JOURNAL OF MICROBIOLOGY AND BIOTECHNOLOGY
·
2024
·
IF 2.5
Lu Huang,Ye Zhang,Jie Liu,Dalin Zhang,Li Li
This study presents a fluorescent mechanism for two-step amplification by combining two widely used techniques, exponential amplification reaction (EXPAR) and catalytic hairpin assembly (CHA). Pseudomonas aeruginosa ( P. aeruginosa ) engaged in competition with the complementary DNA in order to attach to the aptamer that had been fixed on the magnetic beads. The unbound complementary strand in the liquid above was utilized as a trigger sequence to initiate the protective-EXPAR (p-EXPAR) process, resulting in the generation of a substantial quantity of short single-stranded DNA (ssDNA). The amplified ssDNA can initiate the second CHA amplification process, resulting in the generation of many double-stranded DNA (dsDNA) products. The CHA reaction was initiated by the target/trigger DNA, resulting in the release of G-quadruplex sequences. These sequences have the ability to bond with the fluorescent amyloid dye thioflavin T (ThT), generating fluorescence signals. The method employed in this study demonstrated a detection limit of 16 CFU/ml and exhibited a strong linear correlation within the concentration range of 50 CFU/ml to 10 5 CFU/ml. This method of signal amplification has been effectively utilized to create a fluorescent sensing platform without the need for labels, enabling the detection of P. aeruginosa with high sensitivity.
研究领域:
分子生物学
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病原微生物学
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生物化学
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产品关联
DOI
PubMed
高影响因子
FOOD CONTROL
·
2022
·
IF 6.652
Jinru Zhou,Yanbo Wang,Chaosheng Zhou,Lei Zheng,Linglin Fu
Shellfish allergy represents a crucial public health problem, resulting in an urgent requirement for an accurate, simple, and reliable method to quantify shellfish allergens in the complex food system. Tropomyosin (TM) is known as the major allergen in shellfish. Herein, a novel switch-conversional ratiometric fluorescent biosensor, using TM-specific aptamer as the recognition probe, was developed to detect TM in the complex food system. The proposed aptasensor exhibited good accuracy, stability, and selectivity for the quantification of TM in foodstuffs. Furthermore, the applicability of this proposed aptasensor in complex food samples was verified using seven different shellfish samples. Therefore, this ratiometric fluorescent aptasensor enables accurately detecting the presence of TM traces in foods, providing technical support for allergen label supervision.
研究领域:
食品科学
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分析化学
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生物技术
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过敏与免疫学
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DOI
高影响因子
ANALYTICA CHIMICA ACTA
·
2020
·
IF 5.977
Yu Wang,Xudong Zhao,Man Zhang,Xuan Sun,Jialei Bai,Yuan Peng,Shuang Li,Dianpeng Han,Shuyue Ren,Jiang Wang,Tie Han,Yifei Gao,Baoan Ning,Zhixian Gao
Environmental endocrine disruptors in the environment and food, especially 17 β-estradiol (E2), are important factors affecting the growth and development of organisms. In this research, we constructed a fluorescence strategy for two-step amplification that combined two currently popular methods, exponential amplification reaction (EXPAR) and hybridization chain reaction (HCR). E2 competed with the complementary DNA (cDNA) to bind the aptamer modified on the magnetic beads. The free complementary strand in the supernatant was used as a trigger sequence to activate EXPAR, producing a large amount of short single-stranded DNA (ssDNA). The amplified ssDNA can trigger the second HCR amplification, producing many long double-stranded DNA (dsDNA) analogues. According to the principle of fluorescence resonance energy transfer , the carboxyfluorescein (FAM) signals in H1 and H2 hairpins were quenched by black hole quencher (BHQ-1). After the addition of E2 and initiation of amplification, the initially quenched fluorescent signal would be restored. This strategy with a detection limit of 0.37 pg mL −1 (S/N = 3) showed a good linear relationship in the range of 0.4–800 pg mL −1 . In addition, the recovery rates of the method for milk and water samples were 98.55%–116.95% and 92.32%–107.00%, respectively. This is the first report of the combined detection of EXPAR and HCR, providing a reference for rapid and highly sensitive detection using multiple isothermal amplification methods.
