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1.
Anal Chem ; 2021 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-34613709

RESUMO

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.

2.
Anal Chem ; 2021 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-34609848

RESUMO

Herein, a new field-free and highly ordered spherical nucleic acid (SNA) nanostructure was self-assembled directly by ferrocene (Fc)-labeled DNA tweezers and DNA linkers based on the Watson-Crick base pairing rule, which was employed as an electrochemiluminescence (ECL) quenching switch with improved recognition efficiency due to the high local concentration of the ordered nanostructure. Moreover, with a collaborative strategy combined with the advantages of both self-accelerated approach and pore confinement-enhanced ECL effect, the mesoporous silica nanospheres (mSiO2 NSs) were prepared to be filled with rubrene (Rub) as ECL emitters and Pt nanoparticles (PtNPs) as coreaction accelerators (Rub-Pt@mSiO2 NSs), which demonstrated high ECL response in the aqueous media (dissolved O2 as coreactant). When the SNA nanostructure was immobilized on the Rub-Pt@mSiO2 NSs-modified electrode, it presented a "signal off" state owing to the quenching effect of the Fc molecules. As a proof of concept, the SNA-based ECL switch platform was applied in the detection of microRNA let-7b (let-7b). Impressively, in the presence of the target let-7b, a deconstruction of the SNA nanostructure was actuated, causing the Fc to leave the electrode surface and achieved an extremely high ECL recovery ("signal on" state). Hence, a sensitive determination for let-7b was realized with a low detection limit of 1.8 aM ranging from 10 aM to 1 nM by employing the Rub-Pt@mSiO2 NSs-based ECL platform combined with the target-triggered SNA deconstruction, which also offered an ingenious method for the further applications of biomarker analyses.

3.
ACS Appl Mater Interfaces ; 13(37): 44079-44085, 2021 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-34514796

RESUMO

Polycyclic aromatic hydrocarbons (PAHs) as traditional electrochemiluminescence (ECL) luminophores have been widely applied in the analysis field. However, their ECL intensity and efficiency are still limited due to the aggregation-induced quenching (ACQ) effect of PAHs. Hence, to overcome this limitation, we put forward a new strategy to increase the ECL intensity and efficiency by eliminating the ACQ effect of PAHs through the coordinative immobilization of PAHs within metal-organic frameworks (MOFs). As anticipated, the proof-of-concept experiment indicated that the coordinative immobilization of perylene-3,4,9,10-tetracarboxylate (PTC) into a Zn-PTC MOF could distinctly increase the ECL intensity and efficiency compared with H4PTC aggregates and H4PTC monomers. The reason for the ECL enhancement of Zn-PTC was that the immobilization of PTC within the MOF effectively amplified the distance between perylene rings of PTC ligands and thus eliminated the ACQ effect. Furthermore, the PTC into Zn-PTC was stacked in an edge-to-edge mode to form J-aggregation, which was also conducive to ECL enhancement. On the basis of the excellent ECL performance, we utilized Zn-PTC as a new ECL emitter combined with exonuclease III-stimulated target cycling and DNAzyme-assisted cycling dual amplification strategies to construct an ECL sensor for microRNA-21 detection, which had a wide signal response (100 aM to 100 pM) with a detection limit of 29.5 aM. Overall, this work represents a new and convenient method to overcome the ACQ effect of PAHs and boost the ECL performance, which opens a new horizon for developing high-performance ECL materials, thus offering more opportunities for building highly sensitive ECL biosensors.

4.
Anal Chem ; 93(36): 12400-12408, 2021 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-34469691

RESUMO

The electrochemiluminescence (ECL) ratiometric assay is usually based on two different ECL luminophores, and the choice of two suitable luminophores and shared co-reactant makes its construction challenging. The single-emitter-based ECL ratio mode could overcome the limitation of two luminophores and simplify the construction process, so it is an ideal choice. In this work, CdTe quantum dots (CdTe QDs) were modulated using black phosphorus (BP) nanosheet to simultaneously emit the cathodic and anodic ECL signals, and H2O2 and tripropylamine (TPrA) served as the cathodic and anodic co-reactants, respectively. MicroRNA-126 (miRNA-126) was selected as the template target to exploit the application of BP-CdTe QDs in the single-emitter-based ECL ratio detection. Through the target recycling triggering rolling-circle amplification (RCA) reaction, a large amount of glucose oxidase (GOx)-modified single strand 1 was introduced. GOx catalyzed glucose to produce H2O2 in situ, which acted as a dual-role moderator to quench the anodic ECL emission with TPrA as the co-reactant while enhancing the cathodic emission, thereby realizing the ratiometric detection of miRNA-126 with a low detection limit of 29 aM (S/N = 3). The dual-ECL-emitting BP-CdTe QDs with TPrA-H2O2 as dual co-reactant provide a superior ECL ratio platform involving enzyme catalytic reaction, expanding the application of single-emitter-based ratio sensing in the diverse biological analysis.


