JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2023
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IF 6.1
Yayun Jiang,Chaochuan Zheng,Ming Jin,Ruolan Zhou,Qiaoli Wu,Fuyuan Huang,Yongliang Lou,Laibao Zheng
Accurate, rapid, and sensitive pathogenic detections play an important role in food safety. Herein, we developed a novel CRISPR/Cas12a mediated strand displacement/hybridization chain reaction (CSDHCR) nucleic acid assay for foodborne pathogenic colorimetric detection. A biotinylated DNA toehold is coupled on avidin magnetic beads and acts as an initiator strand to trigger the SDHCR. The SDHCR amplification allowed the formation of long hemin/G-quadruplex-based DNAzyme products to catalyze the TMB-H2O2 reaction. In the presence of the DNA targets, the trans-cleavage activity of CRISPR/Cas12a was activated to cleave the initiator DNA, resulting in the failure of SDHCR and no color change. Under optimal conditions, the CSDHCR has a satisfactory linear detection of DNA targets with a regression equation Y = 0.0531*X – 0.0091 (R2 = 0.9903) in the range of 10 fM to 1 nM, and the limit of detection was determined as 4.54 fM. In addition, Vibrio vulnificus, one foodborne pathogen, was used to verify the practical application of the method, and it showed satisfactory specificity and sensitivity with a limit of detection at 1.0 × 100 CFU/mL coupling with recombinase polymerase amplification. Our proposed CSDHCR biosensor could be a promising alternative method for ultrasensitive and visual detection of nucleic acids and the practical application of foodborne pathogens.
研究领域:
食品安全
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分子生物学
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生物技术
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病原体检测
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核酸分析
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产品关联
DOI
PubMed
高影响因子
Small
·
2025
·
IF 12.1
Chenggong Zhu,Qunfang Huang,Ruijie Fu,Zhen Xun,Qishui Ou,Yunlei Xianyu,Can Liu
Hepatitis B virus (HBV) infection remains a significant global public health issue, and rapid detection of HBV DNA is crucial for disease prevention and control. However, traditional methods for HBV DNA detection are limited by their reliance on precise instruments and single readout, which can hardly meet the requirements of on-site detection. In this study, the Mg 2+ -enhanced trans-cleavage activity of clustered regularly interspaced short palindromic repeats/associated protein 12a (CRISPR/Cas12a) is reported and coupled with loop-mediated isothermal amplification (LAMP) and Au@Pt nanoparticles as a signaling reporter for on-site detection of HBV DNA. This triple-modal biosensing strategy enables multiple signal readouts including UV–vis spectrum, RGB value, and temperature with high sensitivity and accuracy. The linear detection ranges using UV–vis spectroscopy, RGB color recognition, and photothermal modes are from 50 to 10,000 copies µL −1 , with the limitation of detection of 24.07, 39.65, and 23.33 copies µL −1 , respectively. This biosensing strategy is further employed for the qualitative detection of HBV DNA in 48 serum samples, achieving sensitivities of 100%, 100%, and 95.24% for triple modes. This work offers a promising tool of next-generation LAMP-CRISPR/Cas12a for the rapid and portable detection of nucleic acids.
研究领域:
生物传感器
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病毒学
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纳米技术
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分子诊断学
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CRISPR技术
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分析化学
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产品关联
DOI
PubMed
高影响因子
TALANTA
·
2021
·
IF 6.057
Kai Zhang,Zhenqiang Fan,Yue Huang,Yuedi Ding,Minhao Xie
Early diagnosis and timely management of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) are the keys to preventing the spread of the epidemic and controlling new infection clues. Therefore, strengthening the surveillance of the epidemic and timely screening and confirming SARS-CoV-2 infection is the primary task. In this work, we first proposed the idea of activating CRISPR-Cas12a activity using double-stranded DNA amplified by a three-dimensional (3D) DNA walker. We applied it to the design of an electrochemiluminescent (ECL) biosensor to detect the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) gene. We first activated the cleavage activity of CRISPR-Cas12a by amplifying the target DNA into a segment of double-stranded DNA through the amplification effect of a 3D DNA walker. At the same time, we designed an MXene based ECL material: [email protected] 3 C 2 @AuNPs, and constructed an ECL biosensor to detect the RdRp gene based on this ECL material as a framework. Activated CRISPR-Cas12a cleaves the single-stranded DNA on the surface of this sensor and causes the ferrocene modified at one end of the DNA to move away from the electrode surface, increasing the ECL signal. The extent of the change in electrochemiluminescence reflects the concentration of the gene to be measured. Using this system, we detected the SARS-CoV-2 RdRp gene with a detection limit of 12.8 aM. This strategy contributes to the rapid and convenient detection of SARS-CoV-2-associated nucleic acids and promotes the clinical application of ECL biosensors based on CRISPR-Cas12a and novel composite materials.