研究领域:
环境科学
·
分析化学
·
生物技术
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DOI
PubMed
高影响因子
ANALYTICAL CHEMISTRY
·
2026
·
IF 7.3
Ye Tian,Xinxi Li,Jingdong Tang,Lei Zhang,Shuai Jiang,Halimulati Muertizha,Zhenwei Yang
Diabetic foot ulcer (DFU), a severe complication of diabetes, necessitates sensitive detection of pathogenic microRNAs (miRNAs) as emerging diagnostic biomarkers. The exponential amplification reaction (EXPAR) offers high efficiency and rapid kinetics for miRNA detection, but its application is limited by persistent nonspecific amplification arising from spurious template–template interactions and interference from partially complementary sequences in complex samples. To address this, we present an integrated strategy combining a blocking probe prehybridized to the EXPAR template to sterically hinder nonspecific hybridization, magnetic bead-based isolation of target-bound probes to remove interferents, and a self-priming mechanism that regenerates the EXPAR primer only upon target recognition. This design ensures high specificity and minimizes primer-independent amplification. Coupling this optimized EXPAR platform with a G-quadruplex-based signal transduction system enables facile colorimetric readout for instrument-free visual detection. The method achieves a detection limit of 41.4 aM across a dynamic range of 100 aM to 50 pM for DFU-associated miRNAs, with excellent selectivity against nontarget miRNAs and consistent reproducibility in fibroblasts and serum. This work provides a generalizable solution to the specificity issue in EXPAR and delivers a reliable, sensitive platform for miRNA profiling in clinical and point-of-care settings.
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产品关联
DOI
PubMed
高影响因子
ACS Applied Nano Materials
·
2024
·
IF 5.3
Runzi Zhang,Shunbi Xie,Xiang He,Ying Gao,Mengjun Wang,Minghang Jiang,Xiaojun Luo,Yi He
Chloramphenicol (CAP), as a broad-spectrum antibiotic, is genotoxic to humans. In this paper, a “Y-shaped” SERS biosensor based on magnetic plasmonic superstructures Fe3O4@SiO2@Ag flowers and the highly efficient isothermal amplification strategy of exponential amplification reaction (EXPAR) technology was proposed for ultrasensitive detection of CAP in milk. This was achieved by forming a stable “Y-shaped” structure on the abundant sharp tips of Fe3O4@SiO2@Ag flowers through specific three-part assembly (Probe 1 connected to the magnetic substrate, Signal Probe 2 Au@4-NTP, and the EXPAR amplification product Amplicon). When the CAP was present, the Amplicon obtained from EXPAR amplification could form a “Y-shaped” structure with the magnetic material and the signal probe. Benefiting from the abundant branching, extensive surface area, and rough surface of the magnetic Fe3O4@SiO2@Ag flower, it offers a multitude of “hot spots” for plasmonic coupling in the electromagnetic field gap region and significantly reduces the nanogap between the signal probe and the silver nanoflowers by constructing a “Y”-shaped structure, thereby maximizing the amplification of the Raman signal. Enabling the detection range of CAP to be between 10–6 and 10–13 M with a detection limit of 17.9 fM. This strategy provided an avenue for trace detection of CAP in food.
研究领域:
食品安全检测
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纳米生物技术
·
分析化学
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产品关联
DOI
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2023
·
IF 8.4
Jiaxin Peng,Xiaomin Hang,Huiyi Wang,Kairen Zhao,Honghong Wang,Haoming Yu,Li Wang
MicroRNAs (miRNAs) have emerged as powerful biomarkers for various diseases diagnosis, yet their clinical application is greatly plagued by the lack of reliable and rapid detection methods. Here, a simple, fast, and accurate point-of-care testing (POCT) platform for miRNA-155 detection was developed by interfacing self-primer exponential amplification reaction (SPEXPAR) with colorimetric and POCT lateral flow assay (LFA). In this platform, a hairpin template was designed for exponential amplification reaction, which could effectively avoid nonspecific template interactions and eventually effectively avoid undesirable background amplification; the colorimetric assay is based on the SPEXPAR amplicons induced gold nanoparticles (AuNPs) color change; and the LFA detection principle involves the amplicons hybridization with designed probes. Under the optimized conditions, the colorimetric assay could detect miRNA-155 as low as 0.41 pM in less than 35 min; and LFA could detect as low as 100 pM in less than 40 min. Furthermore, the method was adopted to detect miRNA-155 in human serum samples with satisfactory recoveries. In summary, this study offered a promising analytical tool for molecular diagnostic testing application.
