ANALYTICA CHIMICA ACTA
·
2023
·
IF 6.2
Yuwan Du,Shuaifeng Ji,Qingyang Dong,Jiang Wang,Dianpeng Han,Zhixian Gao
Nucleic acid markers have been widely used in the detection of various virus-related diseases, including hepatitis B virus (HBV), which is spreading worldwide. The trans -activated CRISPR-Cas system has shown excellent sensitivity and specificity in nucleic acid detection. However, nucleic acid testing usually requires amplification of the target nucleic acid for more accurate and specific detection; furthermore, current nucleic acid assays are time-consuming, costly, and are limited by non-specific cross-reactivity. We developed an amplification-free viral DNA biosensor-based diagnostic method that uses a clustered regularly interspaced short palindromic repeats-associated system (CRISPR/Cas)-based approach with surface enhanced Raman spectroscopy . This method can specifically identify the target site by changing the crRNA sequence. In addition, the incubation period and development of the disease can be determined by quantitative detection of viral DNA. This system could achieve rapid and highly sensitive detection of HBV DNA within 50 min and vast detection range from 0.1 pM to 1 nM. Therefore, a combined CRISPR/Cas12a-SERS-based assay would improve the sensitivity of detection in assays using multiple biomarkers. In conclusion, our CRISPR/Cas12a-based biosensor would enable rapid, simple, and sensitive detection of HBV nucleic acids.
研究领域:
病毒学
·
分子诊断
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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
IMMUNITY
·
2023
·
IF 32.4
Lina Wang,Siru Li,Kai Wang,Na Wang,Qiaoling Liu,Zhen Sun,Li Wang,Lulu Wang,Quentin Liu,Chengli Song,Qingkai Yang
Self-nonself discrimination is vital for the immune system to mount responses against pathogens while maintaining tolerance toward the host and innocuous commensals during homeostasis. Here, we investigated how indiscriminate DNA sensors, such as cyclic GMP-AMP synthase (cGAS), make this self-nonself distinction. Screening of a small-molecule library revealed that spermine, a well-known DNA condenser associated with viral DNA, markedly elevates cGAS activation. Mechanistically, spermine condenses DNA to enhance and stabilize cGAS-DNA binding, optimizing cGAS and downstream antiviral signaling. Spermine promotes condensation of viral, but not host nucleosome, DNA. Deletion of viral DNA-associated spermine, by propagating virus in spermine-deficient cells, reduced cGAS activation. Spermine depletion subsequently attenuated cGAS-mediated antiviral and anticancer immunity. Collectively, our results reveal a pathogenic DNA-associated molecular pattern that facilitates nonself recognition, linking metabolism and pathogen recognition.
研究领域:
免疫学
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分子生物学
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病毒学
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癌症研究
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DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
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2013
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IF 5.437
Lichun Zheng,Xiaoyan Li,Panpan Liu,Guofan Wu,Xiaoquan Lu,Xiuhui Liu
We present a novel strategy for simultaneous electrochemical detection of multiple DNA targets based on the use of different encoding metal ions as tags. The principle of this scheme is that metal ions bound to metallothionein (MT) molecules can be released down after hybridization with DNA targets and then be detected by stripping voltammetry . The novel detection probes, ssDNA/MT conjugates , covered with different metal ions were synthesized for the first time, then three encoding metal ions (Zn 2+ , Cd 2+ , and Pb 2+ ) were used to differentiate the signals of three virus DNA due to their well-defined anodic stripping peaks at −1.13 V (Zn), −0.78 V (Cd), and −0.52 V (Pb) at BiFE , respectively. The anodic peak currents increased linearly with the concentrations of DNA targets in the range from 0.1 nM to 10 nM with a detection limit of 33 pM. In addition, the one-base mismatched target was effectively discriminated from the complementary target. The described results demonstrated that this method possesses high sensitivity and selectivity for multi-target DNA assay and has great potential in applications for detection of even more targets in biological assays , particularly immunoassays .