Assuntos
Técnicas Biossensoriais , Compostos de Cádmio , MicroRNAs , Pontos Quânticos , Peróxido de Hidrogênio , Fósforo , Telúrio
5.
Anal Chem ; 93(38): 12981-12986, 2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34515473

RESUMO

In this work, a DNA three-way junction (TWJ) with multiple recognition regions was intelligently engineered, which could be applied as an unconfined DNA walker with a rapid walking speed and high sensitivity for electrochemical detection of microRNA (miRNA-182-5p). Once the target miRNA was presented, the hairpins on TWJ could be successively opened to form an annular DNA walker, which could walk on the entire scope of the electrode surface without the confine for the length of DNA walker legs compared with the traditional DNA walker, greatly improving the walking efficiency. In addition, this DNA walker with multirecognition segments could obviously increase the local concentration of recognition sites, which significantly enhanced the detection speed and sensitivity. As a result, this proposed biosensor with annular DNA as a walker could dexterously achieve the ultrasensitive and fast detection of miRNA-182-5p from 0.1 fM to 1 nM with a detection limit of 31.13 aM. Meaningfully, this strategy explored an innovative path in the design of a new DNA walker nanostructure for accomplishing speedy and sensitive detection of biomarkers.


Assuntos
Técnicas Biossensoriais , MicroRNAs , DNA/genética , Técnicas Eletroquímicas , Limite de Detecção , MicroRNAs/genética
6.
Anal Chem ; 93(38): 12995-13000, 2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34524810

RESUMO

Herein, a photoelectrochemical (PEC) assay was designed for a highly sensitive DNA determination relying upon the SnO2/BiOBr p-n heterojunction as a photoactive material and SiO2 as a signal quencher. Compared with most traditional heterojunctions, the SnO2/BiOBr p-n heterostructure not only lessened the recombination of the photogenerated electron-hole pairs but also promoted the light-harvesting in the ultraviolet-visible (UV-vis) region, leading to further enhanced photoelectric conversion efficiency and photocurrent, which demonstrated 12.1-fold and 6.4-fold increments versus those of pure SnO2 and BiOBr, respectively. Additionally, the limited quantity of target DNA (a fragment of p53 gene) could be transformed into abundant output DNA-SiO2 by employing the Nt·BstNBI enzyme-assisted signal amplification procedure, leading to a highly improved detection sensitivity of the biosensor. Then, output DNA-SiO2 hybridized with the capture DNA anchored on the modified electrode surface, remarkably diminishing the PEC signal and thus achieving sensitive DNA determination. The elaborated PEC biosensor demonstrated outstanding performance within the linear range between 0.5 fM and 5 nM and a low limit of detection down to 0.18 fM, paving a new way for fabricating heterojunction with exceptional photoactive performance and demonstrating the enormous potential for detecting multitudinous biomarkers in bioanalysis and clinical therapy.


Assuntos
Técnicas Biossensoriais , Dióxido de Silício , Bismuto , DNA , Técnicas Eletroquímicas , Limite de Detecção
7.
Anal Chem ; 93(39): 13334-13341, 2021 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-34553919