研究领域:
生物技术
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医学诊断
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分子生物学
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产品关联
DOI
高影响因子
NEW JOURNAL OF CHEMISTRY
·
2018
·Vol. 42
·期 7
·5178-5184
·
IF 3.201
Xingxing Zhang,Yan Jin,Baoxin Li
A copper nanoclusters (CuNCs)-based fluorescence biosensing platform was developed for label-free DNA detection through a cascade isothermal exponential amplification reaction (EXPAR). It is based on target-triggered isothermal dual-cycling and a cascade EXPAR, which generates a large amount of long double-stranded DNA (dsDNA). The resultant long dsDNA can act as an efficient template for fluorescent CuNC synthesis, and the produced CuNC is an excellent fluorescence indicator to quantify target DNA concentration. This assay allows the detection of the model DNA with a detection limit of 7 pM. This label-free assay occurs in a homogenous solution, and the whole assay process, including hybridization, amplification and detection, can be completed in a single-tube. This method is simple, low-cost and convenient to operate, and may offer potential applications in related biomedical research.
研究领域:
分析化学
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生物医学工程
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分子生物学
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DOI
高影响因子
CHEMICAL ENGINEERING JOURNAL
·
2022
·
IF 16.744
Jingyang Zhang,Mengmeng Chen,Yuan Peng,Shuang Li,Dianpeng Han,Shuyue Ren,Kang Qin,Sen Li,Tie Han,Yu Wang,Zhixian Gao
The nucleic tweezers-based detection strategies have shown excellent application prospects in molecular detection and diagnosis thanks to the specific target responsiveness and good biocompatibility. However, the limited detection efficiency and unsatisfactory sensitivity make their applications in complex diagnosis difficult, and few reports focused on it. Here, we explored the molecular dynamics simulations and biosensing performance of the Tetrahedral Nucleic Acid Tweezers, including the Antennae like-Tetrahedron Nucleic Acid Tweezer (ATNAT) and the Covered-Tetrahedron Nucleic Acid Tweezer (CTNAT), revealing the molecular dynamics of ATNAT and CTNAT reporters visually and molecularly. After that, we compared the multi-target detection capabilities of DNA tetrahedral tweezers, and combined the CTNAT reporter with better multi-target detection performance with the aptamer and Exponential amplification reaction (EXPAR), developing an efficient and sensitive EXPAR-cDNA-CTNAT strategy. Then we applied the EXPAR-cDNA-CTNAT strategy to detect testosterone, cortisol, and creatine kinase isoenzymes, realizing sensitive and accurate fatigue diagnosis. Compared with the traditional detection strategies, the EXPAR-cDNA-CTNAT strategy showed improved sensitivity and detection efficiency with excellent specificity, and the limits of detection (LODs) for the multi-target detection were as low as 41, 68, and 8 pM, respectively. The EXPAR-cDNA-CTNAT strategy was reliable for multi-target detection, which had great potential in biological science, food safety, and medical diagnosis.
研究领域:
生物传感
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分子生物学
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纳米技术
·
诊断学
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DOI
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2023
·
IF 8.4
Bing Yang,Lu Shi,Qiaorong Tang,Wei Liu,Baoxin Li,Chaoyong Yang,Yan Jin
Automated and sensitive detection of nucleic acid is crucial for reliable and safe disease diagnostics because the minimum manual operation can effectively avoid sample contamination and infection risk. Herein, a sensitive, label-free and automated method was developed for detecting trace DNA by integrating digital microfluidics (DMF) with chemiluminescence (CL) detection. DMF can automatically manipulate discrete droplets. Taking advantage of sensitive and simple CL detection, DMF-CL integration can automatically and sensitively detect HIV DNA via the preset pathway. To further improve the sensitivity, on-chip exponential amplification reaction (EXPAR) was explored to produce large quantities of double-stranded DNA (dsDNA) in the presence of HIV DNA. Then, SYBR Green I (SG) intercalated into dsDNA to generate singlet oxygen upon light irradiation , which in situ oxidized potassium ferrocyanide into potassium ferricyanide . Finally, the CL reaction of potassium ferricyanide and luminol ensured the label-free, reliable and specific detection of HIV DNA in the range of 10 pM to 2 nM. Saving time-consuming and laborious manual operations, the DMF-CL integration greatly reduced the reagent consumption and avoided the contamination of DNA sample as well. Therefore, DMF-CL offers a promising and useful technology for developing automated and reliable biosensing and bioanalysis.
