ACS Sensors
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2025
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IF 9.1
Yapeng Wu,Bei Lv,Xintian Ni,Sheng Zhu,Dawei Li
Strand displacement amplification (SDA) is an isothermal DNA amplification technique. Herein, we developed a novel SDA system, designated All-U-Want SDA (AUW-SDA), which was used as a signal amplification strategy for the construction of nucleic acid detection biosensors. AUW-SDA is capable of target turnover and can be utilized for detection of nucleic acid sequences without available 3′-ends. Of particular significance is the ability of AUW-SDA to generate a substantial number of programmable sequences in accordance with the specifications of the sensor signal output methods, irrespective of the sequence of the target nucleic acid. We used the N gene of SARS-CoV-2 as a model target to develop a sensing platform with dual signal outputs. The colorimetric signals were generated by the G-quadruplex/hemin DNAzyme, in which the G-rich sequences were produced by AUW-SDA with a C-rich primer. On the other hand, by altering the sequence within the replaceable region of the primer, an activator sequence was obtained from AUW-SDA, which could trigger the activity of CRISPR/Cas12a, cleaving the probes modified with a fluorophore and quencher at each end and subsequently yielding the fluorescent signals. After the DNA sequences and reaction conditions were optimized, the limit of detection (LOD) values of the fluorescent and colorimetric assays were estimated to be 0.672 fM and 13.3 fM, respectively. The biosensors were utilized for biological sample detection. The reliability of the proposed method was validated against RT-qPCR results. In addition, a portable scanner-assisted high-throughput RGB analysis (PSHRA) method was developed. This method was applied to our biosensor for multilocus detection of SARS-CoV-2. The results obtained were satisfactory, indicating the potential of this approach for field testing or point-of-care (POC) diagnostics.
研究领域:
生物化学
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分子生物学
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诊断学
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DOI
PubMed
高影响因子
ANALYTICAL CHEMISTRY
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2022
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IF 8.008
Yun Chen,Dehui Qiu,Xiaobo Zhang,Yuan Liu,Mingpan Cheng,Jianping Lei,Jean-Louis Mergny,Huangxian Ju,Jun Zhou
G-quadruplex/hemin (G4/hemin) DNAzymes are biosensing systems, but their application remains limited by an overall low activity and a rather high level of unwarranted background reactions. Here, these issues were addressed through the rational design of F3T-azaC-hemin, a G4-based construct in which the hemin is covalently linked to the G4 core and its binding site flanked with a nucleotide activator, here d(T-azaC). This design led to a G4-DNAzyme whose performances have been ca. 150-fold increased compared to the parent G4-based system. The utility of F3T-azaC-hemin was demonstrated here through the ultrasensitive chemiluminescent detection of miRNA-221. The limit of detection (LOD) has been decreased to the femtomolar range, making it a new and highly efficient molecular tool in the biosensing technology field.
研究领域:
生物传感技术
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分子生物学
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分析化学
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DOI
PubMed
高影响因子
MICROCHIMICA ACTA
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2025
·Vol. 192
·期 6
·1-12
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IF 5.3
Chen Yafei,Zhang Xuhui,Huang Wanjin,Jin Zhenhuan,Wei Xiaoping,Li Jianping
Atrazine (ATZ) is used extensively, resulting in residues in food and the environment, posing a serious threat to human health. Herein, Cd 0.5 Zn 0.5 S/Ti 3 C 2 photoelectric material was synthesized and immobilized on a FTO electrode as a photoanode. A photoelectrochemical (PEC) aptamer sensor was constructed for the highly sensitive and selective determination of ATZ based on signal amplification via cascade catalysis of CRISPR/Cas12a and G-quadruplex/hemin DNAzyme (G4/hemin DNAzyme). G4/hemin DNAzyme catalyses the oxidation reaction between H 2 O 2 and dopamine (DA) to form polydopamine (PDA) deposit. This process, in turn, inhibits the photocurrent at the photoanode, leading to a decrease in photocurrent. Concurrently, the depletion of DA as an electron donor for the PEC reaction at the photoelectrode further contributes to the decrease in photocurrent. ATZ can hybridize with ATZ aptamer (Apt) in Apt/cDNA to release