Strand Displacement Reaction Research Articles

Observation of Enantiomeric Switching of Individual Plasmonic Metamolecules.

Active plasmonic metamolecules under microscopic observation are promising for optical reporters in single molecule sensing applications. While self-assembled reconfigurable chiral plasmonic metamolecules can be conveniently engineered with sensing functionalities, their observation is usually based on ensemble measurements, where the chiroptical response of enantiomers tend to cancel each other in ensemble circular dichroism. Herein, we demonstrate microscopic observation of enantiomeric switching of individual active DNA origami-assembled plasmonic metamolecules. The metamolecules are immobilized on a glass substrate in a microfluidic chamber, in which the plasmonic metamolecule can maintain their activities upon certain local stimuli as in solution. In circular differential scattering, two enantiomeric states controlled by the strand-displacement reaction display opposite spectral signals to each other, representing successful enantiomeric switching of the chirality. Moreover, in a close-to-racemic mixture of chiral metamolecules controlled by pH-sensitive strands, the coexistence of enantiomeric individuals, which is concealed in ensemble measurements, is clearly identified.

Temperature-boosted PAM-less activation of CRISPR-Cas12a combined with selective inhibitors enhances detection of SNVs with VAFs below 0.01%

The precise identification of rare single nucleotide variations (SNVs) concomitant with excess wild-type DNA is a valuable method for minimally invasive disease diagnosis and early prediction of drug responsiveness. Selective enrichment of mutant variants via strand displacement reaction offers an ideal approach of SNVs analysis but fails to differentiate wildtype from mutants with variant allele fraction (VAF) < 0.01%. Here, we demonstrate that integration of PAM-less CRISPR-Cas12a and adjacent mutation-enhanced inhibition of wild-type alleles enables highly sensitive measurement of SNVs well below the 0.01% VAF threshold. Raising the reaction temperature to the upper limit of LbaCas12a helps to boost PAM-less activation of collateral DNase activity, which can be further enhanced using PCR additives, leading to ideal discriminative performance for single point mutations. Along with selective inhibitors bearing additional adjacent mutation, it allowed detection of model EGFR L858R mutants down to 0.001% with high sensitivity and specificity. Preliminary investigation on adulterated genomic samples prepared in two different ways also suggests that it can accurately measure ultralow-abundance SNVs extracted directly from clinical samples. We believe that our design, which combines the superior SNV enrichment capability of strand displacement reaction and unparalleled programmability of CRISPR-Cas12a, has the potential to significantly advance current SNV profiling technologies.

Targeted Selection of Aptamer Complementary Elements toward Rapid Development of Aptamer Transducers.

Biosensing using aptamers has been a recent interest for their versatility in detecting many different analytes across a wide range of applications, including medical and environmental applications. In our last work, we introduced a customizable aptamer transducer (AT) that could successfully feed-forward many different output domains to target a variety of reporters and amplification reaction networks. In this paper, we explore the kinetic behavior and performance of novel ATs by modifying the aptamer complementary element (ACE) chosen based on a technique for exploring the ligand-binding landscape of duplexed aptamers. Using published data, we selected and constructed several modified ATs that contain ACEs with varying length, position of the start sites, and position of single mismatches, whose kinetic responses were tracked with a simple fluorescence reporter. A kinetic model for ATs was derived and used to extract the strand-displacement reaction constant k1 and the effective aptamer dissociation constant Kd,eff, allowing us to calculate a relative performance metric, k1/Kd,eff. Comparing our results with the predictions based on the literature data, we provide useful insight into the dynamics of the adenosine AT's duplexed aptamer domain and suggest a high-throughput approach for future ATs to be developed with improved sensitivity. The performance of our ATs showed a moderate correlation to those predicted by the ACE scan method. Here, we find that predicted performance based on our ACE selection method was moderately correlated to our AT's performance.

