3D DNA Nanomachine Research Articles

Bimodal 3D DNA nanomachines coupling with bioconjugates powered electrochemical and visual-driven dual-enzyme cascade sensing of Let-7a

A dual-enzyme self-powered biosensor utilizing bimodal 3D DNA nanomachines and gold nanoparticle (AuNPs)/carbon/disulfide molybdenum hollow nanorods bioconjugates is developed for the electrochemical and visual detection of the tumor biomarker let-7a. DNA probes are immobilized on a flexible electrode substrate of AuNPs/graphdiyne/carbon cloth to improve probe loading capacity and electron transfer efficiency. The presence of let-7a initiates the work of dual-enzyme 3D DNA nanomachine, and the output DNA single strand cleaved by Nb.BbvCI is served as a bridge to connect glucose oxidase (GOD)-DNA conjugate. The GOD coordinates with the cathodic enzyme bilirubin oxidase and produces an increased open circuit voltage signal, which is attributed to the hexaammine ruthenium in the double-stranded adsorption system of the biocathode. By utilizing the H2O2 generated during the oxidation process of GOD, a linear relationship between the color of 3,3′,5,5′-tetramethylbenzidine and the concentration of let-7a is established, enhancing the reliability of the detection results through the independent signal conversion mechanisms of electrochemical/colorimetric modes. The results demonstrate a wide dynamic range of 0.0001–0.1 pM with detection limits of 31.04 aM (S/N=3) in electrochemical mode and 59.99 aM (S/N=3) in the colorimetric mode. This dual-mode biosensing strategy can be expanded to detect additional nucleic acid biomarkers, providing a promising support and reference for biological analysis and medical diagnosis.

A dual-core 3D DNA nanomachine based on DNAzyme positive feedback loop for highly sensitive MicroRNA imaging in living cells

A double 3D DNA walker nanomachine by DNAzyme self-driven positive feedback loop amplification for the detection of miRNA was constructed. This method uses two gold nanoparticles as the reaction core, and because of the spatial confinement effect the local concentration of the reactants increase the collision efficiency was greatly improved. Meanwhile, the introduction of positive feedback loop promotes the conversion efficiency. In presence of miRNA-21, a large amount of DNAzyme was released and hydrolyze the reporter probe, resulting the recovery of fluorescence signal. The linear range for miRNA-21 is 0.5–60 pmol/L, and the detection limit is 0.41 pmol/L (S/N = 3). This nanomachine has been successfully used for accurate detection of miRNA-21 expression levels in cell lysates. At the same time, it can enter cells for intracellular miRNA-21 fluorescence imaging, distinguishing tumor cells from normal cells. This combination of in vitro detection and imaging analysis of living cells can achieve the goal of jointly detecting cancer markers through multiple pathways, providing new ideas for early diagnosis and screening of diseases.

Dual-responsive 3D DNA nanomachines cascaded hybridization chain reactions for novel self-powered flexible microRNA-detecting platform

The microRNA-21 is closely related to chromatin remodeling and epigenetic regulation. In this work, an efficient double-response 3D DNA nanomachine (DRDN) was assembled by co-immobilizing two different lengths of hairpin DNA on the surface of gold nanoparticles (AuNPs) to capture microRNA-21 (miRNA-21), recycle miRNA-21, and trigger hybridization chain reactions (HCR). This work reports the fabrication of a laser-scribed graphene (LSG) electrode with excellent flexibility and electrical conductivity by laser-scribing commercial polyimide films (PI). The as-proposed self-powered biosensing platform presents significantly increased instantaneous current to in real-time monitor miRNA-21 by a capacitor. The biosensing platform exhibited highly sensitive detection of miRNA-21 with a detection limit of 0.142 fM in the range of 0.5 fM to 1 × 104 fM, and demonstrated high efficiency in the analysis of the tumor markers.

Thermodynamic Allosteric Switch-Actuated 3D DNA Nanomachine for Ultrasensitive Electrochemical/Fluorescent Dual-Mode Biosensing of a Transcription Factor.

