FOOD CHEMISTRY
·
2025
·
IF 9.8
Ruonan Wang,Baoshan He,Zhengyong Liang,Yao Liu,Jun Yang,Huali Jin,Min Wei,Wenjie Ren,Zhiguang Suo,Yiwei Xu
This study presents an integrated dual-mode sensing strategy, in which qualitative fluorescence screening is followed by quantitative electrochemical detection, improving detection efficiency and enabling rapid PCM analysis. It develops a novel fluorescence electrochemical aptasensor that combines in vitro “Dark-box” applications with CRISPR/Cas12a system electrode surface sensing technology. PCM activates the DNA walker, and the DNAzyme induces cyclic cleavage of DNA strands bearing the Carboxyfluorescein (FMA) group. After magnetic separation, the fluorescence reaction combined with the “Dark box” enables the preliminary qualitative analysis of procymidone (PCM). Following the preliminary qualitative detection, the solution is introduced to the electrochemical aptasensor platform integrated with the CRISPR/Cas12 system. The Cas12a system triggers cyclic cleavage, producing a signal change that enables the electrochemical quantitative detection of PCM. An fluorescence (FL) response occurs when the PCM concentration in spiked samples is at or above 1 pg·mL −1 , allowing for qualitative fluorescence detection. The EC platform's detection limit is 8.51 × 10 −6 ng·mL −1 , with a range of 1 × 10 −2 ng·mL −1 to 1 × 10 4 ng·mL −1 . The designed dual-mode sensor provides reliable monitoring of PCM in real samples.
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高影响因子
ANALYTICAL CHEMISTRY
·
2025
·
IF 6.7
Kai Shi,Wenjie Luo,Ying Cheng,Honglei Li,Liai Peng,Xiangrui Luo,Yu Hu,Jiaheng Zhang,Jiaxuan Chen
CRISPR/Cas12a-powered biosensors with guanine (G)-rich sequence reporters (e.g., G-quadruplex and G-triplex) are widely used in detection applications due to their simplicity and sensitivity. However, when these biosensors are employed for molecular detection in complex samples, they may encounter difficulties such as high background signal and susceptibility to interference because of the “turn-off” signal output. Herein, we explore, for the first time, a set of phosphorothioate (ps)-modified G-quadruplex (G4) and G-triplex (G3) sequences that can bind with thioflavin T (ThT) in an active split CRISPR/Cas12a system (SCas12a) to generate a “turn-on” fluorescent signal. To apply this new phenomenon, we develop a universal SCas12a-powered biosensor for “turn-on” fluorescent detection of nucleic acid (miRNA-21) and non-nucleic acid (kanamycin) targets by using ps-modified hairpin G3 as a reporter (SCas12a/psHG3). Target recognition activates SCas12a’s trans-cleavage activity, leading to cleavage at the loop region of the psHG3 reporter. The released prelocked psG3 DNA binds ThT to produce a strong fluorescence signal. Without preamplification, this strategy can detect miRNA-21 with a detection limit of 100 fM. Moreover, the SCas12a/psHG3 system was further utilized for detecting kanamycin by incorporating its aptamers, enabling the detection of kanamycin at concentrations as low as 100 pM. This work is the first to develop a “turn-on” SCas12a/psHG3 system, showcasing its improved performance and wide range of applications in synthetic biology-based sensing technology.
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ANALYTICAL CHEMISTRY
·
2026
·
IF 6.7
Miao He,Jinhua Chen,MuRilege Chao,Feifan Yin,Han Pan,Qiufeng Wang,Wenting Cheng,Zhongyun Wang,Yang Xiang
mRNA vaccines, as a new form of vaccine, are gradually revealing their potential in the field of tumor treatment. With the development of mRNA vaccines, monitoring their levels has become particularly important. In this work, a novel and innovative strategy was proposed for the first time, which directly used the target as crRNA and simultaneously triggered an entropy-driven split Cas12a system for triple signal amplification for the high-sensitivity detection of mRNA vaccines. This study broke the traditional full-length crRNA and innovatively designed a split crRNA structure, directly using the target mRNA as the variable sequence of the crRNA. This design avoided the need for additional introduction of variable sequences, thereby eliminating nonspecific signal leakage and multicomponent interference from the source. At the same time, the target molecule served as the initiating chain for the EDC reaction, allowing the split Cas12a system to be coupled with the entire entropy-driven reaction process, achieving efficient collaboration and triple signal amplification and significantly improving the detection sensitivity and specificity. Moreover, the entire detection process required only one reaction step, which was simple, fast, and efficient. The aim was to provide strategies for screening and pharmacokinetic studies of mRNA vaccines.