研究领域:
生物技术
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纳米技术
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分子生物学
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病毒学
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分析化学
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产品关联
DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2021
·
IF 10.618
Kai Zhang,Zhenqiang Fan,Bo Yao,Yuedi Ding,Jing Zhao,Minhao Xie,Jianbin Pan
Sialic acid-binding immunoglobulin (Ig)-like lectins (Siglecs) is a type I transmembrane receptor on the cell surface. Siglec-5, as one of the Siglecs family, play an important role as an inhibitory receptor for leukocytes in the human body. The development of novel siglec-5 assays can help to study the pathogenesis of related diseases as well as to develop novel therapeutic drugs. We use catalytic hairpin assembly (CHA) amplification strategy combined with CRISPR-Cas12a′s side-cutting feature to build a 2D ultra-thin Ti 3 C 2 T x (MXene) based electrochemiluminescence (ECL) biosensor for the detection of Siglec-5. By using this ECL biosensor, the cleavage of CRISPR-Cas12a is reasonably combined with CHA-mediated isothermal amplification, thereby realizing the sensitive amplification assay Siglec-5 with 20.22 fM sensitivity. By introducing pairs of sites that are not in the same double-stranded DNA into the DNA duplex, the hybridization sequence of CRISPR-Cas12a complements the targeting mechanism to enhance indirect Siglec-5 amplification assay. Also, the double-strand DNA (dsDNA) design based on CRISPR-Cas12a amplification allows the same CRISPR RNA (crRNA, also known as guide RNA (gRNA)) to detect the output of DNA duplexes from different intermediate DNAs, which provides a common way for biomarker detection based on the conversion of protein analytes to intermediate DNA strategy. This work extends the application scope of CRISPR-Cas12a to the construction of ECL biosensors, evaluates the role of lectins, which can be used for the biochemical research and clinical diagnosis of protein markers. This is the first investigative work exploring the Trans-Cleavage activity of CRISPR-Cas12a for Mxene-based ECL biosensor establishment to the best of our knowledge.
研究领域:
生物技术
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生物化学
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纳米技术
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分子生物学
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临床诊断
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产品关联
DOI
PubMed
高影响因子
ANALYTICA CHIMICA ACTA
·
2024
·
IF 5.7
Kai Shi,Yi Tian,Sujun Liu,Wenjie Luo,Keer Liu,Lin Zhang,Ying Zhang,Jiali Chang,Jiaheng Zhang,Shuo Wang
Background Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas12a-powered biosensor with a G-quadruplex (G4) reporter offer the benefits of simplicity and sensitivity, making them extensively utilized in detection applications. However, these biosensors used for monitoring pollutants in environmental water samples may face the problem of high background signal and easy interference due to the “signal-off” output. It is obvious that a biosensor based on the CRISPR/Cas12a system and G4 with a “signal on” output mode needs to be designed for detecting environmental pollutants. Results By using phosphorothioate-modified G4 as a reporter and catalytic hairpin assembly (CHA) integrated with Cas12a as an amplification strategy, a “signal-on” colorimetric/photothermal biosensor (psG4-CHA/Cas) for portable detection of environmental pollutants was developed. With the help of functional nucleotides, the target pollutant (kanamycin or Pb 2+ ) triggers a CHA reaction to produce numerous double-strand DNA, which can activate Cas12a′s trans -cleavage activity. The active Cas12a cleaves locked DNA to release caged psG-rich sequences. Upon binding hemin, the psG-rich sequence forms a psG4/hemin complex, facilitating the oxidation of the colorless 3,3′,5,5′-tetramethylbenzidine (TMB) into the blue photothermal agent (oxTMB). The smartphone was employed for portable colorimetric detection of kanamycin and Pb 2+ . The detection limits were found to be 100 pM for kanamycin and 50 pM for Pb 2+ . Detection of kanamycin and Pb 2+ was also carried out using a portable thermometer with a detection limit of 10 pM for kanamycin and 8 pM for Pb 2+ . Significance Sensitive, selective, simple and robust detection of kanamycin and Pb 2+ in environmental water samples is achieved with the psG4-CHA/Cas system. This system not only provides a new perspective on the development of efficient CRISPR/Cas12a-based “signal-on” designs, but also has a promising application for safeguarding human health and environmental monitoring.
