CHEMICAL ENGINEERING JOURNAL
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2026
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IF 12.5
Shuang Guo,Jiashuo Xu,Huiquan Wang,Wen Li,Tao Meng,Zhe Zhao,Zhiguang Niu,Xiaoqun Gong
We reported a TdT combined with CRISPR/Cas12a driven cascade amplification platform for the sensitive detection of both heavy metal ions and viral targets. • Integrated with reverse fluorescence-enhanced lateral flow strip for visual readout, this platform achieves LODs of 2.5 × 10 −15 M for Pb 2+ and 3.6 × 10 −16 M for DENV-RNA. • The modular design allows flexible replacement of recognition modules, offering a universal solution for on-site rapid monitoring of diverse pollutants in complex environmental matrices. Currently, constructing flexible detection platforms capable of identifying diverse hazardous substances remains a significant challenge. Conventional detection methods are limited by target specificity and lack cross-class universality. Herein, we developed a general platform for sensitive detection of lead ions (Pb 2+ ) and dengue virus (DENV) as two distinct model targets. This platform specifically converted different types of targets into detectable nucleic acid molecules, which would be further recognized and signal amplified by the cascade cycling isothermal amplification system composed of terminal deoxynucleotidyl transferase (TdT) and CRISPR/Cas12a system. TdT has the property of extending the 3′ end of DNA molecule in a template-independent manner, providing long poly-T nucleic acid sequence for binding lots of crRNA, so as to activate the trans-cleavage activity of Cas12a to cleave the hairpin DNA reporters with exposing numerous novel 3′ ends for the same cycle. By combining this strategy with the reverse fluorescence-enhanced lateral flow test strip, the platform achieves a limit of detection (LOD) of 2.5 × 10 −15 M and 3.6 × 10 −16 M for Pb 2+ and DENV-RNA, respectively. Moreover, the detection platform for Pb 2+ in environmental water samples showed excellent agreement with standard methods. It also demonstrated high sensitivity for DENV-RNA detection in complex samples, confirming its reliability and broad applicability potential in practical scenarios. Download: Download high-res image (103KB) Download: Download full-size image
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2022
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IF 9.221
Zhirun Ji,Zhaoyang Shang,Muhammad Sohail,Pei Wang,Bingzhi Li,Xing Zhang,Guoxiang Chen
Protamine, an arginine-rich polycationic peptide, is a natural food preservative and the only approved antidote to heparin overdose. The detection of protamine is meaningful; however, the detection of protamine is difficult due to the lack of aromatic amino acids in its composition . Herein, we proposed a fluorometric biosensor for the sensitive detection of protamine based on the protamine-mediated regulation of CRISPR/Cas12a collateral cleavage. In the absence of protamine, the DNA probe specifically hybridizes with the Cas12a-crRNA complex, causing the activation of Cas12a. In the presence of protamine, positively charged protamine tightly binds to negatively charged DNA probes through electrostatic interaction , so that the DNA probe cannot hybridize with the Cas12a-crRNA complex. Using a short ssDNA modified with fluorophore and quencher as a reporter, the information of protamine concentration is transduced into DNA information and sensed in a signal turn-off manner. The linear range of this method in protamine detection is 0.04–3.2 μg/mL, yielding a detection limit of 0.03 μg/mL, and the entire sampling-to-response time is about 1 h. This biosensor provides a rationale for using the programmable nuclease Cas12a to detect protamine, and further advances the application of CRISPR technology in bioanalyses.
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高影响因子
ANALYST
·
2022
·Vol. 147
·期 24
·5687-5693
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IF 5.227
Yuyuan Zhang,Jing Lei,Wei Liu,Yan Jin,Baoxin Li
Various CRISPR/Cas12a-based biosensing systems have been developed in the past few years, and most of these systems used Taqman probes to report signals through fluorescence resonance energy transfer (FRET). In this study, we explored chemiluminescence resonance energy transfer (CRET) as the readout mode for CRISPR/Cas12a-based biosensing. The chemiluminescence (CL) reaction of bis(2,4,6-trichlorophenyl) oxalate (TCPO) and H2O2 was used to excite the fluorophore dye of the Taqman probe. Different from FRET, CRET does not need external excitation light, which can effectively avoid autofluorescence and photobleaching. The detection limit of this CRET readout mode was estimated to be 10 pM for target DNA, which was about 8 times lower than that of the widely used FRET readout mode. These results suggest that CRET can serve as a rapid, sensitive and simple readout mode of CRISPR/Cas12-based biosensing, and can further enrich the toolbox of CRISPR/Cas12-based biosensing.