RESUMO

Herein, a versatile ECL biosensor was fabricated for ultrasensitive detection of microRNA-21 (miRNA-21) from cancer cells based on a novel H2O2-free electrochemiluminescence (ECL) system (luminol/dissolved oxygen/Fe@Fe2O3 nanowires). Compared with the previously reported coreaction accelerator that needed a negative potential to produce reactive oxygen species (ROS), these newly discovered Fe@Fe2O3 nanowires could generate ROS in the detection solution immediately without the application of voltage, which narrowed down the detection potential range to avoid side reactions, favoring their practical application in biological systems. Especially, the Fe@Fe2O3 nanowires could produce H• for activating dissolved oxygen into ROS to improve the ECL intensity dramatically, which initiates a novel pathway to promote the generation of ROS for the ECL system. In addition, an original strand displacement amplification coupled with strand displacement reaction (SDA-SDR) was developed to improve the conversion efficiency of the target for sensitive detection of miRNA-21. By virtue of the SDR, a quadruple quenching effect was achieved through each output DNA strand of SDA; hence, the nucleic acid signal amplification efficiency was effectively enhanced. As expected, on account of the superb activation performance of Fe@Fe2O3 nanowires and the outstanding amplification efficiency of the SDR-SDA strategy, the fabricated ECL biosensor realized ultrasensitive detection of miRNA-21 with a detection limit down to 52.5 aM. The established ECL sensing platform ushered a new route for H2O2-free detection and a promising biomarker assay method for clinical diagnosis.


Assuntos
MicroRNAs , Nanofios , Luminol , Oxigênio
8.
Biosens Bioelectron ; 195: 113662, 2021 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-34571484

RESUMO

Herein, the sequence-specific short-stranded biomarker DNA (hDNA, 21-nt) is acted as targeting out-primer to implement the loop-mediated isothermal amplification for releasing hydrogen ions (LAMP-H+). Using LAMP-H+ as signaling transducer, we report a highly sensitive electrochemical ratiometric biosensor for hDNA with minimized background signal, which is achieved via magnetic separation using AuNPs-modified Fe3O4 (Au@Fe3O4) as micro-reactor. In Au@Fe3O4, a double-stranded complex of a pH-responsible strand (I*) and a substrate strand (S*) is bound via Au-N bonds, where the treatment with LAMP-H+ leads to I* folding into i-motif conformation and S* dehybridization. The S* further hybridizes a catalytic strand (C*) to assemble Mg2+-DNAzymes that are cleaved by Mg2+, releasing C* for repeated formation and robust nicking of Mg2+-DNAzymes. The resultant output fuel strands (F*) are introduced in a modified electrode to drive the strand displacement of two hairpins individually labeled with two electron mediators. Through F*-mediated recycled amplification, the ratio of their electrochemical currents changed in opposite is highly sensitive to the varied hDNA down to 2.1 fM. By integrating LAMP-H+-stimulated i-motif switching with Mg2+-DNAzyme cleavage, this logic transduction of LAMP-H+(i-motif/Mg2+-DNAzyme)F* efficiently minimizes the inherent background of traditional LAMP-based assays. Resultantly, our electrochemical ratiometric strategy would be applicable to diverse short-stranded DNAs or even RNAs as targeting primers of LAMP.

9.
Anal Chem ; 93(33): 11634-11640, 2021 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-34378382

RESUMO

Exploring the ratiometric fluorescence biosensing of DNA-templated biemissive silver nanoclusters (AgNCs) is significant in bioanalysis, yet the design of a stimuli-responsive DNA device is a challenge. Herein, using the anti-digoxin antibody (anti-Dig) with two identical binding sites as a model, a tweezer-like DNA architecture is assembled to populate fluorescent green- and red-AgNCs (g-AgNCs and r-AgNCs), aiming to produce a ratio signal via specific recognition of anti-Dig with two haptens (DigH). To this end, four DNA probes are programmed, including a reporter strand (RS) dually ended with a g-/r-AgNC template sequence, an enhancer strand (ES) tethering two same G-rich tails (G18), a capture strand (CS) labeled with DigH at two ends, and a help strand (HS). Initially, both g-AgNCs and r-AgNCs wrapped in the intact RS are nonfluorescent, whereas the base pairing between RS, ES, CS, and HS resulted in the construction of DNA mechanical tweezers with two symmetric arms hinged by a rigid "fulcrum", in which g-AgNCs are lighted up due to G18 proximity ("green-on"), and r-AgNCs away from G18 are still dark ("red-off"). When two DigHs in proximity recognize and bind anti-Dig, the conformation switch of these tweezers resultantly occurs, taking g-AgNCs away from G18 for "green-off" and bringing r-AgNCs close to G18 for "red-on". As such, the ratiometric fluorescence of r-AgNCs versus g-AgNCs is generated in response to anti-Dig, achieving reliable quantization with a limit of detection at the picomolar level. Based on the fast stimulated switch of unique DNA tweezers, our ratiometric strategy of dual-emitting AgNCs would provide a new avenue for a variety of bioassays.