研究领域:
生物医学工程
·
分析化学
·
分子生物学
·
诊断学
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产品关联
DOI
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2021
·
IF 7.46
Bin Zhou,Lei Lin,Bingzhi Li
Flap endonuclease 1 (FEN1) is a nuclease overexpressed in cells with a high replicate rate, which has been recognized as a potential biomarker in the molecular diagnosis of cancers. In this work, we developed a fluorescent biosensor based on exponential amplification reaction (EXPAR) to realize the rapid and sensitive detection of FEN1. A DNA probe specific for FEN1 is used for the generation of reporter DNA (rDNA), and then the rDNA hybridizes with a template, triggering EXPAR to produce substantial rDNA exponentially. However, when no FEN1 is in the system, the EXPAR will not be initiated because of the lack of rDNA. Derived from the high amplification efficiency of EXPAR, the rapid quantification of FEN1 with a limit of detection (LOD) of 0.5 fM was realized. It is also revealed that this biosensor exhibits good selectivity towards FEN1 under the challenge from real samples including extracts of cells and solid tumors. Furthermore, this sensor was reengineered for the analysis of FEN1 inhibitors, which shows promise in the screening and evaluation of potential drugs targeting FEN1. This work integrates EXPAR into the biosensing of FEN1 for the first time, which provides a promising tool for FEN1-related biomedical investigations.
研究领域:
癌症诊断
·
分子生物学
·
生物医学工程
·
分析化学
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DOI
高影响因子
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
·
2023
·Vol. 415
·期 12
·2271-2280
·
IF 4.3
An Yaqian,Yu Zhiqi,Liu Di,Han Lirong,Zhang Xian,Xin Xuelian,Li Cuiping
The detection of methyltransferase (MTase) activity is of great significance in methylation-related disease diagnosis and drug screening. Herein, a HpaII-assisted and linear amplification-enhanced exponential amplification strategy is proposed for sensitive and label-free detection of M.SssI MTase activity. The P1 probe contains self-complementary sequence 5′-CTAGCCGGCTAG-3′ at 3′-terminal. After denaturation and annealing, P1 probes hybridize with itself to generate P1 duplexes. M.SssI MTase induces methylation of cytosine at 5′-CG-3′ in P1 duplexes, and thus, HpaII fails to cleave at 5′-CCGG-3′ due to methylation sensitivity, leaving P1 duplex intact. Then, these intact P1 duplexes are extended along 3′-terminal through Vent (exo − ) DNA polymerase to generate dsDNA, which is recognized and nicked at the recognition sites by Nt.BstNBI, releasing two copies of primer X. Primer X hybridizes with X′ at the amplification template T1 (X′-Y′-X′) and then serves as primers to trigger the exponential amplification reaction (EXPAR). The point of inflection (POI) values of real-time fluorescence curves is linearly correlated with the logarithm of M.SssI MTase concentration in the range of 0.125 \(-\) 8 U mL −1 with a low detection limit of 0.034 U mL −1 . In the absence of M.SssI, P1 duplexes are cut by HpaII and separated into ssDNA under the executed temperature of EXPAR and thus unable to trigger the amplification. The strategy provides good selectivity against other types of MTases and protein and is able to detect M.SssI activity in human serum. Furthermore, the analytical method has the generality and can be extended to the analysis of other types of DNA MTases. Graphical Abstract
研究领域:
生物化学
·
分子生物学
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分析化学
·
生物技术
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产品关联
DOI
PubMed
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2024
·
IF 8
Di Liu,Xuelian Chen,Yaping Li,Guiping Sun,Jingwen Gao,Hailiang Nie,Cuiping Li
The abuse of 17β-estradiol (E2) in breeding and the excretion into the environment by human beings result in exposure of E2 to humans. Excessive levels of E2 can cause an adverse effect on the human endocrine systems. Therefore, an efficient and sensitive method for the analysis of E2 is required. Herein, based on the specific capture of aptamer, the digestion of Exonuclease III, and efficient amplification of isothermal exponential amplification reaction (EXPAR), a highly sensitive and selective strategy is established for E2 analysis. E2 binds to the aptamer due to their high affinity, preventing the complementary DNA (cDNA) of the aptamer from hybridizing with it. Thus, the cDNA remains unbound and freely available in the solution. Then, Exonuclease III digests the cDNA-aptamer complex, leaving the free cDNA intact. cDNA serves as primers to trigger EXPAR with the help of Bst 2.0 WarmStart DNA polymerase and Nt.BstNBI nicking enzyme, achieving sensitive detection of E2. Without E2, cDNA hybridizes with the aptamer to form double-stranded DNA, which is degraded by Exonuclease III. As a result, the amplification reaction fails to initiate. The approach shows a linear range of 100 aM-10 pM, spanning 5 orders of magnitude, with the limit of detection as low as 36 aM. In addition, the selectivity and reproducibility are great, and the recoveries in tap water and milk ranges from 99.4 % to 117.3 %, suggesting good practicability. This method may also provide a new possibility for sensitive detection of other small-molecule targets via using the corresponding aptamers.
研究领域:
分析化学
·
生物化学
·
环境科学
·
分子生物学
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DOI
高影响因子
危险品化学品经营许可证（带存储）