ACS Sensors
·
2019
·
IF 6.944
Liping Liu,Zhen Fang,Xiangjiang Zheng,Dongmei Xi
A straightforward yet efficient, nanopore-based strategy that enables the sensitive detection of copper(II) ion (Cu2+) and real-time monitoring of a click reaction is provided. Two single-stranded DNAs (ssDNAs) are designed to act as the preprobes, one being modified with an azide and the other an alkyne. The presence of Cu2+ induces the ligation of two ssDNAs via click reaction, leading to the formation of a forked DNA which can quantitatively generate characteristic current signatures when interacts with α-hemolysin (α-HL) nanopore. The assay facilitates a highly selective and sensitive measurement of Cu2+ without the need for labels or signal amplification. More importantly, this nanopore platform exhibits excellent performance in real-time monitoring of a copper(I) ion (Cu+)-catalyzed click reaction at the single-molecule level, by recording the current signals of the forked DNA generated by click chemistry. The proposed strategy is believed to play an important role in both nanopore sensing and characterization of chemistry reactions, especially coupling reactions.
研究领域:
纳米孔传感
·
化学反应表征
·
生物分析化学
·
分子生物学
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DOI
PubMed
高影响因子
TALANTA
·
2021
·
IF 6.057
Xianfeng Wang,Xiaolong Chen,Chengxiang Chu,Yuanyi Deng,Mei Yang,Danqun Huo,Faliang Xu,Changjun Hou,Jiayi Lv
Accurate and effective detection of single-stranded nucleic acids is vital in both disease diagnosis and pathological studies. Hence, we develop a PAMmer-assisted CRISPR/Cas9 system mediated G4-EXPAR (Cas-G4EX) strategy for site-specific detection of ssRNA and ssDNA . PAMmer-assisted CRISPR/Cas9 executes the site-specific cleavage of target ssRNA or ssDNA and released product fragment with the desired sequence at the 3′-terminal. This fragment serves as a primer to activate subsequent sequence-dependent exponential amplification reaction (EXPAR). The G-rich EXPAR products assembles with hemin to form a G-Quadruplex (G4/hemin). G4/hemin catalyzes ABTS-H 2 O 2 system with the appearance of vivid green color, realizing naked-eye analysis. Cas-G4EX integrates the superiority of CRISPR/Cas9 and EXPAR, presenting outstanding site-specific recognition and high-performance amplification efficiency. Meanwhile, the programmability of CRISPR/Cas9 system makes the proposed method become a universal detection paradigm for any ssRNA or ssDNA. Cas-G4EX assay shows the linear relationship from 250 aM to 2.5 nM for ssRNA detection with the actual LOD of 250 aM, and that ranges from 100 aM to 1 nM for ssDNA detection with the actual LOD of 100 aM. Additionally, the acceptable recoveries of 101.48%–109.61% for ssRNA and 93.25%–111.98% for ssDNA in real detection of human serum are obtained for detection of single-strand nucleic acid in real samples. Cas-G4EX also exhibits the excellent discrimination for single-base mutation of single-stranded nucleic acids. Therefore, Cas-G4EX assay provides a promising platform in the applications of molecular diagnosis and pathological analysis.
研究领域:
生物技术
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分子生物学
·
诊断医学
·
病理学
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产品关联
DOI
PubMed
高影响因子
MOLECULES
·
2018
·Vol. 23
·期 9
·2337
·
IF 3.098
Xixia Liu,Qi Lu,Sirui Chen,Fang Wang,Jianjun Hou,Zhenlin Xu,Chen Meng,Tianyuan Hu,Yaoyao Hou
We describe a multiple combined strategy to discover novel aptamers specific for clenbuterol (CBL). An immobilized ssDNA library was used for the selection of specific aptamers using the systematic evolution of ligands by exponential enrichment (SELEX). Progress was monitored using real-time quantitative PCR (Q-PCR), and the enriched library was sequenced by high-throughput sequencing. Candidate aptamers were picked and preliminarily identified using a gold nanoparticles (AuNPs) biosensor. Bioactive aptamers were characterized for affinity, circular dichroism (CD), specificity and sensitivity. The Q-PCR amplification curve increased and the retention rate was about 1% at the eighth round. Use of the AuNPs biosensor and CD analyses determined that six aptamers had binding activity. Affinity analysis showed that aptamer 47 had the highest affinity (Kd = 42.17 ± 8.98 nM) with no cross reactivity to CBL analogs. Indirect competitive enzyme linked aptamer assay (IC-ELAA) based on a 5′-biotin aptamer 47 indicated the limit of detection (LOD) was 0.18 ± 0.02 ng/L (n = 3), and it was used to detect pork samples with a mean recovery of 83.33–97.03%. This is the first report of a universal strategy including library fixation, Q-PCR monitoring, high-throughput sequencing, and AuNPs biosensor identification to select aptamers specific for small molecules.Keywords:clenbuterol;systematic evolution of ligands by exponential enrichment;real-time quantitative PCR;high-throughput sequencing technology;aptamers;gold nanoparticles biosensor
研究领域:
生物技术
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分析化学
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纳米技术
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分子生物学