Herein, we reported an innovative thermodynamic allosteric switch-actuated 3D DNA nanomachine for selective, sensitive, and accurate electrochemical (EC)/fluorescent (FL) dual-mode biosensing of a microphthalmia-associated transcription factor (MITF). The thermodynamic allosteric switch was ingeniously customized as a hairpin probe (HP) that was in dynamic equilibrium but rapidly interconverting conformations. At the "inactive state", the MITF-binding region and the switch part were "sequestered". Upon the introduction of MITF, an MITF-HP complex promptly formed, and the equilibrium of HP thermodynamically inclined from the "inactive state" toward the "active state" conformation. Immediately, the exposed switch on HP effectively actuated the 3D DNA nanomachine and synchronously produced the restriction site for Nb.BbvCI nicking endonuclease. After the autonomous conveying of the 3D DNA nanomachine by means of the high-efficiency circularly nicking endonuclease signal amplification (NESA), not only was MB-S1 in the supernatant used for FL measurements but also MB-SP/MNs/S2 in the precipitate was adapted for EC analysis, significantly improving the utilization of output products derived from the 3D DNA nanomachine. Accordingly, benefiting from the efficient DNA nanomachine signal amplification manner and the self-calibration function of a dual-mode bioassay, the constructed biosensor exhibits superior sensitivity and accuracy for MITF determination.

CRISPR-Cas12a-enhanced mass spectrometric DNA nanomachine for HPV-16 detection in human serum.

Cervical cancer is the second most harmful cancer to women, while human papillomavirus (HPV)-16 is the main cause of cervical cancer. Early virus screening can reduce the risk of death, but is sometimes challenged by the low concentration and complicated matrix in physiological samples such as human serum. Herein, we report a CRISPR-Cas12a-enhanced mass spectrometric 3D DNA nanomachine for HPV-16 detection in human serum. Thanks to the high sensitivity of the CRISPR-Cas12a-enhanced 3D DNA nanomachine, and the high matrix-tolerance of elemental mass spectrometry, the proposed method was successfully applied for direct human serum analysis with a low detection limit of 1.2 pM.

Integrating Reliable Pt-S Bond-Mediated 3D DNA Nanomachine with Magnetic Separation in a Homogeneous Electrochemical Strategy for Exosomal MicroRNA Detection with Low Background and High Sensitivity.

Precise and sensitive analysis of exosomal microRNA (miRNA) is of great importance for noninvasive early disease diagnosis, but it remains a great challenge to detect exosomal miRNA in human blood samples because of their small size, high sequence homology, and low abundance. Herein, we integrated reliable Pt-S bond-mediated three-dimensional (3D) DNA nanomachine and magnetic separation in a homogeneous electrochemical strategy for the detection of exosomal miRNA with low background and high sensitivity. The 3D DNA nanomachine was easily prepared via a facile and rapid freezing method, and it was capable of resisting the influence of biothiols, thus endowing it with high stability. Notably, the as-developed magnetic 3D DNA nanomachine not only enabled the detection system to have a low background but also coupled with liposome nanocarriers to synergistically amplify the current signal. Consequently, by ingeniously combining the low background and multiple signal-amplification strategies in homogeneous electrochemical biosensing, highly sensitive detection of exosomal miRNA was successfully achieved. More significantly, with good anti-interference ability, the as-proposed method could effectively discriminate plasma samples from cancer patients and healthy subjects, thus showing a high potential for application in the nondestructive early clinical diagnosis of disease.

Efficient Three-Dimensional DNA Nanomachine Guided by a Robust Tetrahedral DNA Nanoarray Structure for the Rapid and Ultrasensitive Electrochemical Detection of Matrix Metalloproteinase 2.

Herein, a giant-sized DNA nanoarray was subtly assembled by two kinds of independent tetrahedral DNA structures as the DNA track for a multi-armed three-dimensional (3D) DNA nanomachine to perform signal transduction and amplification efficiently, which was developed as an electrochemical biosensor for the rapid and ultrasensitive detection of matrix metalloproteinase 2 (MMP-2). Impressively, in contrast to conventional DNA walkers with inefficiency, which walked on random DNA tracks composed of a two-dimensional (2D) probe or a one-dimensional (1D) single-stranded (ss)DNA probe, the multi-armed 3D DNA nanomachine from exonuclease III (Exo III) enzyme-assisted target recycling amplification would be endowed with faster reaction speed and better walking efficiency because of the excellent rigidity and orderliness of the tetrahedral DNA nanoarray structure. Once the hairpin H3-label with the signal substance ferrocene (Fc) was added to the modified electrode surface, the multi-armed 3D DNA nanomachine would be driven to move along the well-designed nanoarray tracks by toehold-mediated DNA strand displacement, resulting in most of the ferrocene (Fc) binding to the electrode surface and a remarkable increase in electrochemical signals within 60 min. As a proof of concept, the prepared biosensor attained a low detection limit of 11.4 fg/mL for the sensitive detection of the target MMP-2 and was applied in Hela and MCF-7 cancer cell lysates. As a result, this strategy provided a high-performance sensing platform for protein detection in tumor diagnosis.