activation strand (cDNA), which activates the activity of CRISPR/Cas12a and triggers cleavage of G4, causing the cleaving of G4/hemin DNAzyme immobilized on the electrode surface. This process leads to a decrease of G4/hemin DNAzymes amount on the electrode, consequently reducing both the PDA generation and the DA consumption. As a result, the photocurrent is restored. The cascade catalysis of CRISPR/Cas12a and G4/hemin DNAzyme has been demonstrated to result in photocurrent amplification. The photocurrent change was linear with the logarithmic value of ATZ concentration in the range 1.00 × 10 –12 to 1.00 × 10 –5 mol/L. The limit of detection was 3.47 × 10 –13 mol/L. The sensor has been successfully applied to the determination of trace ATZ in environmental and food samples. Graphical abstract
研究领域:
环境科学
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分析化学
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分子生物学
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DOI
PubMed
高影响因子
ANALYTICA CHIMICA ACTA
·
2024
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IF 5.7
Mengmeng Wu,Bing Yang,Lu Shi,Qiaorong Tang,Jing Wang,Wei Liu,Baoxin Li,Yan Jin
Background The human immunodeficiency virus (HIV) remains a major worldwide health problem. Nowadays, many methods have been developed for quantitative detecting human immunodeficiency virus DNA (HIV-DNA), such as fluorescence and colorimetry. However, these methods still have the disadvantages of being expensive and requiring professional technicians. Early diagnosis of pathogens is increasingly dependent on portable instruments and simple point-of-care testing (POCT). Therefore, it is meaningful and necessary to develop portable and cheap methods for detecting disease markers. Results In this work, a label-free chemiluminescence (CL) method was developed for detecting HIV-DNA via a handheld luminometer. To achieve label-free target detection, the CL catalyst, G-triplex-hemin DNAzyme (G3-hemin DNAzyme), was in-situ assembled in the presence of HIV-DNA. For improving sensitivity, HIV-DNA induced the cyclic strand displacement reaction (SDR), which can form three G3-hemin DNAzymes in one cycle. So, the chemiluminescence reaction between luminol and H 2 O 2 was highly effectively catalyzed, and the CL intensity was linearly related with the concentration of HIV-DNA in the range of 0.05–10 nM with a detection limit of 29.0 pM. Due to the high specificity of hairpin DNA, single-base mismatch can be discriminated, which ensured the specific detection of HIV-DNA. Significance In-situ formation of G3-hemin DNAzyme led to label-free and selective detection without complex synthesis and functionalization. Therefore, it offers a cheap, selective, sensitive and portable method for detecting disease-related genes, which is promising for POCT of clinical diagnosis in resource-limited settings.
研究领域:
生物医学
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分子诊断
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临床检验
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DOI
PubMed
高影响因子
ANALYTICA CHIMICA ACTA
·
2021
·
IF 6.558
Guoyan Zhao,Yongmei Liu,Jie Du,Huizi Zhang,Hanqing Feng,Xiaoquan Lu
Extracellular ATP (eATP) is an important biological signal transduction molecule. Although a variety of detection methods have been extensively used in ATP sensing and analysis, accurate detection of eATP remains difficult due to its extremely low concentration and spatiotemporal distribution. Here, an eATP measurement strategy based on tetrahedral DNA (T-DNA)-modified electrode sensing platform and hybridization chain reaction (HCR) combined with G-quadruplex/Hemin (G4/Hemin) DNAzyme dual signal amplification is proposed. In this strategy, ATP aptamer and RNA-cleaving DNAzyme were combined to form a split aptazyme . In the presence of ATP, this aptazyme hydrolyzes the cleaving substrate strand with high selectivity , releasing cleaved ssDNA , which are captured by the T-DNA assembled on the electrode surface, triggering an HCR on the electrode surface to form numerous linker sequences of the HCR dsDNA product. When [email protected] (G4) spherical nucleic acid enzymes (SNAzymes) with other linkers are used as nanocatalyst tags, they are captured by HCR dsDNA through sticky linkers present on the electrode surface. An amplified electrochemical redox current signal is generated through SNAzyme-mediated catalysis of H 2 O 2 , enabling easy detection of picomole amounts of ATP. Using this strategy, eATP levels released by tobacco suspension cells were accurately measured and the distribution and concentration of eATP released on the surface of an Arabidopsis leaf was determined.
