ANALYST
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2017
·Vol. 142
·期 11
·2013-2019
·
IF 3.885
Hui Zhang,Zhihui Guo,Huilei Dong,Hongfei Chen,Chenxin Cai
In this work, a highly sensitive electrochemiluminescence (ECL) assay was fabricated for the detection of human DNA (cytosine-5)-methyltransferase1 (DNMT1) activity in cancer cells. The ECL assay coupled hybridization chain reaction with a G-quadruplex/hemin DNAzyme biosensing strategy. The ECL intensity changes (ΔI) allowed detection of DNMT1 activity down to 0.09 U mL−1, and ΔI was proportional to the logarithm of the activity of DNMT1 within the range of 1.0 to 30.0 U mL−1 in buffer solution. It also showed high sensitivity to DNMT1 activity in A549 cells, with a detection limit of about 2 cells. This ECL assay provides a promising platform for profiling of the mutational cells of tumors and shows a great potential for application to DNA methylation-related clinical diagnostics.
研究领域:
生物化学
·
分子生物学
·
临床诊断学
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高影响因子
Food Analytical Methods
·
2014
·Vol. 8
·期 6
·1556-1564
·
IF 1.802
Liu Tao,Zhang Xiao,Hao Jianpeng,Zhu Wenxin,Liu Wei,Zhang Daohong,Wang Jianlong
Herein, we for the first time report an acetylcholinesterase (AChE)-free colorimetric method for the detection of chlorpyrifos (CP), which is based on the oxidation reaction of H 2 O 2 with CP and 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS 2− ) catalyzed by hemin/G-quadruplex DNAzyme. In the absence of CP, ABTS 2− is directly oxidized by H 2 O 2 with hemin/G-quadruplex DNAzyme as a biocatalyst to produce the blue-green-colored free radical anion (ABTS ·− ), accompanied with an obvious UV-visible absorbance at 418 nm. However, the amount of H 2 O 2 used in H 2 O 2 -DNAzyme-ABTS 2− system will be decreased if CP is first reacted with H 2 O 2 , leading to the decrease of the amount of ABTS ·− and a change in the UV-visible absorbance of ABTS ·− at 418 nm. Thus, an indirect dependence relation between CP concentration and the absorbance of ABTS ·− at 418 nm could be constructed and used for CP detection. Under optimal conditions, the detection range of the method is 0.04–1 μg/mL, with a detection limit of 0.01 μg/mL. Application of the method in real juice samples shows good analytical features in terms of recoveries (95–107.5 %), the relative standard deviation (RSD) (0.6–4.7 %), rendering it as an attractive candidate to current methodologies for the determination of CP.
研究领域:
分析化学
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环境科学
·
食品安全
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高影响因子
SCIENCE OF THE TOTAL ENVIRONMENT
·
2021
·
IF 7.963
Nuanfei Zhu,Xuesong Li,Ye Liu,Jingfu Liu,Yawei Wang,Xiangyang Wu,Zhen Zhang
Dibutyl phthalate (DBP) is considered as one of the most widely used phthalate esters (PAEs), which has attracted worldwide concerns because of its potential threats to eco-environments and human health. Systematic investigations of DBP environmental occurrence contribute to the further risk assessment, which depends on effective and available analytical methods. In this study, an amplified ratiometric fluorescence ELISA was established for sensitive and high-throughput detection of DBP in the aquatic system based on a novel tetrahedral DNA nanostructure (TDN)-scaffolded-DNAzyme (Tetrazyme). Wherein, Tetrazyme was prepared by the precise folding of G-quadruplex sequence on three vertex angles of the TDN, together with hemin as the horseradish peroxidase (HRP)-mimicking enzyme. The rigid TDN avoided the local overcrowding effect to provide a reasonable spatial spacing on the interface for G-quadruplex sequence, increasing the collision chance between DNAzyme and substrates, improving the catalytic ability of DNAzyme effectively. Besides, streptavidin (SA) and biotin (bio) were used to anchor TDN and antibody, in which the specific binding of SA/bio could make more Tetrazyme conjugate on each signal element, resulting in the dual signal amplification. Meanwhile, the accuracy and precision were enhanced owing to the inherent built-in rectification to the environment from the dual output ratiometric fluorescence assay. Under the optimized conditions, the detection limit of this proposed method was 0.17 ng/mL (16 times lower than that of conventional ELISA using the same antibody) with a satisfactory accuracy (recoveries, 79.0%- 116.2%; CV, 2.1–6.5%). Overall, this platform provides a promising way for accurate, sensitive and rapid determination of DBP from environmental waters.
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BIOMACROMOLECULES
·
2022
·
IF 6.978
Hao Sun,Haifeng Wu,Qiao Teng,Yuanxi Liu,Hui Wang,Zhen-Gang Wang
In enzymatic active sites, the essential functional groups are spatially arranged as a result of the enzyme three-dimensional folding, which leads to remarkable catalytic properties. We are inspired to self-assemble the polylysine peptides with guanine-rich DNA and hemin as cofactor to fabricate the peroxidase-mimicking catalytic nanomaterials. The DNA can fold into G-quadruplex to provide a supramolecular scaffold and a nucleobase for supporting and coordinating hemin, and the polylysine provides amine as distal groups to promote the H2O2 adsorption to the iron of hemin. The polylysine and DNA components synergistically accelerated the hemin-catalyzed reactions, and the complex containing ε-polylysine exhibited higher activity than α-polylysine. This activity difference is attributed to the higher pKa value and more susceptible protonation of amine of ε-polylysine than α-polylysine. The ε-polylysine/DNA/hemin had similar coordination states of hemin and conformations of the components to α-polylysine/DNA/hemin but accelerated the formation of the intermediate compound I faster than α-polylysine. Theoretical simulation reveals that the unprotonated NH2 behaved like a base catalyst, similar to His-42 residue in the natural heme pocket, while the protonated NH3+ acted as an acid, which indicated that the base catalyst on the distal side of the hemin pocket is more active than the acid. This work provides an avenue to control the distribution of the catalytic residues in an enzyme-like active site and to understand the roles of the key residues of native enzymes.