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PubMed
高影响因子
FOOD CONTROL
·
2022
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IF 6.652
Baoqing Zhou,Qinghua Ye,Moutong Chen,Chufang Wang,Xinran Xiang,Ying Li,Jumei Zhang,Youxiong Zhang,Juan Wang,Shi Wu,Qihui Gu,Yu Ding,Qingping Wu
Rapid and accurate screening of foodborne pathogens in contaminated food is with great significance for food safety. AuNPs-based colorimetric assay is an attractive strategy in the field of food safety inspection, but is limited by tedious probe preparation, low color rendering resolution, and narrow color gamut. Herein, we proposed an e nhanced c olorimetric p latform based on the C RISPR/Cas12a system and label-free D NA-AuNP p robe (CDP-ECP) for high-throughput detection of Staphylococcus aureus ST398. In this strategy, a novel and specific gene targeting S. aureus ST398 was identified and efficient AuNP-bioprobe was successfully prepared. Through a PCR/RAA amplification, the trans -ssDNA cleavage ability induced by target DNA product triggered an aggregation performance for the prepared AuNP-bioprobe pairs, which could realize target colorimetric detection within 50 min. Under optimal conditions, the colorimetric assay could detect as low as 6.4 × 10 2 cfu/mL of S. aureus ST398 in pure culture. Specific gene mining and target-induced trans -cleavage character gave this method a superior specificity and anti-interference. More importantly, this colorimetric method can be extended to a high-throughput platform for rapid detection of spiked and natural sample by using a common microplate reader. Since it is simple to operate, low cost, high sensitivity and specificity, this new platform is fully qualified for clinical diagnosis, environmental monitoring onsite testing requirements.
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高影响因子
BIOSENSORS & BIOELECTRONICS
·
2022
·
IF 12.545
Feng Qiu,Xiaoyan Gan,Jianglong Yao,Bingying Jiang,Ruo Yuan,Yun Xiang
Transcription factors (TFs) are key substances in regulating the transcription, replication and expression of genes, and the detection of TFs can provide valuable information to diagnose a variety of diseases. By integrating hybridization chain reaction (HCR)-activated Cas12a enzyme with bio-responsive DNA hydrogels, we propose a dual amplification and label-free homogeneous electrochemical detection method to realize sensitive nuclear factor-kappa B p50 (NF-κB p50) detection. The presence of the target molecules protects the DNA duplex probes from digesting by exonuclease III and initiates HCR to generate long double stranded DNAs that can activate the activity of RNA-guided Cas12a enzymes. The single-stranded region of the DNA linkers that crosslink the DNA hydrogels can be cleaved by the activated Cas12a to release a large number of electroactive substances embedded in the gels, which exhibit highly enhanced electrochemical signals for detecting target molecules at the detection limit of 54.1 fM. In addition, the successful interrogation of NF-κB p50 spiked into lysate of HeLa cells by such method is also verified. The established method thus shows new opportunities for sensitive and convenient monitoring of other transcription factors and biomarkers.
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PubMed
高影响因子
LWT-FOOD SCIENCE AND TECHNOLOGY
·
2023
·
IF 6
Yingchun Li,Yaqin Zhang,Fange Kong,Chunxia Wang,Shanshan Chen,Jiasi Wang,Di Wang
Fusarium temperatum ( F. temperatum ) causes maize stalk rot disease, reducing grain yield and producing multiple harmful mycotoxins that threaten food safety and quality. However, current detection methods are limited by the requirement of well-trained personnel, complicated operations and expensive instruments. To address this challenge, gold nanoparticles (AuNPs)-based colorimetric assay using loop-mediated isothermal amplification (LAMP) coupled with CRISPR-Cas12a (Au-CRISPR) was tested for F. temperatum detection. The assay utilizes DNA-modified AuNPs as a colorimetric probe and the results can be readily observed by the naked eye. After optimization, the method can specifically detect as low as 100 copies/μL of target DNA or 10 −8 ng/μL of extracted DNA from F. temperatum . The detection platform allows high-throughput testing by integrating microtiter plates and microplate reader. Furthermore, a portable smartphone-based device was developed to realize point of care (POC) detection of F. temperatum in resource-limited settings. This simple, inexpensive and sensitive detection platform has great potential for the applications in field detection.