Bifunctional three-way DNA junction-based strand displacement recycling for amplifiable electrochemical bivariate biosensing

It’s intriguing to utilize the branched arms of three-way DNA junction (3WDJ) for modifying specific recognizing and/or sensing elements of multivariate analytes. Herein, by using two targeting DNA segments (T and T*) specific to SARS-CoV-2 as analyte models, an electrochemical bivariate biosensor was created based on a functional 3WDJ including two-NH2-labeled recognizable probes (RP and RP*) and an assistant probe (AP), while its two branched arms hybridized with four helping DNA blockers. In the electrode surface electrodeposited in HAuCl4, the 3WDJ was stably immobilized via Au-N bonds to specifically recognize and bind T and T*, with which two modified signaling probes by electroactive methylene blue (SP-MB) and ferrocene (SP*-Fc) were introduced to initiate two strand displacement reactions. Resultantly, SP-MB and SP*-Fc were guided to be complementarily hybridized in two arms of 3WDJ, replacing T and T * to execute two individual repeatable recycling for signal amplification. Thus, MB and Fc were oriented proximal to the modified electrode surface for significantly increased electrochemical current signals, respectively dependent on T and T*. With the branched arms of rapidly assembled 3WDJ, the discernible detection of bivariate targets was achievable, showing superb simplification, high sensitivity, and potentially more accurate electrochemical assay of multivariate targets.

Throwing and manipulating and cheating with a DNA nano-dice

Artificial molecular machines have captured the imagination of researchers, given their clear potential to mimic and influence human life. Key to behavior simulation is to reproduce the specific properties of physical or abstract systems. Dice throwing, as a stochastic model, is commonly used for result judgment or plan decision in real life. In this perspective we utilize DNA cube framework for the design of a dice device at the nanoscale to reproduce probabilistic events in different situations: equal probability, high probability, and low probability. We first discuss the randomness of DNA cube, or dice, adsorbing on graphene oxide, or table, and then explore a series of events that change the probability through the way in which the energy released from entropy-driven strand displacement reactions or changes in intermolecular forces. As such, the DNA nano-dice system provides guideline and possibilities for the design, engineering, and quantification of behavioral probability simulation, a currently emerging area of molecular simulation research.

Multi-Reconfigurable DNA Origami Nanolattice Driven by the Combination of Orthogonal Signals.

The progress of the scaffolded DNA origami technology has enabled the construction of various dynamic nanodevices imitating the shapes and motions of mechanical elements. To further expand the achievable configurational changes, the incorporation of multiple movable joints into a single DNA origami structure and their precise control are desired. Here, we propose a multi-reconfigurable 3 × 3 lattice structure consisting of nine frames with rigid four-helix struts connected with flexible 10-nucleotide joints. The configuration of each frame is determined by the arbitrarily selected orthogonal pair of signal DNAs, resulting in the transformation of the lattice into various shapes. We also demonstrated sequential reconfiguration of the nanolattice and its assemblies from one into another via an isothermal strand displacement reaction at physiological temperatures. Our modular and scalable design approach could serve as a versatile platform for a variety of applications that require reversible and continuous shape control with nanoscale precision.

CRISPR-Cas12a Coupled with DNA Nanosheet-Amplified Fluorescence Anisotropy for Sensitive Detection of Biomolecules.