Dual-layer 3D DNA nanostructure: The next generation of ultrafast DNA nanomachine for microRNA sensing and intracellular imaging

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

A label-free and ratiometric fluorescent immunosensor by integrating CRISPR/Cas 12a and 3D DNA nanomachine

Sensitive, selective, especially label-free detection of disease relevant protein biomarkers is highly valuable for clinical diagnosis and disease process monitoring but remains a challenge. Herein, with prostate specific antigen (PSA) as a model target, we have designed a label-free ratiometric fluorescent immunosensor based on three-dimensional (3D) DNA walker and the powerful clustered regularly interspaced short palindromic repeat (CRISPR)/Cas system, with poly(thymine) templated Cu nanoclusters and DAPI as signals. The aptamer-linked activator DNA was designed to convert target recognition event into activate deoxyribonuclease activity of CRISPR/Cas12a system and further cleave oligonucleotides. The cleavage behavior leads to the quenching of copper nanoclusters and stagnation of the cascade amplification progress of 3D DNA nanomachine. The DNA machine is constructed from primer DNA and silica nanospheres loaded by double-stranded DNA (dsDNA) with prominent 3′ end, and movement of the primer DNA is powered by exonuclease (Exo) III via a burnt-bridge mechanism. Without primer DNA, the dyes (DAPI) were embedded in the complementary DNA and emit strong fluorescence. The practical applicability of the biosensor was demonstrated by assay of PSA in human serum with satisfactory result. In addition, this completely label-free method, reduced consumption of antibody, greatly simplify the procedures and saving cost.

A novel nanoparticle surface-constrained CRISPR-Cas12a 3D DNA walker-like nanomachines for sensitive and stable miRNAs detection

The CRISPR-Cas system has broad prospects as a new type of nucleic acid signal amplification technology based on the trans-cleavage activity of Cas12a to single-stranded DNA, but the trans-cleavage reaction efficiency is relatively low in solution. In order to overcome this negative factor, a new 3D DNA nanomachine whose CRISPR-Cas12a is limited to the surface of nanoparticles is used for sensitive and stable detection of miRNA. By loading Cas12a activator onto spherical nucleic acid (SNA), the CRISPR-Cas12a activator system on the surface of Au nanoparticles (AuNPs) acts as a walker to carry out continuous recognition-walking-cutting reaction on the surface of AuNPs, which enhances the trans-cleavage activity of Cas12a to SNAs. Benefiting from the confinement effect of spherical nucleic acids surface, a 3D DNA nanomachine has been developed for the detection of miRNA-21, which has achieved high sensitivity and accuracy, and the detection limit is able to reach 8.0 pM. This new 3D DNA walker-like nanomachine provided another insight for future bioanalysis and early clinical diagnoses of disease and liquid biopsy.

Electrochemiluminescence detection of miRNA-21 based on dual signal amplification strategies: Duplex-specific nuclease -mediated target recycle and nicking endonuclease-driven 3D DNA nanomachine

DNA nanomachines have shown potential application in the construction of various biosensors. Here, an electrochemiluminescence biosensor for the sensitive detection of miRNA-21 were reported based on three-dimensional (3D) DNA nanomachine and duplex-specific nuclease (DSN)-mediated target recycle amplification strategy. First, the bipedal DNA walkers were obtained by DSN-mediated digestion reaction initiated by target miRNA-21.3D DNA tracks were prepared by modifying Fe3O4 magnetic beads (MBs) with ferrocene-labeled DNA (Fc-DNA). The produced DNA walkers autonomously moved along 3D DNA tracks powered by nicking endonuclease. During the movement, ferrocene-labeled DNA was cleaved, resulting in large amounts of Fc-labeled DNA fragments away from the MBs surface. Finally, the liberated Fc-labeled DNA fragments were dropped on the C-g-C3N4 modified electrode surface, leading to the quenching of C-g-C3N4 electrochemiluminescence (ECL). Benefiting from the dual amplification strategy of 3D DNA nanomachine and DSN-mediated target recycling, the developed ECL biosensor exhibited an excellent performance for miRNA-21 detection with a wide linear range of 10 fM to 10 nM and a low detection limit of 1.0 fM. This work offers a new thought for the application of DNA walkers in the construction of various biosensors.