研究领域:
生物化学
·
分子生物学
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DOI
PubMed
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2022
·
IF 9.221
Yu Tao,Ke Yi,Haixia Wang,Kai Li,Mingqiang Li
Since hepatitis B virus (HBV) infection can cause chronic viral hepatitis, increase the risk of liver fibrosis and cirrhosis, and ultimately result in hepatoma, it is of great significance to explore convenient, rapid, and sensitive diagnostic strategies for HBV DNA detection. In this study, we developed a novel and sensitive fluorescent biosensor for HBV detection based on the sensing system consisting of CRISPR-Cas12a enzymes and metal nanoclusters as the luminescent nanoprobes, involving gold nanoclusters (AuNCs), silver nanoclusters (AgNCs), and copper nanoclusters (CuNCs). The DNA probes could effectively be applied as the template for the preparation of fluorescent metal nanoclusters. In the presence of the HBV target, the trans-cleavage capability of Cas12a was able to be initiated to degrade DNA probes, which brought about the inhibition of metal nanoclusters formation, thereby giving minimal fluorescence signals. In comparison with AuNCs and AgNCs, the CuNCs-based nanosensor exhibited much higher sensitivity. In addition, the fluorescence response of CuNCs to HBV detection was significantly faster, which could be completed within 25 min without the requirements of sophisticated equipment or harsh reaction conditions. Moreover, the practical application of this analysis method was also demonstrated to detect HBV DNA target in human serum samples with satisfactory recovery, which was promising for simple, rapid, sensitive, and label-free HBV DNA bioanalysis .
研究领域:
分子生物学
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医学诊断
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纳米技术
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产品关联
DOI
高影响因子
ELECTROANALYSIS
·
2023
·Vol. 35
·期 9
·e202300052
·
IF 3
Qiaoyu Ding,Xi Bai,Jiali Ju,Jiajun Shi,Guixin Li
A sensitive electrochemiluminescence (ECL) biosensor for the specific DNA sequence of hepatitis C virus (HCV) was developed based on the efficient quenching effect of the ferrocene cluster functionalized gold nanoparticles (Fc@AuNPs) on the ECL of electrodeposited silica@Ru(bpy) 3 2+ -chitosan-graphene oxide nanocomposite (SiO 2 @Ru−CS−GO). Graphene oxide (GO) can accelerate electron transfer rate, thus improving the ECL of Ru(bpy) 3 2+ on electrode surface. The molecular beacons (MB) was fixed to SiO 2 @Ru−CS−GO by glutaraldehyde (GA) using the Schiff reaction between amino groups of chitosan (CS) and MB. The ECL of SiO 2 @Ru−CS−GO was depressed greatly by the Fc@AuNPs labelled at the end of MB, then, a stronger ECL was observed when the distance between Fc@AuNPs and SiO 2 @Ru−CS−GO increased after the hybridization of target DNA with MB. Under optimum conditions, the restored ECL intensity increased linearly with the target DNA concentration in the range of 1.0×10 −16 ∼1.0×10 −10 mol ⋅ L −1 , and the limit of detection (LOD) is 1.4×10 −17 mol ⋅ L −1 . The proposed method exhibits acceptable stability and reproducibility. In general, the constructed HCV biosensor can be used for the sensitive detection of HCV in human serum, suggesting potential application prospects in bioanalysis.
研究领域:
分析化学
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生物传感器
·
医学检测
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DOI
高影响因子
TALANTA
·
2018
·
IF 4.244
Li Zou,Tingting Li,Ruidi Shen,Shaozhi Ren,Liansheng Ling
A label-free light-up fluorescent sensing strategy using hybridization chain reaction (HCR) amplification and DNA triplex assembly has been developed. Remarkably, the proposed fluorescence assay is successfully applied to the determination of avian influenza A (H7N9) virus DNA and thrombin. Herein, in the presence of targets, the target DNA/initiator triggers a cascade of hybridization events between H1 and H2 that yields nicked double helices analogous to alternating copolymers . With the additions of triplex-forming oligonucleotide (TFO) and berberine , the triplex structures form between HCR products and TFO. Then, a large amount of berberine can bind to the triplex structures and the sensing system exhibits a dramatic increase in the fluorescence intensity at 530 nm. Under optimal conditions, the label-free fluorescent sensing platform shows sensitive responses to H7N9 virus DNA and thrombin in the range of 0.2–100 nM and 0.5–200 nM, respectively. The detection limits of H7N9 virus DNA and thrombin are as low as 0.14 nM and 0.32 nM, respectively. Owing to the simplicity and low cost, the proposed strategy can be used in other biomarkers assays, providing a promising tool for clinical diagnosis and biomedical detection.