DNA nanosheets (DNSs) have been utilized effectively as a fluorescence anisotropy (FA) amplifier for biosensing. But, their sensitivity needs to be further improved. Herein, CRISPR-Cas12a with strong trans-cleavage activity was utilized to enhance the FA amplification ability of DNSs for the sensitive detection of miRNA-155 (miR-155) as a proof-of-principle target. In this method, the hybrid of the recognition probe of miR-155 (T1) and a blocker sequence (T2) was immobilized on the surface of magnetic beads (MBs). In the presence of miR-155, T2 was released by a strand displacement reaction, which activated the trans-cleavage activity of CRISPR-Cas12a. The single-stranded DNA (ssDNA) probe modified with a carboxytetramethylrhodamine (TAMRA) fluorophore was cleaved in large quantities and could not bind to the handle chain on DNSs, inducing a low FA value. In contrast, in the absence of miR-155, T2 could not be released and the trans-cleavage activity of CRISPR-Cas12a could not be activated. The TAMRA-modified ssDNA probe remained intact and was complementary to the handle chain on the DNSs, and a high FA value was obtained. Thus, miR-155 was detected through the obviously decreased FA value with a low limit of detection (LOD) of 40 pM. Impressively, the sensitivity of this method was greatly improved about 322 times by CRISPR-Cas12a, confirming the amazing signal amplification ability of CRISPR-Cas12a. At the same time, the SARS-CoV-2 nucleocapsid protein was detected by the strategy successfully, indicating that this method was general. Moreover, this method has been applied in the analysis of miR-155 in human serum and the lysates of cells, which provides a new avenue for the sensitive determination of biomarkers in biochemical research and disease diagnosis.

Metal-mediated Fe3O4@polydopamine-aptamer capture nanoprobe coupling multifunctional MXene@Au@Pt nanozyme for direct and portable photothermal analysis of circulating breast cancer cells

Breast cancer (BC) is the most common cancer in the world and circulating tumor cells (CTCs) are reliable biomarkers for early breast cancer diagnosis in a non-invasive manner. However, effective isolation and sensitive detection of BC-CTCs by portable devices in human blood samples are extremely challenging. Herein, we proposed a highly sensitive and portable photothermal cytosensor for direct capture and quantification of BC-CTCs. To achieve efficient isolation of BC-CTCs, aptamer functionalized Fe3O4@PDA nanoprobe was facilely prepared through Ca2+-mediated DNA adsorption. To further detect the captured BC-CTCs with high sensitivity, multifunctional two-dimensional Ti3C2@Au@Pt nanozyme was synthesized, which not only possessed superior photothermal effect but also exhibited high peroxidase-like activity for catalyzing 3,3′,5,5′-tetramethylbenzidine (TMB) to produce TMB oxide (oxTMB) with a strong photothermal characteristic, combining with Ti3C2@Au@Pt to synergistically amplify the temperature signal. Moreover, numerous Ti3C2@Au@Pt nanocomposites would be selectively attained on the BC-CTCs surface through multi-aptamer recognition and binding strategy, which further enhanced the specificity and facilitated signal amplification. Therefore, direct separation and highly sensitive detection of BC-CTCs was successfully achieved in human blood samples. More significantly, the controlled release of the captured BC-CTCs without affecting cell viability could be straightforwardly realized by a simple strand displacement reaction. Thus, with the distinct features of portability, high sensitivity, and easy operation, the current method holds great promise for early diagnosis of breast cancer.

Microswimmer-Based Ultrasensitive Dual-Signal Ratiometric Electrochemical Homogeneous Aptasensor with Functionalized Co-UiO-66 for the Single-Step Detection of HER2 in Whole Blood.