研究领域:
植物学
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分子生物学
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DOI
PubMed
高影响因子
JOURNAL OF FOOD COMPOSITION AND ANALYSIS
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2025
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IF 4.6
Dian Yu,Yingying Sun,Xin Lin,Wenjie Liu,Jianna Yu,Guoxing Jing,Wen Liu,Xiaoyun Zhang,Cheng Jiang,Wenshan Li
G-quadruplex/hemin DNAzyme (G-DNAzyme) is widely utilized in biosensors due to its ability to catalyze peroxidase-like reactions. However, its relatively low catalytic efficiency can limit detection sensitivity. To address this limitation and enable trace detection, this study presents a G-DNAzyme-mediated biosensor that combines entropy-driven catalytic reactions (EDC) and catalytic hairpin assembly (CHA) to enhance sensitivity and broaden its applicability for detecting trace levels of aflatoxin B1 (AFB1). In this biosensor, AFB1 specifically binds to aptamer, triggering the release of a complementary initiator chain, which activates both EDC and CHA amplification processes. This dual amplification results in the generation of numerous free G-quadruplex sequences, which then bind to hemin, catalyzing a color change in the chromogenic substrate and significantly amplifying the signal. The biosensor demonstrated excellent linearity between the absorbance intensity and logarithm of AFB1 concentrations, with a detection limit (LOD) of 0.47 pM. Furthermore, spiked recovery tests of AFB1 in actual samples showed recovery rates between 96.12 % and 108.50 %, indicating strong accuracy. The method is simple, rapid, and does not require complex instrumentation, making it an efficient tool for detecting AFB1 in food safety applications.
研究领域:
生物化学
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食品安全检测
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分子生物学
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产品关联
DOI
高影响因子
Frontiers in Chemistry
·
2025
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IF 4.2
Ting Yuan,Jianbo Yuan,Jian Huang,Nana Li,Huan Cai,Yan Yang,Juan Li,Rui Chen,Xun Min
Introduction: The rapid and accurate identification of Mycobacterium tuberculosis (MTB) is essential for effective tuberculosis (TB) control. However, conventional diagnostic methods for MTB suffer from limitations such as low sensitivity, poor specificity, high cost, reliance on specialized instruments, and complex, time-consuming procedures. To address these challenges, there is an urgent need for a simple, rapid, and highly sensitive detection method that can be deployed in point-of-care settings.Methods: We developed a one-pot biosensing system combining recombinase polymerase amplification (RPA) and CRISPR/Cas12a-driven G4-hemin self-assembly for the colorimetric detection of MTB. Glycerol was employed as a phase-separation barrier to prevent interference between RPA amplification and CRISPR/Cas12a trans-cleavage. A single-stranded DNA (ssDNA) probe, designed to self-assemble with ssDNA-hemin into G4-hemin nanozymes upon CRISPR/Cas12a-mediated cleavage, served as the reaction substrate. The ssDNA-hemin further enhanced the catalytic activity of the generated G4-hemin DNAzyme. The entire assay was completed in a single step within 60 min without requiring complex instrumentation.Results and Discussion: Under optimized conditions, the biosensing system achieved ultrasensitive naked-eye detection of MTB with a limit of detection (LOD) of 10 copies/μL, comparable to traditional four-step fluorescent assays. Clinical validation using 104 patient samples demonstrated high concordance with standard diagnostic methods. This approach combines the advantages of recombinase polymerase amplification (RPA), CRISPR/Cas12a specificity, and G4-hemin DNAzyme-based colorimetric signal amplification, enabling simple, equipment-free visual detection. Given its speed, sensitivity, and ease of use, this biosensing system holds significant promise for point-of-care MTB nucleic acid testing in resource-limited settings.
研究领域:
生物传感器
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分子诊断
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CRISPR技术
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核酸扩增
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DNA酶技术
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结核病检测
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产品关联
DOI
PubMed
高影响因子
TALANTA
·
2024
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IF 5.6
Changjiang Li,Yuqiang Hu,Tianzi Shi,Kejun Dong,Tongbo Wu
The expression level of human apurinic/apyrimidinic endonuclease 1 (APE1) is closely associated with the onset of various diseases, establishing it as a crucial clinical biomarker and a target in anti-cancer efforts. This study accomplished colorimetric and visual detection of APE1 by harnessing its endonuclease activity through catalytic hairpin self-assembly (CHA) and G-quadruplex/hemin DNAzyme. Optimization of the freedom degrees of the G-rich sequence significantly improved the detection performance of the strategy by influencing DNAzyme formation. Additionally, we replaced the signal reporting system with a molecular beacon to develop a fluorescence detection strategy, which served as an extension of the signal amplification system for validation and signal readout. The fluorescent probe method achieved a detection limit of 3.37 × 10 −4 U/mL, while the colorimetric method yielded a detection limit of 6.5 × 10 −3 U/mL, with a linear range spanning from 0.01 to 0.25 U/mL. Subsequently, the colorimetric approach effectively assessed APE1 activity in biological samples and facilitated the screening of APE1 activity inhibitors. Furthermore, this CHA/G-quadruplex/hemin DNAzyme strategy was adapted for the colorimetric detection of adenosine, showcasing its broad applicability across various biomarkers. The developed colorimetric analytical strategy represents a pivotal biosensing platform for diagnosing and treating diseases.