Assuntos
Técnicas Biossensoriais , Nanopartículas Metálicas , Anticorpos , DNA , Fluorescência , Prata , Espectrometria de Fluorescência
10.
Anal Chem ; 93(35): 12075-12080, 2021 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-34427443

RESUMO

Herein, with skillfully engaging stable T-Hg2+-T bonding, a "Crab Claw"-like DNA nanomachine with concise and highly efficient assembly and enhanced recognition/conversion efficiency was engineered as a super signal amplifier, which was united with Pd@Cu@Pt multimetallic mesoporous nanomaterials (Pd@Cu@Pt MMNs) for ultrasensitive electrochemical assay of mercury ions (Hg2+). Specifically, the formed "Crab Claw"-like DNA nanomachine could simultaneously trigger four same cascade DNAzyme cleavage reactions with the help of Mg2+ DNAzyme for markedly converting target Hg2+ to enormous DNA segments labeled with ferrocene (Fc), improving the detection sensitivity. Subsequently, the prepared Pd@Cu@Pt MMNs could not only show commendable electrochemical catalysis to Fc but also act as an excellent immobilization matrix for capturing and accumulating abundant Fc around them to further strengthen the electrochemical signal. As a result, the well-designed electrochemical sensor could achieve a low limit of detection of 3.58 fM in the range from 10 fM to 100 nM for Hg2+detection. This strategy offers a simple and rapid avenue to detect heavy metal ions and shows promising application potential for environmental pollutant monitoring.


Assuntos
Técnicas Biossensoriais , DNA/química , Mercúrio , Nanotecnologia , Técnicas Eletroquímicas , Limite de Detecção , Mercúrio/análise
11.
Talanta ; 234: 122643, 2021 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-34364452

RESUMO

Herein, an amplified and renewable electrochemical biosensor was developed via bienzymatic cascade catalysis of glucose oxidase (GOx) and horseradish peroxidase (HRP), which were confined in a functional Y-shaped DNA nanostructure oriented by a dual-thiol-ended hairpin probe (dSH-HP) with a paired stem as a rigid scaffold and unpaired loop as enclosed binding platform. For proof-of-concept assay of sequence-specific biomarker DNA related to Alzheimer's disease (aDNA), GOx and redox ferrocene-modified HRP (Fc@HRP) were chemically conjugated in two enzyme strands (GOx-ES1 and Fc@HRP-ES2), respectively. The repeated recycling of aDNA was powered by the displacement of GOx-ES1 by aDNA and exonuclease III (ExoIII)-assisted cleavage reaction for amplified output of numerous GOx-ES1 as dependent transducers, together with Fc@HRP-ES2 which was simultaneously hybridized with dSH-HP to assemble this DNA structure. Rationally, the bienzymatic cascade catalysis was motivated through GOx-catalyzed glucose oxidization to in situ generate hydrogen peroxide (H2O2) and overlapped HRP-catalyzed H2O2 decomposition to promote the electron transfer, producing significantly enhanced electrochemical signal of Fc with an ultrahigh sensitivity down to 0.22 fM of aDNA. Benefited from the unique design of dSH-HP-oriented bienzymatic cascades, this one-step strategy without non-specific blockers passivation was simple and renewable, and would pave a promising avenue for sensitive electrochemical assay of biomolecules.


Assuntos
Técnicas Biossensoriais , Técnicas Eletroquímicas , Catálise , DNA , Glucose Oxidase , Peroxidase do Rábano Silvestre , Peróxido de Hidrogênio
12.
Anal Chem ; 93(31): 10890-10897, 2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34313108

RESUMO

Organic materials with diverse structures and brilliant glowing colors have been attracting extensive attention in optical electronic devices and electrochemiluminescence (ECL) fields and are currently faced with the issue of low ECL efficiency. Herein, a series of tetraphenyl alkene nanocrystals (TPA NCs) with an ordered molecular structure were synthesized to explore regularities in the crystallization-induced enhanced (CIE) ECL emission effects by altering the number and position of vinyl on the backbone of TPA molecules. Among those TPA NCs, tetraphenyl-1,3-butadiene (TPB) NCs exhibit the brightest ECL emission via a coreactant pathway, with the relative ECL efficiency of up to 31.53% versus the standard [Ru(bpy)3]2+/TEA system, which is thousands of times higher than that of free TPB molecules. The high ECL efficiency of TPB NCs originates from the effective electron transfer of unique J-aggregates on the a axis of the nanocrystals to notably promote radiative transition and the restriction on the free rotation of TPB molecules to further suppress the nonradiative transition, which has exhibited great potential in ultrasensitive biosensing, efficient light-emitting devices, and clear ECL imaging fields. As a proof of concept, since dopamine (DA) can form benzoquinone species by electrochemical oxidation to realize intermediate radical quenching and excited-state quenching on the TPB NCs/TEA system, the TPB NCs with the CIE ECL effect are used to construct an ultrasensitive ECL-sensing platform for the determination of DA with a lower detection limit of 3.1 nM.