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2024
·
IF 8
Sirui Liu,Xinlan Bian,Furong Zhao,Min Cao,Zhaoyang Shang,Bingzhi Li,Xing Zhang
High mannose is overexpressed on the cell surface during carcinogenesis; however, there are currently no efficient techniques for its detection. Herein, we propose an assay called RADAR ( r emote high m a nnose d etection on cell surf a ce with C R ISPR) for the quantification of high mannose, leveraging the integration of high mannose-binding aptamer and CRISPR technique. The high mannose-binding aptamer is hybridized with a locker, which is released upon specific binding to high mannose. The released locker then activates CRISPR/Cas12a to produce a fluorescence signal. The "remote" concept in RADAR refers to the recognition of high mannose occurring on the cell surface, while signal generation takes place in a solution separate from the target. This feature makes RADAR suitable for the detection of suspended bio-samples without the need for sophisticated imaging. RADAR can detect high mannose Man 9 with a limit of detection of 2.5 nM, distinguish cancer cells from non-cancerous cells, and profile high mannose level in human serum for potential non-small cell lung cancer (NSCLC) diagnosis. This work represents the first attempt to integrate CRISPR technology into the aptasensing of high mannose, which may serve as a synthetic biology-enabled tool in glycochemistry and glycobiology.
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高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2024
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IF 5.7
Yan Zhao,Qianqian Yu,Miaolin Duan,Qi Zhang,Zhishang Li,Yan Zhang,Yana Liu,Han Wang,Xingmin Li,Ruitong Dai,Fei Jia
The efficient, sensitive, and rapid detection of Salmonella typhimurium (S. typhimurium) in food and food products is important to ensure food safety and health. This study developed a fluorescence biosensing assay that integrated recombinase-aided amplification (RAA) and CRISPR/Cas12a with a zeolitic imidazolate framework-8@fluorescein sodium (ZIF-8@FLS) nanocomposite for the sensitive detection of S. typhimurium. In this approach, using RAA as a preamplification module, CRISPR/Cas12a-AChE as a target recognition and dual-enzyme cascade amplification module, and the prepared ZIF-8@FLS with high porosity and rapid pH responsiveness as a fluorescence signal explosive amplification module, the RAA-CRISPR/Cas12a-ZIF-8@FLS biosensor was constructed. Under optimal conditions, it exhibited an excellent linear relationship for S. Typhimurium, with a sensitive detection limit as low as 1.3 × 102 CFU/mL and could complete sample detection within 2 h relying on the RAA and ZIF-8@FLS explosive fluorescence rapid response, demonstrating its significant advantages in specificity, sensitivity, and reliability in food-borne pathogens detection.
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PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2024
·
IF 10.7
Chuanyu Yang,Caiyi Du,Feiyu Yuan,Peitong Yu,Boxu Wang,Changshun Su,Ruiqi Zou,Junyang Wang,Xu Yan,Chunyan Sun,Hongxia Li
Benefiting from specific target recognition and trans-cleavage capabilities, the CRISPR/Cas12a system has great application prospects in the design of highly sensitive and rapid fluorescence biosensors. The CRISPR/Cas12a-based fluorophore-quencher molecular beacons exhibit single-color emission and are easily exposed to interference from environmental factors. Herein, we design a CRISPR/Cas12a-derived ratiometric fluorescence sensor for Pb 2+ detection based on embedded carbon dots@zeolitic imidazolate framework-8 (CDs@ZIF-8) composites and DNAzyme. The functions of ZIF-8 about encapsulating red emissive CDs in the inner cavity and adsorbing DNA on the outer surface are integrated to establish dual fluorescence signals, thereby reducing the possibility of interference and improving sensing accuracy. The presence of Pb 2+ is converted into the change of activator by the GR5 DNAzyme to activate the CRISPR/Cas12a system, which provides signal amplification through multiple turnovers of side branch cutting, achieving highly sensitive detection of Pb 2+ with a low detection limit of 18 pM. This method has the advantages of simplicity, universality, and excellent quantitative ability, and has broad prospects in sensing applications.