研究领域:
环境科学
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生物技术
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纳米技术
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分析化学
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生物化学
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产品关联
DOI
PubMed
高影响因子
CHEMICAL ENGINEERING JOURNAL
·
2021
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IF 13.273
Kai Zhang,Zhenqiang Fan,Yuedi Ding,Minhao Xie
In this work, we constructed an exonuclease III cleavage reaction-based isothermal amplification of nucleic acids with CRISPR/Cas12a-mediated pH-induced regenerative Electrochemiluminescence (ECL) biosensor for ultrasensitive and specific detection of SARS-CoV-2 nucleic acids for SARS-CoV-2 diagnosis. The triple-stranded nucleic acid in this biosensor has an extreme dependence on pH, which makes our constructed biosensor reproducible. This is essential for effective large-scale screening of SARS-CoV-2 in areas where resources are currently relatively scarce. Using this pH-induced regenerative biosensor, we detected the SARS-CoV-2 RdRp gene with a detection limit of 43.70 aM. In addition, the detection system has good stability and reproducibility, and we expect that this method may provide a potential platform for the diagnosis of COVID-19.
研究领域:
生物技术
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纳米技术
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分子生物学
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分析化学
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病毒学
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产品关联
DOI
PubMed
高影响因子
CHEMICAL ENGINEERING JOURNAL
·
2024
·
IF 13.3
Yuexin Wang,Bo Shen,Nini Luo,Cai Li,Haiping Wu,Yanshuang Wang,Shen Tian,Xuemiao Li,Rui Liu,Xinmin Li,Junman Chen,Wei Cheng,Shijia Ding,Rui Chen,Meifang Xiao,Qianfeng Xia
The simple, rapid, and accurate detection of highly lethal melioidosis is crucial for early clinical diagnosis and improving cure rates. Currently, due to time consumption, low sensitivity and detection rate the existing clinical detection methods cannot satisfy the needs of clinical diagnosis. Herein, a novel self-enhanced porous hydrogel material (Au@PEI-ABEI@Pt) for ultrasensitive ECL strategy of detection B. pseudomallei was report. The novel porous hydrogel material composed of PEI-ABEI porous hydrogel, gold nanoparticles (AuNP) and platinum nanoparticles (PtNP) have large specific surface area and porous structure, which not only fix more ABEI to realize self-enhanced ECL signal amplification, but also facilitate ion diffusion and efficient utilization of catalytic materials, realizing rapid electron transfer and zero-distance catalysis, significantly improving the initial signal of ECL sensor. Moreover, the most attractive aspect is that Au@PEI-ABEI@Pt hydrogels with good biocompatibility can achieve widespread application of CRISPR/Cas12a in solid-phase carriers without affecting the sensitivity, specificity and shearing activity of CRISPR/Cas12a. After coupling with the ECL system and CRISPR/Cas12a signal amplification strategy, the Au@PEI-ABEI@Pt can achieve an ultrasensitive ECL assay of B. pseudomallei with the LOD of 5 CFU mL −1 in complex samples, with high specificity and stability to effectively classify B. pseudomallei and other Gram-negative bacteria. This study shows that the developed porous hydrogel materials not only serve as an excellent ECL signal reporter to significantly improve the detection sensitivity of ECL biosensors, but also provide a new approach for the wide application of CRISPR/Cas systems in solid-phase carriers.