An all-graphdiyne electrochemiluminescence biosensor for the ultrasensitive detection of microRNA-21 based on target recycling with DNA cascade reaction for signal amplification.

Graphdiyne oxide quantum dots (GDYO QDs), as derivatives of graphdiyne (GDY), have excellent electroconductibility and luminous properties and can be applied as a new ECL emitter. Herein, an electrochemiluminescence (ECL) biosensor for miRNA-21 ultrasensitive determination is constructed based on AuNPs/GDY, GDYO QD and oligonucleotide signal amplification strategy that integrates DNA walker and hybridization chain reaction (HCR) amplification. As electrode substrate material, AuNPs/GDY can not only bond with the aptamer CP but can also enhance the conductivity of the interface. When miRNA-21 exists, the DNA walker process is initiated, and the signaling probes are introduced on the electrode surface, producing abundant double-stranded H1/H2; then, H3/H4 undergoes complementary base pairing with H1/H2 through HCR. With the increase in miRNA-21, the 3D DNA nanomachine is actively manipulated, resulting in a gradual increase in ECL signal. This ECL biosensor demonstrates outstanding performance in the determination of miRNA-21 in the linear range from 0.1 fM to 1 nM. This study offers a new sensitive idea for the clinical analysis of cancer biomarkers.

Pyrene-Based Hydrogen-Bonded Organic Frameworks as New Emitters with Porosity- and Aggregation-Induced Enhanced Electrochemiluminescence for Ultrasensitive MicroRNA Assay.

Exploring new electrochemiluminescence (ECL) luminophores with strong ECL emission is highly desirable for developing ultrasensitive ECL sensors. Herein, a pyrene-based hydrogen-bonded organic framework (Py-HOF) featuring prominent ECL performance was prepared by utilizing 1,3,6,8-tetrakis(p-benzoic acid) pyrene (H4TBAPy) with an aggregation-induced enhanced emission (AIEE) property as a building block, exhibiting a stronger ECL emission than those of H4TBAPy monomers, H4TBAPy aggregates, the low-porosity Py-HOF-210 °C and Py-HOF-180 °C. We have coined the term "the porosity- and aggregation-induced enhanced ECL (PAIE-ECL)" for this intriguing phenomenon. The Py-HOF displayed superb and stable ECL intensity, not only because the luminophore H4TBAPy was assembled into the Py-HOF via four pairs of O-H···O hydrogen bonds, which constrained the intramolecular movements to reduce nonradiative transition, but also because the H4TBAPy in Py-HOF was stacked in a slipped face-to-face mode to form J-aggregates that benefited the ECL enhancement. Furthermore, the high porosity of Py-HOF allowed the enrichment of coreactants and facilitated the migration of ions, electrons, and coreactants, which made it possible for the inner and outer H4TBAPy to be electrochemically excited. Considering the remarkable ECL performance, Py-HOF was first employed as an ECL probe combined with a 3D DNA nanomachine amplification strategy to assemble a hypersensitive "on-off" ECL sensor for the microRNA-141 assay, presenting a satisfactory linear range (100 aM to 1 nM) with a detection limit of 14.4 aM. The PAIE-ECL manifested by Py-HOF provided a bright avenue for the design and synthesis of outstanding HOF-based ECL materials and offered new opportunities for the development of ECL biosensors with excellent sensitivity.

Wavelength-resolved photoelectrochemical biosensor triggered by cascade signal amplification reactions for RNA methylation analysis on a single interface

Among ribonucleic acid (RNA) methylation modifications, those of m6A and m5C are the two most abundant forms, and the bias of m6A and m5C modification levels plays an essential role in many biological processes. Therefore, constructing a mechanism that can simultaneously analyze the two modifications would be potentially very significant. Herein, we propose a wavelength-resolved photoelectrochemical (PEC) biosensor based on cascade signal amplification reactions for m6A-RNA and m5C-RNA analysis on a single interface. First, a target sequence was transformed with antibody-functionalized magnetic beads to obtain the initial sequences, triggering the 3D DNA nanomachine to transform and amplify targets by generating many output sequences. Subsequently, the output sequences were combined with rolling-circle-amplification (RCA) primer scaffolds, and then a DNA origami structure was formed based on RCA. A DNA origami could immobilize many photoelectrochemical mediators to amplify photocurrent responses. As expected, the proposed PEC biosensor exhibited excellent analytical performances for both m6A-RNA and m5C-RNA. The detection limits were as low as 1.96 fM and 7.37 pM, respectively. The successful fabrication of the PEC biosensor opened new possibilities for epigenetic research and the diagnosis and treatment of methylation-related diseases.