研究领域:
生物化学
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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
PubMed
高影响因子
TALANTA
·
2017
·
IF 4.162
Lu Ding,Chunlan Xiang,Gang Zhou
Poly(acrylic acid) (PAA) brushes coated onto silica nanoparticles have been widely utilized in bioassays due to their abilities of providing favorable microenvironments and ensuring good biological activities for biomolecules. However, traditional PAA brushes are synthesized by reversible addition–fragmentation chain transfer polymerization. Hence, it is generally difficult to control and characterize the molecular weight of the PAA brushes, which may depress the reproducibility and bring more uncertain results. Herein, atom transfer radical polymerization method is employed to synthesize β -cyclodextrin-cored PAA with uniform and controllable molecular weight. After loading on the surfaces of adamantane-functionalized silica nanoparticles via host-guest interactions, glucose oxidase and probe single strand DNA (ssDNA) are further immobilized on the as-prepared nanoparticles. Meanwhile, capture ssDNA is functionalized on amino modified magnetic beads. In the presence of ssDNA sequence of Hepatitis B Virus (HBV) containing completely matched sequence of both probe and capture ssDNA, a bioconjugate is formed and can be separated by an external magnet. The isolated glucose oxidase can further catalyze glucose into gluconic acid and H 2 O 2 , and then reduce HAuCl 4 on Au seeds. By monitoring the absorption intensity change of the Au NPs at 530 nm, the proposed biosensor with novel signal amplification probes can be used to detect DNA sequence of HBV with high sensitivity and selectivity in both buffer and serum samples. This developed strategy has presented a new way to construct silica nanoparticles coated by PAA brushes for the fields of clinical diagnosis and other life sciences.
研究领域:
生物化学
·
纳米医学
·
临床诊断
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产品关联
DOI
PubMed
高影响因子
TALANTA
·
2019
·
IF 4.916
Tian Wu,Xiang Li,Yuanqi Fu,Xuelian Ding,Zhongjian Li,Guifen Zhu,Jing Fan
Developing the high selectivity and sensitivity strategy for nucleic acid detection is crucial for early diagnosis and therapy of diseases. In this work, a novel low back-ground fluorescent sensor platform for the detection of nucleic acid has been developed based on δ-FeOOH nanosheets integrating with exonuclease III-assisted target-recycling signal amplification. Because of the strong binding ability between the single-strand DNA (ssDNA) and the δ-FeOOH nanosheets, the dye-labeled ssDNA probe would be quenched by δ-FeOOH nanosheets through fluorescence resonance energy transfer (FRET). By using magnetic separate properties of δ-FeOOH, the background signal was separated from the sensor system, and the low background sensor system was obtained. After adding the target DNA, a double-strand DNA complex (dsDNA) would be formed between the target DNA and dye-labeled ssDNA probe. Then, the dye-labeled ssDNA probe in the dsDNA complex would be stepwise hydrolyzed into short fragments from 3′-terminus by Exonuclease III, and the fluorescence signal was recovered due to the weak bind affinity between the short fragments and δ-FeOOH nanosheets. By using the fluorescence quenching ability of δ-FeOOH nanosheets and enzyme-assisted target-recycling signal amplification, this strategy could show an excellent selectivity toward hepatitis C virus DNA with a low detection limit of 10 pM. By simply changing the dye-labeled ssDNA probe sequence, this sensing platform can be developed as a universal approach for the simple, sensitive, and selective detection of different target DNA.
研究领域:
生物技术
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分子生物学
·
分析化学
·
病毒学
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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
·
病毒学
·
纳米技术
·
分子诊断学
·
CRISPR技术
·
分析化学
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产品关联
DOI
PubMed
高影响因子
Science China-Chemistry
·
2025
·
IF 9.7
Lei Chao,Fan Wenjiao,Ren Wei,Duan Xinrui,Liu Chenghui
We propose a novel single microbead “chemiluminescence (CL) super enriching and imaging” strategy that efficiently concentrates highly intense, long-lasting, and ultrastable CL shining on only a single microbead (SMB), allowing for the ultrasensitive quantification of various biomarkers at the attomolar level. Aided by a phenothiazine derivative enhancer, the traditional flash-type CL of horseradish peroxidase (HRP)-H2O2-luminol can be efficiently converted into a significantly enhanced (103-fold), persistent, and ultrastable CL emission (E-CL) that can be exactly confined and accurately imaged on the microbead interface. Then, by employing only one minuscule-sized SMB as the sole reaction and signaling unit, the target as well as the aroused E-CL signal concentrated on the SMB can further achieve ~104 “super-enriching” efficiency. Therefore, by “super enriching” target-responsive E-CL signal on an SMB, the SMB E-CL imaging strategy provides a powerful tool for the attomolar-level detection of various biomarkers including proteins, microRNAs, and virus DNA.