研究领域:
生物技术
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纳米技术
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分析化学
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生物传感器
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分子生物学
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医学诊断
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产品关联
DOI
高影响因子
Environmental Science-Nano
·
2022
·Vol. 9
·期 9
·3417-3426
·
IF 9.473
Fan Yang,Wei Wang,Mei Zhang,Wenxin Tao,Youwang Wang,Jiameng Shi,Yuedi Ding,Minhao Xie,Sai Zhang,Zhenqiang Fan,Kai Zhang
Since the emergence of coronavirus disease 2019 (COVID-19), this highly contagious disease has ravaged the world, cumulatively causing millions of deaths and huge economic losses. As the culprit of COVID-19, severe acute respiratory syndrome β-coronavirus 2 (SARS-CoV-2) is highly infectious and pathogenic, which has caused extreme panic worldwide. Early and rapid monitoring of SARS-CoV-2 has a significant role in suppressing the pandemic and reducing the virus's lethality. In our work, we prepared a self-enhanced ruthenium complex linked to zeolitic imidazole framework-8 (ZIF-8) and used it as an electrochemiluminescence (ECL) emitter. Additionally, a double-stranded specific nuclease (DSN)-assisted target RNA cycling with catalytic hairpin assembly (CHA) signal amplification technology was used to achieve the conversion of target RNA concentration to double-stranded DNA (dsDNA) output which significantly improved the detection sensitivity of target RNA under environmental conditions and in real human serum samples. In addition, we also combined the trans-cleavage property of CRISPR–Cas12a with the adsorption property of C3N4 on a ferrocene (Fc)-labeled DNA probe and obtained target RNA-dependent ECL signals. The reliable detection protocol achieved the transformation of SARS-CoV-2 RNA concentration to ECL responses, obtaining a limit of detection (LOD) of 0.67 fM with high specificity and reproducibility, which was of guiding significance for current detection methods of mutant SARS-CoV-2 and universal RNA.
研究领域:
生物技术
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分析化学
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分子生物学
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病毒学
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产品关联
DOI
高影响因子
SCIENCE OF THE TOTAL ENVIRONMENT
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2023
·
IF 9.8
Junhua Chen,Gu Shi,Chong Yan
On-site and cost-effective monitoring of antibiotic residue in water samples using a ubiquitous device that is readily available to the general public is a big challenge. Herein, we developed a portable biosensor for kanamycin (KAN) detection based on a glucometer and CRISPR-Cas12a. The aptamer-KAN interactions liberate the trigger C strand, which can initiate the hairpin assembly to produce numerous double-stranded DNA. After recognition by CRISPR-Cas12a, Cas12a can cleave the magnetic bead and invertase-modified single-stranded DNA. After magnetic separation, the invertase can convert sucrose into glucose, which can be quantified by a glucometer. The linear range of the glucometer biosensor is from 1 pM to 100 nM and the detection limit is 1 pM. The biosensor also exhibited high selectivity and the nontarget antibiotics had no significant interference with KAN detection. The sensing system is robust and can work in complex samples with excellent accuracy and reliability. The recovery values were in the range of 89–107.2 % for water samples and 86–106.5 % for milk samples. The relative standard deviation (RSD) was below 5 %. With the advantages of simple operation, low cost, and easy accessibility to the public, this portable pocket-sized sensor can realize the on-site detection of antibiotic residue in resource-limited settings.