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PubMed
高影响因子
TALANTA
·
2024
·
IF 5.6
Qiang Tang,Xiaoling Huang,Guangxing Liu,Hongran Huang,Jiayi Zhang,Jihua Wei,Yuanxun Gong,Qianli Tang,Kai Zhang,Xianjiu Liao
This study introduces a novel electrochemical biosensor for detecting Matrix Metalloproteinase-2 (MMP-2), a key biomarker in cancer diagnostics and tissue remodeling. The biosensor is based on a dual-amplification strategy utilizing T7 RNA polymerase isothermal amplification and CRISPR-Cas12a technology. The principle involves the release of a DNA template in the presence of MMP-2, leading to RNA synthesis by T7 RNA polymerase. This RNA activates CRISPR-Cas12a, which cleaves a DNA probe on the electrode surface, resulting in a measurable electrochemical signal.The biosensor demonstrated exceptional sensitivity, with a detection limit of 2.62 fM for MMP-2. This high sensitivity was achieved through the combination of transcriptional amplification and the collateral cleavage activity of CRISPR-Cas12a, which amplifies the signal. The sensor was able to detect MMP-2 across a wide dynamic range from 2 fM to 1 nM, showing a strong linear correlation between MMP-2 concentration and the electrochemical signal. In practical applications, the biosensor accurately detected elevated levels of MMP-2 in cell culture supernatants from HepG2 liver cancer cells, distinguishing them from normal LO2 liver cells. The use of an MMP-2 inhibitor confirmed the specificity of the detection. These results underscore the biosensor's potential for clinical diagnostics, particularly in early cancer detection and monitoring of tissue remodeling activities. The biosensor's design allows for rapid, point-of-care testing without the need for complex laboratory equipment, making it a promising tool for personalized healthcare and diagnostic applications.
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2024
·
IF 8
Honglian Wu,Ruolin Zhang,Huang Lai,Luogen Lai,Yi Liu,Yadi Fan,Yu Zhang,Jingyu Shi,Mo Yang
Endotoxins or lipopolysaccharides (LPS) are potent inflammatory agents that pose serious health risks even at low concentrations, thus requiring sensitive and reliable detection methods. Current methods for endotoxin detection, primarily the Limulus Amebocyte Lysate (LAL) assay and Enzyme-linked Immunosorbent Assays (ELISA) are limited by complex procedures, prolonged assay time, and susceptibility to interference. To address these challenges, we developed a single-reporter probe-based dual-mode biosensor integrating MXene-Au nanosheets with the CRISPR/Cas12a system for endotoxin detection. The sensing platform was constructed by conjugating ssDNA-Cy5 reporters onto MXene-Au nanosheets. MXene-Au nanosheets served dual functions, including efficient quenching of fluorescence and strong enhancement of SERS signals. In the presence of target endotoxins, CRISPR/Cas12a remains inactive, leaving the ssDNA-Cy5 reporters intact on the MXene-Au surface. The close proximity of Cy5 to the MXene-Au surface results in quenched fluorescence (“FL off”) but enhanced SERS signals (“SERS on”). Conversely, in the absence of endotoxins, activated CRISPR/Cas12a cleaves the ssDNA-Cy5, releasing Cy5 from the surface, thus leading to fluorescence recovery (“FL on”) but diminished SERS signals (“SERS off”). By measuring the changes in SERS and FL signals and analysing the ratiometric I SERS /I FL signals, the biosensor achieved a lower limit of detection (LOD) of 15.9 pg/mL within 30 minutes compared to single detection modes. Validation studies in complex matrices, including spiked water and milk samples, showed performance comparable to commercial endotoxin detection kits. Overall, our CRISPR-Cas12a-mediated fluorescent-SERS dual-mode biosensor not only improves detection sensitivity but also provides built-in quality control, improving the reliability of results through cross-validation.
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高影响因子
LWT-FOOD SCIENCE AND TECHNOLOGY
·
2025
·
IF 6.6
Xingyue Tang,Yafang Shen,Jiangqi Wang,Zhaohui Qiao,Fangkai Han,Yanna Cui,Qiuyue Hu,Yuxin Wu,Jiahua Wang,Xiaodan Liu,Huang Dai,Guijie Hao
Sensitive and accurate detection of enrofloxacin (ENR) residues is essential for food safety. Herein, a competitive immunosensor was developed for the detection of ENR, using a gold nanoparticle (AuNP)-enhanced CRISPR/Cas12a system. In this method, bovine serum albumin-ENR- and single-stranded DNA-modified AuNPs (AuNPs-BSA-ENR/ssDNA) was used to act as both a competitor and CRISPR/Cas12a activator. The use of AuNPs-BSA-ENR/ssDNA helped transduce the target recognition information to the cleavage activity of the CRISPR/Cas12a system. Moreover, benefiting from the high ssDNA loading capability of AuNPs, it enabled one-to-multiple CRISPR/Cas12a activation. The synergetic signal amplification from AuNPs and the CRISPR/Cas12a system guaranteed the sensitivity of this method with a limit of detection of 1 ng/mL. Its feasibility for the detection of food and environmental samples was also investigated with recovery rates ranging from 91.0% to 115.2%. The developed immunosensor offered a promising alternative for ENR detection.
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高影响因子
危险品化学品经营许可证（带存储）