The ability to efficiently detect trace disease biomarkers in whole blood remains an enormous challenge. Researchers have paid more attention to the homogeneous electrochemical ratio biosensor due to its self-calibration capability and improved detection accuracy. However, proportional homogeneous electrochemical sensing is difficult to achieve and typically requires functional modification of the electrode or the preparation of complex materials. Herein, a dual-signal ratiometric aptamer homogeneous electrochemical microswimming detection device with active capture capability and one-step detection of human epidermal growth factor receptor-2 (HER2) is proposed. The homogeneous electrochemical biosensor is fabricated based on a functionalized nanocomposite double-stranded DNA({single-stranded DNA-ferrocene (Fc)-aptamer})@Co-UiO-66 with catalase properties and adsorptive properties to electroactive toluidine blue (TB) molecules. Encapsulation of Co-UiO-66 material with dsDNA (ssDNA-Fc-Apt) containing HER2 aptamer as a gate switch inhibited its ability to adsorb TB molecules. This functionalized Co-UiO-66 material can catalyze hydrogen peroxide. Using hydrogen peroxide as a fuel, it breaks down to release oxygen bubbles, creating a propulsion force that drives dsDNA(ssDNA-Fc-Apt)@Co-UiO-66 target HER2 through whole blood. When the surface dsDNA (ssDNA-Fc-Apt)@Co-UiO-66 recognizes HER2, a strand displacement reaction occurs, and the ssDNA-Fc is released into solution. The HER2 aptamer is coiled because it targets HER2, and the ability to adsorb TB molecules is restored due to the exposed surface of Co-UiO-66. A certain negative voltage is applied to the ITO electrode, and due to the electrostatic attraction, the TB molecules and ssDNA-Fc are adsorbed and enriched on the surface of the electrode by electrostatic attraction, which produces two strong and oppositely changing electrochemical signals, and the electrochemical signals depend on the HER2 concentration. It can sensitively detect HER2 biomarkers in only 40 min with the detection range of 0.0001-10 ng/mL and detection limits as low as 10 fg/mL. The electrochemical microswimmer for the detection of trace disease biomarkers involves a one-step process of capture, signal change, and detection.

Enzyme-Free Nucleic Acid Circuits for Fold-Change Detection.

Fold-change detection is widespread in sensory systems of various organisms. Dynamic DNA nanotechnology provides an important toolbox for reproducing structures and responses of cellular circuits. In this work, we construct an enzyme-free nucleic acid circuit based on the incoherent feed-forward loop using toehold-mediated DNA strand displacement reactions and explore its dynamic behaviors. The mathematical model based on ordinary differential equations is used to evaluate the parameter regime required for fold-change detection. After selecting appropriate parameters, the constructed synthetic circuit exhibits approximate fold-change detection for multiple rounds of inputs with different initial concentrations. This work is anticipated to shed new light on the design of DNA dynamic circuits in the enzyme-free environment.

A Theoretical Framework for Implementable Nucleic Acids Feedback Systems.

Chemical reaction networks can be utilised as basic components for nucleic acid feedback control systems' design for Synthetic Biology application. DNA hybridisation and programmed strand-displacement reactions are effective primitives for implementation. However, the experimental validation and scale-up of nucleic acid control systems are still considerably falling behind their theoretical designs. To aid with the progress heading into experimental implementations, we provide here chemical reaction networks that represent two fundamental classes of linear controllers: integral and static negative state feedback. We reduced the complexity of the networks by finding designs with fewer reactions and chemical species, to take account of the limits of current experimental capabilities and mitigate issues pertaining to crosstalk and leakage, along with toehold sequence design. The supplied control circuits are quintessential candidates for the first experimental validations of nucleic acid controllers, since they have a number of parameters, species, and reactions small enough for viable experimentation with current technical capabilities, but still represent challenging feedback control systems. They are also well suited to further theoretical analysis to verify results on the stability, performance, and robustness of this important new class of control systems.

Modular, Articulated Models of DNA and Peptide Nucleic Acids for Nanotechnology Education

ABSTRACT Dynamic and flexible nucleic acid models can provide current and future scientists with physical intuition for the structure of DNA and the ways that DNA and its synthetic mimics can be used to build self-assembling structures and advanced nanomachines. As more research labs and classrooms dive into the field of structural nucleic acid nanotechnology, students and researchers need access to interactive, dynamic, handheld models. Here, we present a 3D-printable kit for the construction of DNA and peptide nucleic acid (PNA). We have engineered a previous modular DNA kit to reduce costs while improving ease of assembly, flexibility, and robustness. We have also expanded the scope of available snap-together models by creating the first 3D-printable models of γPNA, an emerging material for nuclease- and protease-resistance nanotechnology. Building on previous research, representative nucleic acid duplexes were split into logical monomer segments, and atomic coordinates were used to create solid models for 3D printing. We used a human factors approach to customize 3 types of articulated snap-together connectors that allow for physically relevant motion characteristic of each interface in the model. Modules are easy to connect and separate manually but stay together when the model is manipulated. To greatly reduce cost, we bundled these segments for printing, and we created a miniaturized version that uses less than half the printing material to build. Our novel 3D-printed articulated snap-together models capture the flexibility and robustness of DNA and γPNA nanostructures. Resulting handheld helical models replicate the geometries in published structures and can now flex to form crossovers and allow biologically relevant zipping and unzipping to allow complex demonstrations of nanomachines undergoing strand displacement reactions. Finally, the same tools used to create these models can be readily applied to other types of backbones and nucleobases for endless research and education possibilities.