研究领域:
生物技术
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环境科学
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分析化学
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即时诊断
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产品关联
DOI
PubMed
高影响因子
ANALYTICAL CHEMISTRY
·
2025
·
IF 6.7
Fei Hu,Kaihui Liu,Yunyun Zhang,Shuhao Zhao,Tianyi Zhang,Cuiping Yao,Xing Lv,Jing Wang,Xiaolong Liu,Niancai Peng
Infectious diseases, especially respiratory infections, have been significant threats to human health. Therefore, it is essential to develop rapid, portable, and highly sensitive diagnostic methods for their control. Herein, a short-time preamplified, one-pot clustered regularly interspaced short palindromic repeats (CRISPR) nucleic acid detection method (SPOC) is developed by combining the rapid recombinase polymerase amplification (RPA) with CRISPR-Cas12a to reduce the mutual interference and achieve facile and rapid molecular diagnosis. SPOC can reduce the detection time and stably detect up to 1 copy/μL of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA without affecting the detection sensitivity. A highly sensitive one-pot assay integrated with reverse transcription RPA is achieved by wrapping paraffin with a specific melting point on the lyophilized CRISPR reagent surface. A self-heating pack is designed based on thermodynamic principles to melt the paraffin and release CRISPR reagents, enabling low-cost and time-saving detection. Notably, the designed system, coupled with RNA extraction-free technology, can achieve “sample-in-answer-out” detection of the SARS-CoV-2 Orf1ab gene within 22 min using smartphone imaging. The developed assay is validated on 12 clinical samples, and the results 100% correlate with real-time polymerase chain reaction. SPOC is time-saving, is easy to operate, and can eliminate centrifugal and complex hardware devices, satisfying the demand for point-of-care diagnostics in resource-constrained settings.
研究领域:
分子诊断学
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传染病学
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生物技术
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产品关联
DOI
PubMed
高影响因子
ANALYTICA CHIMICA ACTA
·
2025
·
IF 6
Fei Ding,Xiaomin Hang,Sen Tian,Wei Cao,Jingwen Wu,Li Wang
Primer exchange reaction (PER) is a novel and simple nucleic acid-templated extension technique that has recently attracted much attention in the field of biosensing. However, current PER reactions have shown relatively slow rates and low amplification performances, resulting in long assay times and limited detection sensitivities. Here we report a nicking endonuclease-mediated PER reaction (named NEPER) that rapidly releases amplified DNA products by adding a nicking endonuclease to hydrolyze the hybridized double-stranded DNA (dsDNA), and consequently has a maximum speed that is thirty orders of magnitude greater than the maximum for conventional PER. We further combined a CRISPR/Cas12a signal readout technique and developed a cascade NEPER-CRISPR/Cas12a method that can detect miRNA-155 with a limit of detection (LOD) down to 3.1 fM. We also show that the NEPER-CRISPR/Cas12a can be used to detect targets in serum samples.
研究领域:
分子生物学
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生物传感
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诊断学
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产品关联
DOI
PubMed
高影响因子
Current Research in Food Science
·
2025
·
IF 7
Lijuan Yin,Lijing Sun,Yueqi Wang,Gailing Shi,Liangcai Lin,Cuiying Zhang
Baijiu is the product of the cooperation of a variety of core functional organisms in an open fermentation system. The differences in the structure of these core functional microorganisms are directly related to the quality of the base liquor. Acetilactobacillus jinshanensis is the dominant functional microorganism involved in the process of making Jiang-flavour Baijiu. To quantitatively evaluate the concentration of A. jinshanensis in the fermentation system, a CRISPR/Cas12a-coupled gold nanoparticle visual biosensor was developed. The limit of detection for A. jinshanensis was as low as 1 copy/μL, and its sensitivity exceeded that of the existing quantitative real-time PCR method. Furthermore, this system was employed to determine the concentration of A. jinshanensis during actual Baijiu fermentation. The average fluorescence value of A. jinshanensis in excellent-grade workshop was 4838, while that in ordinary workshops was 2145. The results revealed a linear relationship (R 2 =0.845) between the quality of the base liquor and the concentration of A. jinshanensis was obtained. Therefore, the concentration of A. jinshanensis might be used as a new parameter to predict the quality of base liquor and as a regulatory target in the fermentation process, thereby achieving a controllable fermentation process and high quality.
研究领域:
微生物学
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分子生物学
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食品科学
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DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）