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SENSORS AND ACTUATORS B-CHEMICAL
·
2018
·
IF 5.667
Liyan Zhang,Chuan Fan,Min Liu,Fengjuan Liu,Shanshan Bian,Shuyue Du,Shuyun Zhu,Hua Wang
Catalytic hemin was first remolded with biomineralized gold to form Hemin-Au core and further encapsulated into the Tb metal organic framework (MOF) matrix. The so obtained Hemin-Au@MOF composites could possess high environmental stability and especially double catalysis activities of enhanced peroxidase-like and gold catalysis. Steady state kinetics studies indicate that Hemin-Au@MOF could present much higher intrinsic catalysis and substrate affinity (lower Michaelis constants) than native hemin and even natural peroxidase . Furthermore, the catalytic composites were employed for labeling antibodies towards the sandwiched immunoassays for alpha-fetoprotein (AFP) as a model biomarker of cancers. The results indicate that the Hemin-Au@MOF-based immunoassays could be conducted by two signal amplification ways of hemin-catalyzed chromogenic reaction and gold-catalyzed silver staining, enabling the detection of AFP in blood with the level down to 0.020 ng mL −1 . Importantly, the proposed fabrication protocol combining the gold biomineralization and MOF encapsulation routes may be extended for the fabrication of a variety of enzymes or enzymatic mimics with the improved environmental stability and multiple catalysis activities for the wide catalysis applications.
研究领域:
生物化学
·
医学诊断
·
材料科学
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高影响因子
MICROCHIMICA ACTA
·
2015
·Vol. 182
·期 15
·2607-2613
·
IF 3.741
Yang Zhiqing,Xie Liyan,Yin Huanshun,Zhou Yunlei,Ai Shiyun
We describe an electrochemical bioassay for the detection of the activity of methyltransferase (MTase), and for screening this enzyme’s inhibitors. The assay is based on the conjugation of a hemin to a G-quadruplex that enables enzymatic signal amplification with the aid of exonuclease III (ExoIII). In the first step, double-stranded DNA containing the quadruplex-forming oligomer is assembled on the surface of a gold electrode and then methylated by DNA adenine methyltransferase (DAM). After cleaved by endonuclease DpnI, the methylated DNA is digested by ExoIII and the quadruplex-forming oligomers are liberated. This leads to the formation of a hemin/G-quadruplex (in presence of hemin and of potassium ions). The hemin/G-quadruplex catalyzes the oxidization of hydroquinone by H 2 O 2 and the benzoquinone was formed to generate electrochemical signal. Finally, the gold electrode modified with reduced graphene oxide was used as working electrode for performing differential pulse voltammetry. The method has a detection limit of 0.31 unit · mL −1 . A study on the inhibition of MTase showed it was inhibited by epicatechin with an IC50 value of 157 μM.
We describe an electrochemical bioassay for the detection of the activity of methyltransferase and for screening for its inhibitors. Due to the conjugation of a hemin to a G-quadruplex, strong enzymatic signal amplification is enabled with the aid of exonuclease III.
研究领域:
生物化学
·
分子生物学
·
电化学分析
·
药物筛选
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高影响因子
Analytical Methods
·
2025
·
IF 2.6
Zheng Chang,Zhenyuan Duan,Junpeng Li,Xiangze Meng
In this study, hemin-functionalized polyethyleneimine/silica nanoparticles (hemin/PEI/SiO2 NPs) served as an electrochemical label for ultrasensitive detection of prostate-specific antigen (PSA), a clinically important prostate cancer biomarker. A poly(thionine)/carbon nanotube (PTh/CNTs) multilayer film was fabricated on a glassy carbon electrode (GCE) via integrated electropolymerization and electrostatic layer-by-layer (LbL) assembly, forming a {PTh/CNTs}2/PTh modified GCE with abundant amino groups. PSA capture antibodies (Ab1) were then covalently immobilized on the modified electrode through glutaraldehyde (GA) crosslinking, leveraging the film's surface-exposed amino groups. Furthermore, hemin/PEI/SiO2 NPs were synthesized through amine-carboxyl interactions between carboxyl groups of hemin and amine groups of PEI/SiO2 NPs. The immunosensor assembly employed a sandwich-type electrochemical platform using PSA detection antibody (Ab2)-conjugated hemin/PEI/SiO2 NPs. The specific antibody-antigen binding enables efficient electron transfer across the nanoscale spacing (several nanometers) between hemin/PEI/SiO2 NPs and the {PTh/CNTs}2/PTh multilayer film. In this architecture, CNTs serve as excellent electron mediators, significantly facilitating electron transport. Moreover, the high-density immobilization of hemin on PEI/SiO2 NPs significantly amplifies the electrochemical redox currents of the multilayer film through proximity effects. This enhanced electrical signal enables highly sensitive detection of target protein antigen–antibody binding events. Consequently, by correlating the enhanced reduction peak current of the immunosensor with PSA concentration, highly sensitive detection of PSA at various concentrations can be achieved. Under optimized conditions, it exhibited a linear response to logarithmic PSA concentrations (0.01 pg mL−1 to 100 pg mL−1, R = 0.9963), with an ultralow detection limit of 3.0 fg mL−1 (S/N = 3). Furthermore, the immunosensor demonstrated outstanding analytical performance in human serum samples (96.0–103% recovery, n = 3), confirming its high accuracy for clinical applications.
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