研究领域:
生物技术
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分子生物学
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分析化学
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癌症研究
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诊断学
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产品关联
DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2025
·
IF 10.5
Hang Ao,Jinshan Xiong,Wencheng Xiao,Wenrui Hu,Jie Wu,Huangxian Ju
Owing to the continuous mutations of the coronavirus, the sensitive and accurate detection of the SARS-CoV-2 nucleocapsid protein is still necessary in clinical diagnosis. Aim to the challenge of sensitivity in chemiluminescence (CL) imaging detection and the need for high-throughput clinical screening, this work used imidazole (Im) as an enhancer for hemin-DNA/luminol/H 2 O 2 CL system to propose a CL imaging method for SARS-CoV-2 nucleocapsid protein detection. In this system Im could reduce the hemin-DNA catalyzed oxidation energy of luminol by H 2 O 2 to promote the CL emission through the accelerated breaking of O-O bond of dianionic cyclic peroxide. The enhancement mechanism was demonstrated with theoretical calculations and several characterization techniques. By combining the proximity-induced initiator release with immunological recognition, an initiator-triggered DNA nanomachine was designed to achieve the release of abundant primers for activating hemin-DNA switch. This signal amplification strategy led to strong CL emission for highly sensitive imaging detection of the target. The Im-enhanced hemin-DNA/luminol/H 2 O 2 CL imaging assay exhibited a linear detection range over 4 orders of magnitude with a detection limit down to 2.75 pg/mL for SARS-CoV-2 nucleocapsid protein, exhibiting promising potential of both Im sensitization and the proposed DNA nanomachine to improve the sensitivity of high-throughput CL biosensing for the early diagnosis and clinical screening of the coronavirus and other diseases.
研究领域:
生物分析化学
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生物传感器
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临床诊断
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病毒学
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纳米生物技术
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产品关联
DOI
PubMed
高影响因子
Advanced Healthcare Materials
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2024
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IF 10
Zhenqiang Wang,Xiyue Xie,Kaifei Jin,Daqing Xia,Jing Zhu,Jixi Zhang
In situ staining of protein dimerization on cell membrane has an important significance in accurate diagnosis during perioperative period, yet facile integration of specific recognition function and local signal conversion/amplification abilities on membrane surface remains a great challenge. Herein, a two-stage catalytic strategy is developed by installing DNA nanomachines and employing. Specifically, dual-aptamer-assisted DNA scaffold perform a “bispecific recognition-then-computing” operation and the output signal initiate a membrane-anchored biocatalysis for self-assembly of DNA catalytic converters, that is, G-quadruplex nanowire/hemin DNAzyme. Then, localized-deposition of chromogenic polydopamine is chemically catalyzed by horseradish peroxidase-mimicking DNAzyme and guided by supramolecular interactions between conjugate rigid plane of G-tetrad and polydopamine oligomer. The catalytic products exhibit nanofiber morphology with a diameter of 80–120 nm and a length of 1–10 µm, and one-to-one localize on DNA scaffold for amplified and specific staining of protein dimers. The bispecific staining leads to a higher (≈3.4-fold) signal intensity than traditional immunohistochemistry, which is beneficial for direct visualization. Moreover, an efficient discrimination ability of the bispecific staining strategy is observed in co-culture model staining. This study provides a novel catalytic method for controlling deposition of chromogens and paves a new avenue to sensitively stain of protein-protein interactions in disease diagnosis.
研究领域:
生物化学
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纳米技术
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癌症生物学
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分子生物学
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生物医学工程
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产品关联
DOI
PubMed
高影响因子
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