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Analytical Methods
·
2026
·
IF 3
Jing Chen,Yanna Le,Luoxian Yang,Hua Qin,Wanli Yang
Chronic exposure to lead ions (Pb2+) disrupts intestinal flora homeostasis, underscoring the need for sensitive and practical detection methods. Herein, we report a dual-mode analytical strategy combining split-DNA re-assembly-driven colorimetric reaction with CRISPR/Cas12a trans-cleavage activity for Pb2+ analysis. A bifunctional magnetic probe is constructed, where Pb2+-specific DNAzyme recognition triggers cleavage and release of crRNA, which subsequently activates Cas12a to generate fluorescence by cleaving a fluorophore-quencher reporter. Concurrently, the remaining magnetic bead-anchored trigger initiates split G-quadruplex re-assembly, catalyzing a visible colorimetric reaction. Under optimized conditions, the fluorescence mode achieves a detection limit of 1.03 fM with a linear range of 5 fM to 100 pM, while the colorimetric mode offers a limit of 62.1 fM from 100 fM to 500 pM. Both modes exhibit excellent specificity against competing metal ions. The platform's clinical feasibility is validated using human serum samples, showing strong correlation with ICP-MS, satisfactory recoveries (97.1–103.4%), and good repeatability (CV = 4.1%). Magnetic separation effectively minimizes matrix interference, making the assay suitable for complex biological specimens such as intestinal contents and fecal samples. This dual-mode design integrates fluorescence for precise quantification and colorimetry for equipment-free visual detection, holding great promise for point-of-care testing, early risk assessment of lead-induced gut microbiota dysbiosis, and environmental monitoring.
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高影响因子
ANALYST
·
2022
·Vol. 147
·期 24
·5623-5632
·
IF 5.227
Jiahai Wang,Le Luo,Yunhui Li,Lanfang Chen,Cenlin Gui,Jianji Zhu,Huizhen Li,Wenlong Wang,Daqi Chen
Solid-state nanopores have been proven as a powerful platform for label-free single-molecule analysis. However, due to its relatively low resolution and selectivity, developing biosensors with good translocation signals faces two significant challenges: (1) small-sized chemical or biological targets show difficulty in producing recognizable translocation signals because of their weak interaction with the nanopore and (2) protein interferents that widely exist in biological samples or buffers would considerably deteriorate the noise level of the nanopore, submerging the translocation signal. Herein, we demonstrate an effective way to overcome both the challenges. DNA cubes were used as signal transducers that can achieve an ultra-high (>50 : 1) signal-to-noise ratio (SNR) translocation signal, which is maintained even in protein interferent-rich buffers. A sensing strategy was constructed via hepatitis B virus (HBV) target-triggered cleavage of the component elements of the DNA cube with the assistance of the CRISPR–Cas12a technology, which caused a great drop in the translocation rate. The elements to cleave were optimized, and the sensor performance was tested in different protein stabilizer-rich buffers and human serum. Coupling with the polymerase chain reaction (PCR) pre-amplification technology, HBV-positive or -negative classification was achieved with the detection limit reaching 5 aM. It is worth noting that in our method, all reaction buffers were directly used without further optimization, which is of great help for the practical application of solid-state nanopores.
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BIOSENSORS & BIOELECTRONICS
·
2026
·
IF 11.8
Lingsong Lu,Yingying Zhang,Bei Liu,Nuo zhou,Dao jun Yu,Ying Wang
Circular RNAs (circRNAs) represent an emerging family of noncoding transcripts defined by closed-loop architecture, which are now established as key participants in the etiology of tumorigenesis and malignant progression. While circRNAs show potential as therapeutic targets and biomarkers, the accurate detection of circRNAs remains challenging due to interference from homologous linear RNAs. In this study, an ultrasensitive method for detecting circ_0051240 based on the sulfhydrated toehold capture probe-initiator assembly-mediated hybridization chain reaction (HCR) and the CRISPR-Cas12a system was developed. The capture probe (with a toehold domain)-initiator strand duplex was ingeniously fabricated to identify the back-splice junction (BSJ) of circRNA. After magnetic enrichment and separation by the Fe 3 O 4 -Au@UIO-66-NH 2 nanocomposite, the initiator strand (H0) was delivered to trigger HCR. The HCR product dsDNA concatemers contained multiply repeated CRISPR-targetable DNA sites that were readily recognized by the CRISPR RNA (crRNA). This specific recognition and binding activated the CRISPR-Cas12a system's collateral endonuclease activity, leading to cleavage of the fluorophore-quencher (FQ) reporters and fluorescence emission at a characteristic wavelength. This design eliminated linear RNA-related interference and enhanced the detection of fluorescence intensity (FI). Under optimal conditions, the proposed HCR/CRISPR-Cas12a method exhibited a wide quantitative measurement range spanning from 45 pM up to 180 nM, achieving a notable limit of detection (LOD) of 0.03 pM. In this study, a novel circRNA sensing strategy capable of accurate and highly sensitive quantification of ovarian cancer-specific circRNA was reported. The proposed method exhibits acceptable performance when compared to present approaches.