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产品关联
DOI
PubMed
高影响因子
RSC Advances
·
2014
·Vol. 5
·期 8
·6100-6105
·
IF 3.708
Xue Gong,Wenjiao Zhou,Yaqin Chai,Yun Xiang,Ruo Yuan
Small molecule/protein interactions have a key role in drug discovery, clinic diagnosis and protein–metabolite interactions in biology. By using the specific interaction between folic acid (FA) and folate receptor (FR) as a model, the development of a label-free and sensitive colorimetric approach for the detection of the FR biomarker is described. The sensing approach relies on the coupling of the FR-induced terminal protection of FA-linked ssDNA strategy with significant signal amplification by self-assembled DNAzyme polymers. The FR binds to the FA-ssDNA and protects the FR/FA-ssDNA from digesting by exonuclease I. The terminal protected ssDNA further triggers autonomous self-assembly of two G-quadruplex sequence-containing hairpin DNAs into DNAzyme polymers, which result in intensified color change of the probe solution for label-free and highly sensitive colorimetric detection of FR. The terminal protection mechanism and the self-assembly formation of the DNAzyme polymers are characterized by using polyacrylamide gel electrophoresis, and the sensing parameters are optimized as well. Under optimal experimental conditions, the detection limit of 0.35 pM for FR can be obtained by using a UV-Vis spectrophotometer and the presence of as low as 5 pM of FR can be directly visualized by the naked eye. The developed method is also selective and can be applied to detect FR in serum samples, which makes this approach a sensitive platform for sensing different types of small molecule/protein interactions.
研究领域:
生物标志物检测
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药物发现
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临床诊断
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分子生物学
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纳米技术
✓ 已验证
产品关联
DOI
高影响因子
NEW JOURNAL OF CHEMISTRY
·
2018
·Vol. 42
·期 4
·2850-2856
·
IF 3.201
Yu Su,Huaguo Xu,Yonghui Chen,Jinxia Qi,Xiang Zhou,Renshan Ge,Zhenkun Lin
A real-time and label-free strategy for bisphenol A (BPA) detection was developed using an ssDNA aptamer modified sensor combined with dual polarization interferometry (DPI). The BPA-specific aptamer, which played the role of a specific probe for recognizing and binding the target BPA, was immobilized on the surface of a sensor chip through biotin–avidin interactions. Conformational changes on the sensor surface when binding the target were monitored in real time by DPI, so that data on mass, thickness and density were obtained. Our results verified that the developed sensor had good specificity for the target, as the changes in mass, thickness and density induced by BPA were much more significant than those induced by an analogue. It was found that, with the increase of BPA concentration, the mass and thickness on the sensor surface increased while the density decreased. Both mass and thickness changes on the sensor surface showed good linearity with the BPA concentration in the range of 1–20 μM, with correlation coefficients (R2) of 0.9946 and 0.9950, respectively. This work provides a demonstration for the first time that DPI is a reliable and efficient platform for real-time and label-free detection of organic pollutants and toxicants.
研究领域:
分析化学
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环境科学
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生物技术
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产品关联
DOI
高影响因子
LUMINESCENCE
·
2022
·Vol. 37
·期 6
·980-986
·
IF 2.613
Weiwei Luo,Zhuoxin Ye,Daqian Song,Pinyi Ma
In this work, an electrochemiluminescence (ECL) sensor chip for sensitive detection of thrombin (TB) was prepared using a screen-printed electrode (SPE) as a working electrode and an aptamer as a specific recognition moiety. To produce an ECL sensor chip, a layer of p L -Cys was immobilized on the surface of the SPE using the cyclic voltammetry scanning method. A layer of gold nanoparticles (AuNPs) was assembled through an Au–S bond and hairpin DNA was further immobilized on the electrode surface. Ru(bpy) 2 (mcpbpy) 2+ , as a luminescent reagent, was covalently bound to single-stranded DNA (ssDNA) to prepare a luminescence probe ssDNA-Ru. The probe was hybridized with TB aptamer to form a capture probe. In the presence of TB, the TB aptamer in the capture probe bound to TB, causing the release of ssDNA-Ru that could bind to hairpin DNA on the electrode surface. The Ru(II) complex as a luminescent reagent was assembled onto the electrode, and p L -Cys was used as a co-reactant to enhance the ECL efficiency. The ECL signal of the sensor chip generated based on the above principles had a linear relationship with log TB concentration at the range 10 fM to1 nM, and the detection limit was 0.2 fM. Finally, TB detection using this method was verified using real blood samples. This work provides a new method using an aptamer as a foundation and SPE as a material for the detection of biological substances.