A novel self-enhancement NCNDs-BPEI-Ru nanocomposite with highly efficient electrochemiluminescence as signal probe for ultrasensitive detection of MTB

Herein, a novel self-enhanced Ru(Ⅱ)-based nanocomposite (NCNDs-BPEI-Ru), synthesized by covalently connecting nitrogen-doped carbon nanodots (NCNDs) and tris (4,4′-dicarboxylicacid-2,2′-bipyridyl) ruthenium (II) dichloride (Ru(dcbpy)3Cl2) through branched polyethyleneimine (BPEI), could be used as an electrochemiluminescence (ECL) signal probe for ultrasensitive detection of mycobacterium tuberculosis (MTB) DNA fragment. Without any extraneous coreactants, NCNDs-BPEI-Ru could generate an ultra-strong initial ECL signal on the glassy carbon electrode (GCE) (signal-on) due to the coreaction from BPEI and NCNDs to Ru(dcbpy)3Cl2. Then, self-assembled DNA nanotubes embedding methylene blue molecules (MB-DNANTs) were modified on the nanocomposite to effectively quench the ECL intensity of NCNDs-BPEI-Ru owing to the quenching effect of MB for Ru(dcbpy)3Cl2 (signal-off). Finally, the 3D DNA nanomachine, formed by a DNA-functionalized magnetic beads (DNA-MBs) could transform trace target MTB into massive output DNA, and the MB-DNANTs could be disintegrated to get ECL signal recovery (signal-on) due to hybridization of output DNA and the S2 in DNANTs. Therefore, the proposed biosensor exhibited predominant performance for the detection of MTB DNA fragment with a low detection limit of 1.4 aM, which showed great application value in the detection of biomarkers in clinical diagnosis.

Dual 3D DNA Nanomachine-Mediated Catalytic Hairpin Assembly for Ultrasensitive Detection of MicroRNA.

Herein, we designed a dual 3D DNA nanomachine (DDNM)-mediated catalytic hairpin assembly (DDNM-CHA) to construct an electrochemical biosensor for ultrasensitive detection of miRNA, which possesses quite a faster reaction rate and much higher amplification efficiency than those of traditional catalytic hairpin assembly (CHA). Impressively, since the DDNM skillfully increases the local concentration of reactants and decreases the steric hindrance of substrates simultaneously, the DDNM-CHA could be endowed with higher collision efficiency and more effective reaction compared with traditional CHA, resulting in a hyper conversion efficiency up to 2.78 × 107 only in 25 min. This way, the developed DDNM-CHA could easily conquer the main predicaments: long reaction time and low efficiency. As a proof of the concept, we adopt the gold nanoparticles (AuNPs) and the magnetic nanoparticle (Fe3O4) as the kernel of DNM-A and DNM-B, respectively, and harness the magnetic electrode to directly adsorb the products H1-H2/Fe3O4 for constructing an immobilization-free biosensor for high-speed and ultrasensitive detection of miRNA with a detection limit of 0.14 fM. As a result, the DDNM-CHA we developed carves out a new insight to design a functional DNA nanomachine and evolve the analysis method for practical amplification in the sensing area and promotes the deeper exploration of the nucleic acid signal amplification strategy and DNA nanobiotechnology.

A core–brush 3D DNA nanostructure: the next generation of DNA nanomachine for ultrasensitive sensing and imaging of intracellular microRNA with rapid kinetics