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Analytical Methods
·
2026
·
IF 3
Yi Tian,Jiaxuan Chen,Fengzheng Chen,Yu Ling,Shengyue Zheng,Daizhen Jian,Qiong Tang,Song Wang,Kai Shi
Split crRNA with Cas12a (SCas12a)-powered biosensor has shown sensitivity and accuracy in nucleic acid detection, but its application to metal ion target detection still needs to be explored. By integrating an SCas12a-mediated cascade strand displacement reaction (CSDR) with a phosphorothioate-modified hairpin G-quadruplex (psHG4) reporter, a portable “turn-on” colorimetric/photothermal system was engineered for the precise and sensitive detection of Hg2+ (termed as SCas12a/CSDR-psHG4). Target Hg2+ triggers SCas12a-mediated CSDR, and then activates SCas12a’s nuclease activity. Activated SCas12a cleaves the loop region of psHG4, thereby liberating the previously silenced psG4 sequence, which subsequently assembles into a psG4/hemin complex in the presence of K⁺ and hemin. This complex catalyzes the typical oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB), generating the colorimetric and photothermal product TMBox. Record the RGB value of the solution with a portable smartphone and track the temperature change using a thermometer. This colorimetric/photothermal biosensor displayed a linear range of 1 pmol/L to 100 nmol/L and 100 fmol/L to 10 nmol/L, with a detection limit of 0.5 pmol/L and 60 fmol/L, respectively. Utilizing the SCas12a technique, the SCas12a/CSDR-psHG4 presents a fresh method for identifying metal ion targets, along with significant prospects for enhancing human health protection and environmental surveillance.
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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.
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ANALYTICAL CHEMISTRY
·
2023
·
IF 7.4
Peiqing Luo,Xiaocui Huang,Fang Luo,Zhonghui Chen,Yu Chen,Cuiying Lin,Jian Wang,Bin Qiu,Zhenyu Lin
In this work, combined with the high amplification efficiency of hybridization chain reaction (HCR), high specificity of the CRISPR/Cas12a system, and convenience of the homogeneous electrochemiluminescence (ECL) assay based on the regulation of negative charge on the reporting probes, a sensitive ECL biosensor for hepatitis B virus DNA (chosen as a model target) had been developed. The initiator chain trigger DNA that can induce HCR amplification is modified on the surface of ruthenium bipyridine-doped silica nanoparticles (Ru@SiO2 NPs) first, and large amounts of negative charges modified on the particles were achieved through the HCR amplification reaction. The efficiency of the nanoparticles reaching the negatively charged working electrode can be regulated and realize the change of the ECL signal. In addition, long DNA on the surface of the luminescent body may prevent the coreactant from entering the pore to react with ruthenium bipyridine. These factors combine to produce a low-background system. The presence of the target can activate the CRISPR/Cas12a system and make trigger DNA disappear from the nanoparticle surface, and strong ECL can be detected. The sensor does not require a complex electrode modification; therefore, it has better reproducibility. Additionally, due to dual signal amplification, the sensor has a high sensitivity. In the range of 10 fM to 10 nM, the ECL intensity exhibits a strong linear relationship with the logarithm of the target concentration, and the detection limit is 7.41 fM. This sensor has shown high accuracy in detecting clinical samples, which holds significant potential for application in clinical testing.
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Environmental Science-Nano
·
2021
·Vol. 9
·期 1
·162-172
·
IF 8.131
Kai Zhang,Zhenqiang Fan,Yuedi Ding,Sha Zhu,Minhao Xie,Nan Hao
In this paper, we present the first idea of using a DNA triple helix structure to inhibit CRISPR-Cas12a activity and apply it to the design of an electrochemiluminescent biosensor for the detection of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA-dependent RNA polymerase (RdRp) gene in real samples and environmental surveillance. We employed a segment from the RdRp gene of SARS-CoV-2 by an entropy-driven reaction, which was paired with double-stranded DNA that can activate CRISPR-Cas12a activity by Hoogsteen pairing to form triple-stranded DNA, thereby inhibiting the binding interaction of the double-stranded DNA with CRISPR-Cas12a, which in turn inhibits the trans cleavage activity of CRISPR-Cas12a. The inhibited CRISPR-Cas12a is unable to cut the nucleic acid modified on the electrode surface, resulting in the inability of the ferrocene (Fc) modified on the other end of the nucleic acid to move away from the electrode surface, and thus failing to cause electrochemiluminescence changes in GOAu–Ru modified on the electrode surface. The extent of the electrogenic chemiluminescence change can reflect the concentration of the gene to be tested. Using this system, we achieved the detection of the SARS-CoV-2 RdRp gene with a detection limit of 32.80 aM.
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