研究领域:
分析化学
·
生物传感器
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纳米技术
·
生物化学
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产品关联
DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2024
·
IF 7.7
Xiuxiu Wu,Mengyuan Huang,Tai Ye,Long Bai,Rui Zhao,Ya Wang,Liling Hao,Min Yuan,Fei Xu
Aptamer conformations are susceptible to environmental conditions, which makes it difficult to achieve stable targets detection in complex environments with aptasensors. Imprinting strategy was proposed to immobilize the specific conformation of aptamers, aiming to enhance their recognition anti-interference. However, it is mechanistically unclear how the imprinted polymers affect aptamers' recognition, which limits application of the strategy. Herein, MD simulation was applied to explore the structural reason why nanogel imprinting improves binding affinity and selectivity of T30695 aptamer to Pb 2+ observed experimentally. Results show the imprinted polymers stabilize the domain-limited T30695 by noncovalent interactions. The coating process undergoes three evolution stages, finally achieving a polymer-aptamer-polymer sandwich-shaped conformation. Notably, it was found the polymers provide additional non-specific binding of Pb 2+ at acylamine group of acrylamide monomers, which accounts for the improved binding affinity with association constant K a 2.5 times larger. More importantly, imprinting enhances selectivity of aptamer to Pb 2+ by changing coordination mode of interfering ions (Ca 2+ , K + , Mg 2+ , NH 4 + and Cu 2+ ), which significantly destroys G-quadruplex conformation and thus its binding ability. This work revealed mechanistic effects of imprinting strategy on aptamers recognition at molecular level, which can guide rational design of high-performance aptamer-based biosensors applied in various detection areas.
研究领域:
生物化学
·
分子生物学
·
纳米技术
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产品关联
DOI
PubMed
高影响因子
FOOD CHEMISTRY
·
2021
·
IF 7.514
Zhijun Liu,Li Wang,Pengfei Liu,Kairen Zhao,Shuying Ye,Guoxi Liang
Hydrogen peroxide (H 2 O 2 ) is usually used as a fungicide in food, it is carcinogenic, accelerates aging or inducing toxic effects such as cardiovascular disease. Herein, to meet the demand for effective and fast detection of H 2 O 2 in food, a novel non-enzymatic electrochemiluminescence (ECL) sensor based on single-stranded DNA (ssDNA)/g-C 3 N 4 nanosheets (NS) was established. The ssDNA/g-C 3 N 4 NS hybrid was prepared by simple mixing g-C 3 N 4 NS and ssDNA solution together. The prepared ssDNA/g-C 3 N 4 NS exhibited improved peroxidase-like activity and was modified on a glassy carbon electrode to catalyze the ECL reaction of luminol-H 2 O 2 to amplify the luminescence signal. Under the optimized conditions, the proposed sensor exhibits high sensitivity with a limit of detection (LOD) as low as 33 aM H 2 O 2 , which is much lower than the vast majority of reported methods. This method enables the reliable responding to H 2 O 2 from the milk samples within 1 min.
研究领域:
分析化学
·
食品科学
·
纳米技术
·
生物化学
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产品关联
DOI
PubMed
高影响因子
MICROCHEMICAL JOURNAL
·
2025
·
IF 5.1
Yongshu Li,Yani Li,Qin Wu,Jianjun Hou,Xixia Liu
As a promising recognition molecule, aptamers are widely used in the field of small molecule detection. However, few reports are available on the key factors governing successful aptamer selection. To improve the success rate, the melamine-specific aptamer served as a model to investigate critical factors in library-immobilization-based SELEX methods (GO-SELEX and Beads-SELEX). The results indicated that non-ionic surfactants significantly enhanced the desorption of ssDNA from the graphene oxide (GO) surface in Mode one of the mimic GO-SELEX (GO-facilitated library immobilization and target-induced aptamer release). In contrast, in Mode two of the mimic GO-SELEX, where GO was used solely for non-target-binding aptamer removal, and significant non-specific binding between ssDNA and GO compromised the enrichment efficiency of target-specific sequences, necessitating the use of high target concentrations during SELEX. Similarly, in Beads-SELEX, high target concentrations were required to release specific ssDNA, which also led to a small amount of non-specific ssDNA co-release during the procedures. Other SELEX conditions were also compared and constructive suggestions were provided for facilitating the successful aptamer selection. In summary, this study provided SELEX condition optimization guidance for the selection of aptamers specific to small molecules, when different library-immobilized SELEX methods were performed.