Aggregation-Induced Electrochemiluminescence Frame of Silica-Confined Tetraphenylethylene Derivative Matrixes for CD44 Detection via Peptide Recognition.

The theory of aggregation-induced electrochemiluminescence (AIECL) has introduced new vitality into preparing new electrochemiluminescence (ECL) emitters. However, the progress in the application of biosensing analysis has been slow owing to the lack of AIECL-based functional nanomaterials. Herein, a biosensor was fabricated using mesoporous silica nanosphere (MSN) matrix-confined 1,1,2,2-tetra(4-carboxylphenyl)ethylene (TPE) as a well-ordered ECL emitter and self-designed WHPWSYC (WC-7) heptapeptide as the target capturer for CD44 detection. TPE and its co-reactant, triethylamine (TEA), were encapsulated in the MSN nanomatrix to enhance the radiation transition by limiting the intramolecular rotation of TPE molecule benefit from the spatial confinement effect, and the ECL intensity is self-enhanced by replacing electron free diffusion in the conventional ECL system. MSN-TPE-TEA can act as satisfactory sensing substrates that improve the reproducibility and batch-to-batch consistency of biosensors and functions as a stable signal label for trace analysis of biomarkers. As a substitute for antibody and hyaluronic acid, the WC-7 heptapeptide significantly reduced the steric hindrance of the sensing interface in CD44 affinity tests. Combined with the DNA strand displacement reaction, this strategy shows a good ECL response to standard CD44 antigen and MCF-7 cells with different concentrations, which is another feasible method for detecting CD44 in body fluids or living cells.

Assembly of DNA triangular pyramid frustum for ultrasensitive quantification of exosomal miRNA

Early screening of biomarkers benefits therapy and prognosis of cancers. MiRNAs encapsulated in tumor-derived exosomes are emerging biomarkers for early diagnosis of cancers. Nevertheless, traditional methods suffer certain drawbacks, which hamper their wide applications. In this contribution, we have developed a convenient electrochemical approach for quantification of exosomal miRNA based on the assembly of DNA triangular pyramid frustum (TPF) and strand displacement amplification. Four single-stranded DNA helps the formation of primary DNA triangle with three thiols for gold electrode immobilization at the bottom and three amino groups on overhangs for the capture of silver nanoparticles. On the other hand, target miRNA induced strand displacement reaction produces abundant specific DNA strands, which help the DNA structural transition from triangle to TPF. Amino groups are thus hidden and the declined silver stripping current can be used for the evaluation of target miRNA concentration. This biosensor exhibits excellent analytical performances and successfully achieves analysis of exosomal miRNAs from cells and clinical serum samples.

Speed and Correctness Guarantees for Programmable Enthalpy-Neutral DNA Reactions†.

Molecular control circuits embedded within chemical systems to direct molecular events have transformative applications in synthetic biology, medicine, and other fields. However, it is challenging to understand the collective behavior of components due to the combinatorial complexity of possible interactions. Some of the largest engineered molecular systems to date have been constructed using DNA strand displacement reactions, in which signals can be propagated without a net change in base pairs (enthalpy neutral). This flexible and programmable component has been used for constructing molecular logic circuits, smart structures and devices, for systems with complex autonomously generated dynamics, and for diagnostics. Limiting their utility, however, strand displacement systems are susceptible to the spurious release of output in the absence of the proper combination of inputs (leak), as well as reversible unproductive binding (toehold occlusion) and spurious displacement that slow down desired kinetics. We systematize the properties of the simplest enthalpy-neutral strand displacement cascades (logically linear topology), and develop a taxonomy for the desired and undesired properties affecting speed and correctness, and trade-offs between them based on a few fundamental parameters. We also show that enthalpy-neutral linear cascades can be engineered with stronger thermodynamic guarantees to leak than non-enthalpy-neutral designs. We confirm our theoretical analysis with laboratory experiments comparing the properties of different design parameters. Our method of tackling the combinatorial complexity using mathematical proofs can guide the engineering of robust and efficient molecular algorithms.