研究领域:
分析化学
·
生物化学
·
生物传感器
·
纳米技术
·
分子诊断
·
生物发光
·
医学诊断
✓ 已验证
产品关联
DOI
高影响因子
ANALYST
·
2022
·Vol. 147
·期 21
·4876-4887
·
IF 5.227
Zengming Zhang,Shuhao Zhao,Lei Jiang,Junjun Wu,Wenhan Zhao,Xiaoniu Guo,Niancai Peng,Fei Hu
As the gold standard for nucleic acid detection, full-process polymerase chain reaction (PCR) analysis often falls into the dilemma of complex workflow, time-consuming, and high equipment costs. Therefore, we designed and optimized a DNA quantification microfluidic system by strategically integrating sample pretreatment and a smartphone-readable gradient plasmonic photothermal (GPPT) continuous-flow PCR (CF-PCR). Through preloading and sequential injection of immiscible extraction reagents, combined with magnetic bead (MB) manipulation, the microfluidic chip successfully purified and concentrated 100 μL of HBV-DNA spiked plasma into a 20-μL purified sample within 14 minutes. With a digital PCR platform, the optimized experiments showed that the DNA extraction efficiency can reach 69% at an immiscible reagent configuration ratio of 10 : 10 : 1 : 12 : 2 (sample : lysis/binding buffer : MB : silicone oil : eluent) and a flow rate of 25 μL min−1. For the first time, we used gold nanorod (AuNR)-doped PDMS to prepare a CF-PCR submodule for the amplification of a 40 μL PCR mixture. Due to the plasmonic photothermal effect of AuNRs and the gradient intensity of an expanded laser spot, the PCR thermal gradient was formed on a coin-sized area. The compact annular thermal-microfluidic layout, optimized DNA dye concentration, and chip transmittance synergistically enable a rarely reported smartphone-based fluorescence CF-PCR, greatly simplifying thermal control and detection setup. Prototype construction and validation experiments show that the microsystem can complete the sample-to-answer quantification of HBV-DNA with a dynamic linear range from 1.2 × 101 to 1.2 × 106 copies per μL in ∼37 minutes. This novel microfluidic solution effectively bridges the technical gap between the CF-PCR, sample pretreatment and result characterization, making the workflow standardized and rapid and requiring <15% of the commercial instrument cost. The simplicity, rapidity and low cost of this work make it promising for applications in decentralized laboratories and low-resource settings.
研究领域:
生物医学工程
·
分子诊断
·
微流控技术
·
纳米技术
·
即时检验
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DOI
PubMed
高影响因子
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
·
2018
·Vol. 411
·期 19
·4569-4576
·
IF 3.307
Wang Kun,Zhai Fu-Heng,He Meng-Qi,Wang Jin,Yu Yong-Liang,He Rong-Huan
A simple fluorescence biosensor is developed based on the enzyme-assisted cascade amplification strategy. The amplification system consists of a hairpin-structure DNA (H-DNA) and exonuclease III. The target DNA can hybridize with the H-DNA and initiate exonuclease III-assisted target recycling amplification to generate abundant G-rich DNA (G-DNA). One region of G-DNA is designed to possess the same sequence as target DNA. Thus, the G-DNA can also hybridize with H-DNA and initiate the digestion of H-DNA. The cascade strategy in this amplification system causes the concentration of G-DNA to grow exponentially. The fluorescence intensity of N -methylmesoporphyrin IX (NMM) is highly enhanced due to the formation of G-quadruplex configuration. Under optimal conditions, the cascade system could achieve an admirable sensitivity with a detection limit of 52 fM for HIV DNA, and guarantees a satisfactory specificity. Moreover, the cascade system could be implemented for other target DNA detections by substituting the recognition region of the H-DNA. In this way, a detection limit of 65 fM for HBV DNA could be achieved by the cascade system. The target DNA analysis in a real serum sample further indicates that this biosensor has potential for future application in clinical diagnosis.
A simple and label-free cascade amplification strategy is developed by exploiting hairpin DNA and EXO III for sensitive DNA detection.
研究领域:
生物化学
·
分子生物学
·
分析化学
·
临床诊断
✓ 已验证
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）