−4mol/l Fe(acac)3 additive served as the structure stabilizer of Zr/Ti monomers, and prevented early hydrothermal reaction route.

Versatile Bovine Serum Albumin as Ingenious Electron Operator-Enhanced Photoelectrochemical Biosensing for Ultrasensitive Detection of miRNA.

Bovine serum albumin (BSA) has been widely used in biosensors as a blocking agent. Herein, conformist BSA was first exploited as an ingenious operator to enhance the photocurrent response of (2Z,2'Z)-2,2'-(1,4-phenylene)bis(3-(4-(bis(4-methoxyphenyl)amino)phenyl)acrylonitrile) (TPDCN)-based photoelectrochemical (PEC) platform via manipulating the electron transfer process of the detection system. Concretely, the presence of target molecules triggered catalytic hairpin assembly reaction and subsequently powered terminal deoxynucleotidyl transferase-mediated signal amplification to produce the AgNP@BSA-DNA dendrimer nanostructure. After being treated with HNO3, a large amount of BSA could be released from the dendrimer nanostructure. When they were transferred to the TPDCN-based PEC platform, the photocurrent response of the biosensor was largely enhanced because BSA can manipulate the electrons of TPDCN via a well-matched energy level to form a new electron transfer track. Meanwhile, tryptophan (Trp) in BSA could be oxidized to quinone Trp-O under photoirradiation, which can facilitate the oxidation of ascorbate and generate more H+ to promote the migration of photogenerated electrons. As a result, the proposed PEC biosensor exhibits excellent analytical performance for detection of miRNA-21 (as a model target) over a wide linear range of 0.01 to 10,000 pM with detection limit as low as 4.7 fM. Overall, this strategy provides a new perspective on constructing efficient PEC biosensors, which expands the potential applications in bioanalysis and clinical diagnosis.

Multiplex Expression Cassette Assembly: A flexible and versatile method for building complex genetic circuits in conventional vectors.

The manipulation of multiple transcription units for simultaneous and coordinated expression is not only key to building complex genetic circuits to accomplish diverse functions in synthetic biology, but is also important in crop breeding for significantly improved productivity and overall performance. However, building constructs with multiple independent transcription units for fine-tuned and coordinated regulation is complicated and time-consuming. Here, we introduce the Multiplex Expression Cassette Assembly (MECA) method, which modifies canonical vectors compatible with Golden Gate Assembly, and then uses them to produce multi-cassette constructs. By embedding the junction syntax in primers that are used to amplify functional elements, MECA is able to make complex constructs using only one intermediate vector and one destination vector via two rounds of one-pot Golden Gate assembly reactions, without the need for dedicated vectors and a coherent library of standardized modules. As a proof-of-concept, we modified eukaryotic and prokaryotic expression vectors to generate constructs for transient expression of green fluorescent protein and β-glucuronidase in Nicotiana benthamiana, genome editing to block monoterpene metabolism in tomato glandular trichomes, production of betanin in tobacco and synthesis of β-carotene in Escherichia coli. Additionally, we engineered the stable production of thymol and carvacrol, bioactive compounds from Lamiaceae family plants, in glandular trichomes of tobacco. These results demonstrate that MECA is a flexible, efficient and versatile method for building complex genetic circuits, which will not only play a critical role in plant synthetic biology, but also facilitate improving agronomic traits and pyramiding traits for the development of next-generation elite crops.