Dual-Mode Logic Gate for Intelligent and Portable Detection of MicroRNA Based on Gas Pressure and Lateral Flow Assay.

Molecular logic gate provides an intelligent option for simultaneous detection of biomarkers. Herein, a dual-mode DNA logic gate was proposed to portably and intelligently detect multiple microRNAs (miRNAs) by gas pressure biosensing and lateral flow assay (LFA). A platinum-coated gold nanoparticle (Au@PtNP) with catalase-like activity was used as a signal reporter to achieve a dual-signal readout. MiRNAs as the input initiated the cyclic strand displacement reaction (SDR) to enrich a large amount of Au@PtNPs. Thus, miRNA can be visually detected by a lateral flow strip (LFS) using the grayish-brown color of Au@PtNPs as output 1. Furthermore, Au@PtNP-catalyzed decomposition of H2O2 resulted in gas pressure as output 2, which was measured by a digital and handheld gas pressure meter. As a consequence, microRNA 21 (miR-21) was sensitively and reliably detected with the limit of detection (LOD) of 7.2 pM. The selectivity and real sample analysis were both satisfactory. Significantly, two-input and three-input AND logic gates were successfully developed to realize multiple detection of two miRNAs and three miRNAs, which provide a promising way for intelligent multi-input analysis. Predictably, with the advantages of portability, simplicity, and affordability, the dual-mode logic gate based on gas pressure biosensing and LFA offers a new perspective on the field of intelligent and portable biosensing and bioanalysis.

A multivalent dual lock-and-key system and its toehold-mediated orthogonal activation for highly specific strand displacement amplification

DNA strand displacement reaction plays an essential role in dynamic DNA nanotechnology and has proven powerful in DNA nano-computers and biosensors. However, the poor specificity and/or kinetics hamper its practical applications. Here we report a multivalent dual lock-and-key (Multi-DLK) system that can be accurately activated to perform a toehold-mediated orthogonal DNA circuitry (ODC) for highly specific and rapid nucleotide polymorphism assay. The “dual lock” can be opened only when target as the unique “key” is introduced and perform matching operation twice in an exact order separately by using the two fragments of target as inputs. The two operations are designed based on toehold-mediated strand displacement reaction, separately named TSDR1 and TSDR2, offering an orthogonal activation strategy for accurate identification. After that, the palindromic end of linker strand is released and taken as self-primer by DNA polymerase to perform a highly target-specific strand displacement amplification (TSSDA). We demonstrate that Multi-DLK system comes with the benefits of orthogonal activation strategy and TSSDA-based amplifier, which make the system ideally suitable for gene typing with a high specificity and efficiency. Comparative analysis and the final application of Multi-DLK system for real samples have shown the great potential for accurate and timely diagnosis.