Assuntos
Técnicas Biossensoriais , Nanopartículas , Alcenos , Cristalização , Técnicas Eletroquímicas , Medições Luminescentes
13.
Anal Chem ; 93(31): 11019-11024, 2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34324804

RESUMO

Au nanoparticles (Au NPs) can be self-assembled in a bottom-up orderly manner at the oil-water interface, which is widely used as SERS platforms, but the stability of the Au NP interface needs to be improved due to shaking or shifting and the Brownian motion. The DNA structure with unique sequence specificity, excellent programmability, and flexible end-group modification capability owns good potential to precisely control the plasmonic structure's distance. In this study, a large area of the SERS substrate is obtained from the DNA structure-stabilized self-assembled ordered Au NPs on the cyclohexane-water interface. Combining with the exonuclease III (exo III)-assisted DNA recycling amplification strategy, we construct a liquid-phase SERS biosensor for efficient detection of microRNA 155 (miRNA 155). Compared with the traditional randomly assembled Au NPs on the two-phase interface, the SERS signal is significantly enhanced and more stable. The detection limit of the SERS biosensor for miRNA 155 reached 1.45 fmol/L, which has a very wide linear range (100 fmol/L-5 nmol/L). This work gives an efficient approach to stabilize the self-assembly Au NPs on the liquid-liquid interface, which can broaden the application of SERS analysis.


Assuntos
Nanopartículas Metálicas , MicroRNAs , DNA , Ouro , Análise Espectral Raman
14.
ACS Appl Mater Interfaces ; 13(27): 32013-32021, 2021 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-34212714

RESUMO

The reported donor donor-acceptor ("DD-A") fluorescence resonance energy transfer (FRET) was typically achieved through random collisions and interactions of DNA molecules in the bulk solution, which has inevitable defects, including weak biological stability, slow reaction kinetics, and low hybridization efficiency. In order to overcome these deficiencies, this work developed a quadrivalent cruciform DNA nanostructure (qCDN)-mediated cascaded catalyzed hairpin assembly (CHA) amplifier for the fluorescence detection of amyloid ß oligomer species (AßOs). First, four H1 and four H2 hairpins were assembled on one qCDN to obtain qCDNH1 and qCDNH2, respectively. In the presence of AßOs, strand C was released from the P1-C hybrid hairpin and then alternately opened qCDNH1 and qCDNH2 to trigger the qCDN-mediated CHA. As a result, double donors in H1 and one acceptor in H2 were mutually closed, and the porous DNA nanonet with a high loading of "DD-A" FRET binary probes was formed. The FRET efficiency was approximately 78%, and the initial reaction rate was 25-fold faster than the conventional CHA. The detection limit of AßOs was as low as 0.69 pM. The combination of the "DD-A" FRET binary probes and qCDN-mediated cascaded amplifier exhibited great promise for detecting biomarkers with trace levels.


Assuntos
Peptídeos beta-Amiloides/química , DNA Cruciforme/química , Transferência Ressonante de Energia de Fluorescência/métodos , Limite de Detecção , Nanoestruturas/química , Multimerização Proteica , Estrutura Quaternária de Proteína
15.
Anal Chem ; 93(27): 9568-9574, 2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34210120