A highly loaded and integrated core–brush three-dimensional (3D) DNA nanostructure is constructed by programmatically assembling a locked DNA walking arm (DA) and hairpin substrate (HS) into a repetitive array along a well-designed DNA track generated by rolling circle amplification (RCA) and is applied as a 3D DNA nanomachine for rapid and sensitive intracellular microRNA (miRNA) imaging and sensing. Impressively, the homogeneous distribution of the DA and HS at a ratio of 1 : 3 on the DNA track provides a specific walking range for the DA to avoid invalid and random self-walking and notably improve the executive ability of the core–brush 3D DNA nanomachine, which easily solves the major technical challenges of traditional Au-based 3D DNA nanomachines: low loading capacity and low executive efficiency. As a proof of concept, the interaction of miRNA with the 3D DNA nanomachine could initiate the autonomous and progressive operation of the DA to cleave the HS for ultrasensitive ECL detection of target miRNA-21 with a detection limit as low as 3.57 aM and rapid imaging in living cells within 15 min. Therefore, the proposed core–brush 3D DNA nanomachine could not only provide convincing evidence for sensitive detection and rapid visual imaging of biomarkers with tiny change, but also assist researchers in investigating the formation mechanism of tumors, improving their recovery rates and reducing correlative complications. This strategy might enrich the method to design a new generation of 3D DNA nanomachine and promote the development of clinical diagnosis, targeted therapy and prognosis monitoring.

A Target-Initiated Autocatalytic 3D DNA Nanomachine for High-Efficiency Amplified Detection of MicroRNA

A Target-Initiated Autocatalytic 3D DNA Nanomachine for High-Efficiency Amplified Detection of MicroRNA

Aggregation-Induced Synergism by Hydrophobic-Driven Self-Assembly of Amphiphilic Oligonucleotides.

The evident contradiction between high local-concentration-based substrate reactivity and free-diffusion-based high reaction efficiency remains one of the important challenges in chemistry. Herein, we propose an efficient aggregation-induced synergism through the hydrophobic-driven self-assembly of amphiphilic oligonucleotides to generate high local concentration whereas retaining high reaction efficiency through hydrophobic-based aggregation, which is important for constructing efficient DNA nanomachines for ultrasensitive applications. MicroRNA-155, used as a model, triggered strand displacement amplification of the DNA monomers on the periphery of the 3D DNA nanomachine and generated an amplified fluorescent response for its sensitive assay. The local concentration of substrates was increased by a factor of at least 9.0×105 through hydrophobic-interaction-based self-assembly in comparison with the traditional homogeneous reaction system, achieving high local-concentration-based reactivity and free-diffusion-based enhanced reaction efficiency. As expected, the aggregation-induced synergism by hydrophobic-driven self-assembly of amphiphilic oligonucleotides created excellent properties to generate a 3D DNA nanomachine with potential as an assay for microRNA-155 in cells. Most importantly, this approach can be easily expanded for the bioassay of various biomarkers, such as nucleotides, proteins, and cells, offering a new avenue for simple and efficient applications in bioanalysis and clinical diagnosis.

A three-dimensional DNA nanomachine with target recycling amplification technology and multiple electrochemiluminescence resonance energy transfer for sensitive microRNA-141 detection

An electrochemiluminescence (ECL) three-dimensional (3D) DNA nanomachine is developed for microRNA-141 (miRNA-141) detection by coupling Pb2+ dependent DNAzyme assisted target recycling amplification technology with multiple ECL resonance energy transfer (ECL-RET) system. Firstly, Pb2+ dependent DNAzyme is formed by three single strand DNA (ssDNA): A1, A2 and the target miRNA-141. In the presence of Pb2+, the specific recognition site of the DNAzyme is cleaved and a large number of secondary targets (A3) are released. Secondly, the 3D DNA nanomachine consists of four ssDNA: H1, H2, H3 and the probe (two ends are labeled with alexa fluor (AF) and a nanocomposite (PtNCs@Ru(dcbpy)32+) which is prepared by polyethyleneimine platinum nanoclusters and tris(4,4′-dicarboxylicacid-2,2′-bipyridyl) ruthenium(II) dichloride). Then, the 3D DNA nanomachine is assembled on the gold nanoparticles modified glassy carbon electrode. Afterwards, A3 is employed to hybridize with the probe, triggering the movement of the nanomachine and forming the multiple ECL-RET system. In this system, AF, serves as an effective energy transfer donor, which can transfer energy to PtNCs and Ru(dcbpy)32+directly. Meanwhile, PtNCs, both as the acceptor and donor, can accept energy from AF and transfer it to Ru(dcbpy)32+. As a result, The biosensor achieves enhanced ECL efficiency, which is 1.78 times that of the classic tris(2,2′-bipyridyl)ruthenium(II) dichloride (Ru(bpy)32+) and exhibits good responses to miRNA-141 in the linear range from 10 aM to 100 nM with a detection limit of 3.3 aM. Also, the obtained biosensor can be employed to detect miRNA-141 in human serum samples, which will be of great significance in bioanalysis.