研究领域:
生物化学
·
分子生物学
·
分析化学
·
生物技术
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产品关联
DOI
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2016
·
IF 4.758
Huayun Chen,Yuangen Wu,Wenping Yang,Shenshan Zhan,Shuyi Qiu,Pei Zhou
This paper reports a new assay platform that performed in polar organic solutions for ultrasensitive and highly selective detection of isocarbophos pesticide. The sensing system just contains hemin molecules and random ssDNAs. Free hemin molecules have a strong absorption peak at Soret band in polar organic solvents, and they can assemble into few small particles with random ssDNAs via their special π–π stacking interaction. However, the subsequent addition of isocarbophos into above sensing system can aggregate hemin molecules to form many larger particles, which result in a great decrease in absorbance value of sensing solution. UV/Vis absorption spectra and scanning probe microscope (SPM) were used to characterize hemin aggregation. Additionally, the effects of some parameters like type of solvents, hemin amount and random ssDNAs on the sensing platform were also investigated. The present assay platform has a detection limit as low as 0.2 μg/L, a dynamic range of 0.5–500 μg/L, and high selectivity over other competitive pesticides. Moreover, the assay platform also displays excellent stability and accuracy in the detection of spiked real samples, suggesting that such sensing platform will play an important role in biological and environmental detection.
研究领域:
分析化学
·
环境科学
·
生物化学
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产品关联
DOI
高影响因子
Science China-Chemistry
·
2016
·Vol. 59
·期 6
·770-775
·
IF 2.429
Wang Min,Li Fangzhen,Jiang Bingying,Xu Junqiang,Li Shuo,Xiang Yun,Yuan Ruo
The detection of protein/small molecule interactions plays important roles in drug discovery and protein/metabolite interactions in biology. In this work, by coupling the terminal protection of small molecule-linked ssDNA strategy with the unmodified and positively charged gold nanoparticle ((+)AuNP) nanoprobes, we have developed a sensitive and simple colorimetric sensor for the detection of folate receptor, a highly expressed protein in many kinds of malignant tumors. The target folate receptor binds the folate moieties of the folate-linked ssDNA through high affinity interactions and protects the protein-bound ssDNA from digestion by exonuclease I. The protected ssDNA thus adsorbs the ((+)AuNP) through electrostatic interactions, leading to a red-to-blue color change of the sensing solution for sensitive colorimetric detection of folate receptor at the sub-nanomolar level. Besides, this colorimetric sensor shows high selectivity toward folate receptor against other control proteins. The developed sensor avoids the modification/conjugation of the AuNP nanoprobes and the involvement of any expensive instruments for signal transduction in protein detection. Featured with these obvious advantages, the colorimetric sensor strategy demonstrated herein can be easily expanded for sensitive and convenient detection of various protein/small molecule interactions.
研究领域:
分析化学
·
生物传感器
·
癌症研究
·
纳米技术
✓ 已验证
DOI
高影响因子
Journal of Materials Chemistry B
·
2018
·Vol. 6
·期 46
·7614-7620
·
IF 4.776
Caihong Wang,Gonge Tang,Hongliang Tan
Nanomaterial/aptamer assembly has been extensively explored in the detection of various targets, but some limitations still exist in its practical applications, especially time consumption and low-efficient detachment of the aptamer from the nanomaterial surface. In this work, we demonstrated the ligand role of pyrophosphate ion (PPi) in the competitive displacement of ssDNA from the nanoscaffold surface. For this purpose, a fluorescein-labeled ssDNA (F-DNA) and a mixed valence state cerium (Ce3+/Ce4+)-based MOF (MVCM) were employed as the signal response unit and nanoscaffold, respectively. Benefiting from the existence of Ce4+, the MVCM exhibits an ultrahigh quenching efficiency (more than 90%) to F-DNA fluorescence, which is 3-fold higher than that of the MOF with Ce3+ only. However, it was found that PPi can effectively suppress the quenching effect of the MVCM by competitive coordination with the MVCM to displace F-DNA. Different from the conventional target-induced conformational change of aptamers, the PPi-triggered displacement assay is independent of the ssDNA sequence and can be rapidly completed in just 2 min. The displacement assay is also highly sensitive, even at a PPi concentration as low as 55 nM. In contrast to PPi, however, the phosphate ions and other anions cannot displace F-DNA from the MVCM surface to switch on the F-DNA fluorescence. Inspired by this fact, the PPi-triggered displacement assay was further applied in the detection of alkaline phosphatase (ALP). The detection limit toward ALP was obtained at 0.18 mU mL−1. Moreover, the accurate determination of ALP concentration in serum samples indicates the applicability of this sensing system in detecting real samples.
研究领域:
分析化学
·
纳米技术
·
生物传感器
✓ 已验证
产品关联
DOI
PubMed
高影响因子
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