RESUMO

In this work, an original rolling-circle strand displacement amplification (RC-SDA) was developed by introducing a circle DNA with two recognition domains as a template instead of the limited liner DNA template in traditional strand displacement amplification (SDA), which displayed much shorter reaction time down to 30 min and quite higher conversion efficiency of more than 1.77 × 108 compared with those of traditional strand displacement amplification (SDA) and could be applied to construct a label-free biosensor for ultrasensitive detection of an HIV DNA fragment. Once the target HIV DNA fragment interacts with the template circle DNA, the RC-SDA could be activated to dramatically output amounts of mimic target DNA with the assistance of the Phi29 DNA polymerase and Nb.BbvCI enzyme. In application, while the output products were captured by the DNA tetrahedral nanoprobe (DTNP) modified electrode, the electrochemical tag silver nanoclusters (AgNCs) on DTNP would be released from the electrode surface, accompanied with an obviously decreased electrochemical signal. This way, the developed signal-off biosensor was successfully applied to realize the rapid and ultrasensitive detection of target HIV DNA fragment with a detection limit down to 0.21 fM, which exploits the new generation of a universal strategy beyond the traditional ones for applications in biosensing assay, clinic diagnosis, and DNA nanobiotechnology.


Assuntos
Técnicas Biossensoriais , Técnicas Eletroquímicas , DNA/genética , Limite de Detecção , Técnicas de Amplificação de Ácido Nucleico , Prata
16.
Anal Chem ; 93(28): 9912-9919, 2021 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-34232629

RESUMO

The simultaneous live-cell imaging of multiple intracellular and disease-related microRNAs (miRNAs) with low abundances is highly important to enhance specificity and accuracy for disease diagnosis. On the basis of the improved cell internalization and accelerated reaction kinetics, we develop a three-dimensional (3D) DNA nanoprobe that integrates intramolecular DNAzyme (intra-Dz) and catalytic hairpin assembly (intra-CHA) amplifications to simultaneously monitor multiple miRNAs in living cells. The sensing components are loaded on a DNA scaffold via the sticky-end hybridization of the DNA sequences to increase the local concentrations of the signal probes. The miRNA-21 and miRNA-155 target sequences can trigger intra-Dz and -CHA amplifications on the nanoprobes to show significantly amplified and distinct fluorescence at different wavelengths for simultaneously monitoring low levels of miRNAs. Real-time fluorescence microscopy reveals that such a 3D DNA nanoprobe design with the intra-Dz and -CHA amplifications can accelerate the reaction rate compared to that of the conventional free Dz and CHA because of the increased local concentrations of the sensing components. Importantly, the 3D DNA nanoprobe has desirable stability and biocompatibility and can be readily delivered into living cells to achieve multiplexed and highly sensitive sensing of intracellular miRNA-155 and miRNA-21 sequences. With the demonstration of its intracellular application, the developed 3D DNA nanoprobe thus holds promising potential for biological studies and accurate disease diagnosis.


Assuntos
Técnicas Biossensoriais , DNA Catalítico , MicroRNAs , DNA/genética , DNA Catalítico/genética , MicroRNAs/genética , Técnicas de Amplificação de Ácido Nucleico , Hibridização de Ácido Nucleico
17.
Anal Chem ; 93(29): 10212-10219, 2021 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-34251187

RESUMO

Herein, a novel Au nanoclusters/Cu2O (Au NCs/Cu2O) heterostructure exhibited exceptionally strong electrochemiluminescence (ECL) emission, in which the p-type semiconductor Cu2O was defined as the electrosensitizer to provide the electrogenerated holes for rapidly transferring the electrogenerated hot electrons of Au NCs. Thus, the fast charge transfer of Au NCs/Cu2O was achieved by the electrosensitizer compared to the sluggish one via intramolecular covalent bond charge transfer of traditional Au NCs, resulting in a greatly higher ECL efficiency (63.8%) than that of pure Au NCs (2.7%) versus the standard [Ru(bpy)3]2+. It solved one main challenge of electrochemiluminophore-based metal NCs: high efficiency with energic charge-transport kinetics. As a proof of concept, Au NCs/Cu2O was successfully employed in an ultrasensitive ECL biosensing platform for determining the biological antioxidant glutathione with a limit of detection (LOD) as low as 6.3 pM. The heterostructure as an ECL emitter is a very promising start for guiding the rational design of efficient electrochemiluminophores in intense light-emitting devices and high-definition ECL imaging.