Extracellular ATP-activated hybridization chain reaction for accurate and sensitive detection of cancer cells

Accurate and sensitive detection of cancer cells is of significant importance for early diagnosis and treatment of cancer. Here, we developed an extracellular ATP-activated hybridization chain reaction (HCR) amplification strategy to meet this purpose. This strategy relies on three DNA probes, Apt-trigger, H1-ATP aptamer duplex and hairpin H2. The Apt-trigger probe consists of two components: an aptamer sequence for specific recognition of the target cells, and a trigger sequence for the HCR assembly. The duplex structure of H1-ATP aptamer causes the toehold in hairpin H1 to be hidden, preventing the strand-displacement reaction between hairpin H1 and Apt-trigger. Upon activation with ATP, the ATP aptamer will bind to ATP to dissociate from hairpin H1, thus leading to an Apt-trigger-induced strand-displacement reaction and subsequent HCR with hairpin H2 on the target cell surface. Benefiting from aptamer recognition and ATP-activated HCR amplification, this strategy can not only perform sensitive quantitative analysis with a detection limit of 25 cells in 200 µL of binding buffer, but also show desirable specificity and accuracy for identifying target cells from control cells and mixed cell samples. Importantly, this method retains stable and good performance for target cell detection in 10% fetal bovine serum, demonstrating great potential for clinical diagnosis in complex biological matrices. Furthermore, this strategy can be adapted to detect various types of cancer cells by changing the corresponding aptamer sequence.

Dumbbell Aptamer Sensor Based on Dual Biomarkers for Early Detection of Alzheimer's Disease.

Finding a timely, sensitive, and noninvasive detection method has become an urgent need for asymptomatic early diagnosis of Alzheimer's disease (AD). MicroRNA-193b (miR-193b) and Aβ42 oligomers (AβO42) in neurogenic exosomes were confirmed to reflect pathological changes in the AD early stage. The combination of two biomarkers is promising for the earlier detection of AD. In this study, a detection system based on the principle of the entropy-driven strand displacement reaction (ESDR) was developed, including a dumbbell detection probe (H), an indicator probe (R), and graphene oxide (GO). In the detection system, the two hairpins of H were opened by the interaction of miR-193b (T1) and AβO42 (T2) with the aptamer. Then R hybridized with H and began to displace T, initiating the next round of ESDR to achieve sensitive detection of T. GO specifically adsorbed free R and quenched the fluorescence, further reducing the intensity of the background signal. Both of these points provided the system with a more sensitive analytical performance. The detection limit of miR-193b was 77 pM and the detection limit of AβO42 was 53 pM. This sensor detected the change of "one increase (AβO42) and one decrease (miR-193b)" in the exosome sample. Additionally, results showed that this detection system could distinguish the model of early AD from the non-AD control, which was sufficient for earlier and more sensitive detection of AD. This strategy has strong specificity, high sensitivity, and easy operation, which provides broad prospects for the early diagnosis of AD.

Cooperative Amplification of Au@FeCo as Mimetic Catalytic Nanozymes and Bicycled Hairpin Assembly for Ultrasensitive Electrochemical Biosensing.

Exploring the cooperative amplification of peroxidase-like metal nanocomposites and cycled hairpin assembly is intriguing for sensitive bioanalysis. Herein, we report the first design of a unique electrochemical biosensor based on mimicking Au@FeCo nanozymes and bicycled hairpin assembly (BHA) for synergistic signal amplification. By loading the enzyme-like FeCo alloy in Au nanoparticles (AuNPs), the as-synthesized Au@FeCo hybrids display great improvement of electronic conductivity and active surface area and excellent mimic catalase activity to H2O2 decomposition into •OH radicals. The immobilization of Au@FeCo in an electrode sensing interface is stabilized via the resulting electrodeposition in HAuCl4 while efficiently accelerating the electron transfer of electroactive ferrocene (Fc). Upon the immobilization of a helping hairpin (HH) via Au-S bonds, a specific DNA trigger (T*) is introduced to activate BHA operation through competitive strand displacement reactions among recognizing hairpin (RH), signaling hairpin (SH), and HH. T* and RH are rationally released to catalyze two cycles, in which the transient depletion of dsDNA intermediates rapidly drives the progressive hairpin assemblies to output more products SH·HH. Thus, the efficient amplification of Au@FeCo mimic catalase activity combined with BHA leads to a significantly increased current signal of Fc dependent on miRNA-21 analogous to T*, thereby directing the creation of a highly sensitive electrochemical biosensor having applicable potential in actual samples.