G-quadruplex embedded in semi-CHA reaction combined with invasive reaction for label-free detection of single nucleotide polymorphisms

G-quadruplex/thioflavin T (G4/THT) is one of the ideal label-free fluorescent light-emitting elements in the field of biosensors due to its good programmability and adaptability. However, the unsatisfactory luminous efficiency of single-molecule G4/THT limits its more practical applications. Here, we developed a G4 embedded semi-catalytic hairpin assembly (G4-SCHA) reaction by rationally modifying the traditional CHA reaction, and combined with the invasive reaction, supplemented by magnetic separation technology, for label-free sensitive detection of single nucleotide polymorphisms (SNPs). The invasive reaction enabled specific recognition of single-base mutations in DNA sequences as well as preliminary signal cycle amplification. Then, magnetic separation was used to shield the false positive signals. Finally, the G4-SCHA was created for secondary amplification and label-free output of the signal. This dual-signal amplified label-free biosensor has been shown to detect mutant targets as low as 78.54 fM. What's more, this biosensor could distinguish 0.01 % of the mutant targets from a mixed sample containing a large number of wild-type targets. In addition, the detection of real and complex biological samples also verified the practical application value of this biosensor in the field of molecular design breeding. Therefore, this study improves a label-free fluorescent light-emitting element, and then proposes a simple, efficient and universal label-free SNP biosensing strategy, which also provides an important reference for the development of other G4/THT based biosensors.

Colorimetric detection of furin based on enhanced catalytic activity of G-quadruplex/hemin DNAzyme

BackgroundRapid and sensitive colorimetric detection methods are crucial for diseases diagnosis, particularly those involving proteases like furin, which are implicated in various conditions, including cancer. Traditional detection methods for furin suffer from limitations in sensitivity and practicality for on-site detection, motivating the development of novel detection strategies. Therefore, developing a simple, enzyme-free, and rapid colorimetric analysis method with high sensitivity for furin detection is imperative. ResultsHerein, we have proposed a colorimetric method in this work for the first time to detect furin, leveraging the assembly of G-quadruplex/hemin DNAzyme with enhanced catalytic activity. Specifically, a peptide-DNA conjugate (PDC) comprising a furin-recognition peptide and flanking DNA sequences for signal amplification is designed to facilitate the DNAzyme assembly. Upon furin treatment, PDC cleavage triggers a cyclic catalytic hairpin assembly reaction to form the complementary double-stranded structures by hairpin 1 (HP1) and hairpin 2 (HP2), bringing the G-quadruplex sequence in HP1 closer to hemin on HP2. Moreover, the resulting G-quadruplex/hemin DNAzymes exhibit robust peroxidase-like activity, enabling the catalysis of the colorimetric reaction of ABTS2− for furin detection. Our method demonstrates high sensitivity, rapid response, and compatibility with complex sample matrices, achieving a detection limit as low as 1.1 pM. SignificanceThe DNAzyme reported in this work exhibits robust catalytic activity, enabling high sensitivity and good efficiency for the detection. By eliminating the requirement for exogenous enzymes, our approach enables visual furin detection without expensive instrumentation and reagents, promising significant utility in biomedical and clinical diagnostic applications. Given the various design of peptide sequence and the programmability of DNA, it can be readily applied to analyzing other useful tumor biomarkers.

High‐Performance Cooperative DNA Nanodevice Enables Sensitive Circular RNA Imaging and Precise Tumor Growth Suppression

The utility of circular RNAs (circRNAs) as emerging biomarkers and regulatory factors in medical diagnostics and therapeutics is hampered by the challenges associated with their sensitive detection and precise modulation. Herein, a high‐performance cooperative DNA nanodevice (HCDN) based on DNA tetrahedron‐confined catalytic DNA assembly reaction (DT‐CDA) that enables both imaging and regulation of circRNAs is developed. Activation of the DT‐CDA is contingent upon the presence of the target circRNA, which, together with a replicative fuel probe, catalyzes the sequential opening of additional DT‐CDAs. This cooperative exponential signal amplification with negligible background interference allows HCDN to effectively detect minute quantities of circRNAs. Employing circSATB2 as a model, the HCDN demonstrates substantial downregulation of Cyclin D1 (CCND1) mRNA and protein levels in cellular and in vivo models, thereby inhibiting tumor growth. The innovative design of HCDN sets the stage for a powerful methodology conducive to enhanced clinical diagnostics and biomolecule manipulation, thereby advancing the capabilities and applications of DNA nanotechnology.