Assuntos
Técnicas Biossensoriais , Nanopartículas Metálicas , Técnicas Eletroquímicas , Ouro , Limite de Detecção , Medições Luminescentes
18.
Analyst ; 146(16): 5067-5073, 2021 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-34297024

RESUMO

Designing antibody-powered DNA nanodevice switches is crucial and fascinating to perform a variety of functions in response to specific antibodies as regulatory inputs, achieving highly sensitive detection by integration with simple amplified methods. In this work, we report a unique DNA-based conformational switch, powered by a targeted anti-digoxin mouse monoclonal antibody (anti-Dig) as a model, to rationally initiate the hybridization chain reaction (HCR) for enzyme-free signal amplification. As a proof-of-concept, both a fluorophore Cy3-labeled reporter hairpin (RH) in the 3' terminus and a single-stranded helper DNA (HS) were individually hybridized with a recognition single-stranded DNA (RS) modified with Dig hapten, while the unpaired loop of RH was hybridized with the exposed 3'-toehold of HS, isothermally self-assembling an intermediate metastable DNA structure. The introduction of target anti-Dig drove the concurrent conjugation with two tethered Dig haptens, powering the directional switch of this DNA structure into a stable conformation. In this case, the unlocked 3'-stem of RH was implemented to unfold the 5'-stem of the BHQ-2-labeled quench hairpin (QH), rationally initiating the HCR between them by the overlapping complementary hybridization. As a result, numerous pairs of Cy3 and BHQ-2 in the formed long double helix were located in spatial proximity. In response to this, the significant quenching of the fluorescence intensity of Cy3 by BHQ-2 was dependent on the variable concentration of anti-Dig, achieving a highly sensitive quantification down to the picomolar level based on a simplified protocol integrated with enzyme-free amplification.


Assuntos
Técnicas Biossensoriais , DNA , Animais , DNA/genética , DNA de Cadeia Simples/genética , Corantes Fluorescentes , Imunoensaio , Limite de Detecção , Camundongos , Hibridização de Ácido Nucleico
19.
Chem Commun (Camb) ; 57(52): 6416-6419, 2021 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-34095911

RESUMO

Herein, based on iron oxyhydroxide (FeOOH) as the photoactive material and exonuclease III (exo III)-aided dual recycling signal amplification, a new photoelectrochemical (PEC) biosensor was successfully developed for human papillomavirus-16 (HPV-16) detection with a wide linear range from 0.5 fM to 1 nM and a low detection limit of 0.17 fM.


Assuntos
Técnicas Biossensoriais/métodos , Exodesoxirribonucleases/metabolismo , Compostos Férricos/química , Papillomavirus Humano 16/isolamento & purificação , DNA de Cadeia Simples/química , DNA de Cadeia Simples/metabolismo , Técnicas Eletroquímicas , Eletrodos , Papillomavirus Humano 16/genética , Humanos , Luz , Limite de Detecção , Técnicas de Amplificação de Ácido Nucleico , Reprodutibilidade dos Testes
20.
Anal Chem ; 2021 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-34133127

RESUMO

The bifunctional moderator is urgently needed in the field of ratiometric electrochemiluminescence (ECL) sensing since it can mediate simultaneously two ECL signals to conveniently realize their opposite change trend. This work designed a novel dual-signal combined nanoprobe with carboxyl-functionalized poly[(9,9-dioctylfluorenyl-2,7-diyl)-co-(1,4-benzo-{2,1',3}-thiadazole)] nanoparticles (c-PFBT NPs) as the anodic ECL probe and L-cysteine capped CdS quantum dots (L-CdS QDs) as the cathodic ECL probe, which performed a dual-signal output capability without any additional coreactants. More importantly, hydrogen peroxide (H2O2) produced in situ by enzyme-catalyzed reaction was developed as a bifunctional moderator for simultaneously regulating two signals. The dual-signal combined nanoprobe (c-PFBT NPs@CdS QDs) served as the matrix to immobilize acetylcholinesterase (AChE) and choline oxidase for organophosphorus (OPs) analysis. In the absence of OPs, H2O2 was produced by catalyzing the substrate acetylthiocholine (ATCl) with enzymes and it quenched the anodic ECL signal from c-PFBT NPs and simultaneously promoted the cathodic ECL signal from L-CdS QDs. When OPs was present, the activity of AChE was inhibited, the anodic signal would increase, and the cathodic signal would accordingly decrease. The integration of the bifunctional moderator H2O2 and dual-signal combined nanoprobe c-PFBT NPs@CdS QDs not only provides an attractive ECL platform for enzymatic sensing involving the generation or consumption of H2O2 but also paves a new pathway for other ratiometric ECL systems involving enzyme catalytic amplification for detecting antigens, antibodies, DNA, RNA, etc.

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