Chemosensors
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2022
·Vol. 10
·期 7
·276
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IF 4.229
Kunhan Nie,Yongjian Jiang,Na Wang,Yajun Wang,Di Li,Lei Zhan,Chengzhi Huang,Chunmei Li
The abnormal expression of miRNA is closely related to the occurrence of pancreatic cancer. Herein, a programmable DNAzyme amplifier for the universal detection of pancreatic cancer-related miRNAs was proposed based on its programmability through the rational design of sequences. The fluorescence signal recovery of the DNAzyme amplifier showed a good linear relationship with the concentration of miR-10b in the range of 10–60 nM, with a detection limit of 893 pM. At the same time, this method displayed a high selectivity for miR-10b, with a remarkable discrimination of a single nucleotide difference. Furthermore, this method was also successfully used to detect miR-21 in the range of 10–60 nM based on the programmability of the DNA amplifier, exhibiting the universal application feasibility of this design. Overall, the proposed programmable DNAzyme cycle amplifier strategy shows promising potential for the simple, rapid, and universal detection of pancreatic cancer-related miRNAs, which is significant for improving the accuracy of pancreatic cancer diagnosis.Keywords:DNAzyme amplifier;signal amplification;miRNA detection;pancreatic cancer
研究领域:
分子生物学
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医学诊断
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生物化学
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DOI
高影响因子
MICROCHIMICA ACTA
·
2017
·Vol. 184
·期 8
·2941-2948
·
IF 4.58
Han Cong,Li Rongguo,Li Hui,Liu Su,Xu Chenggong,Wang Jingfeng,Wang Yu,Huang Jiadong
The authors describe a label-free electrochemical aptasensor for ultrasensitive and highly specific detection of the antibiotic based on cascade enzymatic recycling couple with DNAzyme amplification. The assay involves two sequential reactions: the first reaction is a λ exonuclease-assisted cyclic digestion reaction triggered by target-aptamer binding. The second reaction is a nicking endonuclease-aided cyclic nicking reaction, which produces a large amount of G-rich nucleic acid segments. These form a G-quadruplex/hemin complex in the presence of K(I) ions and hemin. Because the G-quadruplex/hemin complex acts as a horseradish peroxidase-mimicking DNAzyme with excellent redox activity, the electrochemical signal transduction is accomplished due to the electroreduction of H 2 O 2 . It appears that this work is the first example that cascade enzymatic recycling coupled to DNAzyme amplification is used for antibiotic detection. The aptasensor was applied to the quantitation of kanamycin and gave a response that is linear in the 1 pM to 10 nM kanamycin concentration range, with a detection limit as low as 0.5 pM. The working voltage (vs. Ag/AgCl) at which data can be acquired best is −0.35 V. The assay offers the advantages of remarkably improved sensitivity, use of affordable instrumentation, and simplified operation without the need for electrochemical labeling or addition of labile reagents. Thus, cascade enzymatic recycling coupled to DNAzyme amplification represents a versatile platform for highly sensitive and specific antibiotics detection.
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研究领域:
分析化学
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生物医学工程
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分子生物学
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
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2022
·
IF 9.221
Jianmei Yang,Qian Bi,Xinmei Song,Ruo Yuan,Yun Xiang
Sensitive and accurate monitoring of amino acid concentration variations plays an important role for diagnosis of diseases at early stage. Here, the aptamer/organometallic receptor-based methodology for label-free and amplified electrochemical determination of phenylalanine , an amino acid, through multi-pedal DNA walker and DNAzyme dual signal enhancement is demonstrated. The binding of the phenylalanine/organometallic receptor complex with the aptamer results in the release of active DNAzyme walking strands to trigger the multi-pedal DNA walkers to move on the surface of electrode. The substrate hairpins are then cyclically cleaved by the DNA walkers to liberate a large number of free G-quadruplexes, which interact with and confine hemin on the electrode via the G-quadruplex/hemin structures. Significantly amplified current signals can therefore be generated by electrochemical reduction of hemin for the sensitive measurement of phenylalanine down to 1.03 μM. Such an aptasensor also has a high selectivity and can monitor phenylalanine in diluted serums, making it a promising alternative for the development of versatile and robust aptamer-based platforms for detection of various amino acids.
研究领域:
生物化学
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分析化学
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医学诊断
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DOI
高影响因子
BMC Pharmacology & Toxicology
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2019
·Vol. 20
·期 1
·1-7
·
IF 2.103
Shi Xiaoping,Gao Hui,Li Zhong,Li Jinghua,Liu Yang,Li Lujuan,Zhang Qi
Background Methotrexate is an important component in many chemotherapy protocols. The blood concentration of Methotrexate is used to determine the regimen of folinic acid. However, the lower limit of Siemens assay kit is 0.30 μmol/L in China. This study extended the limit from 0.3 to 0.05 μmol/L and reduced the test cost by optimizing the parameters of Enzyme Multiplied Immunoassay Technique assay.Methods Parameters of Enzyme Multiplied Immunoassay Technique assay were modified to decrease the volume of reagents A and B. Then a standard curve with a new custom set of calibrators was prepared to detect low concentration. Intra-day and inter-day imprecision were assessed by control material and samples. The linearity of the modified assay was verified by analyzing a range of quality controls with known concentration from 0.05 to 1.00 μmol/L. At last, the same samples were tested by modified Enzyme Multiplied Immunoassay Technique assay and Liquid Chromatography-tandem Mass Spectrometry assay respectively. A simple linear regression was performed to verify the validity of the modified Enzyme Multiplied Immunoassay Technique assay.Results Intra-day and inter-day imprecision show good reproducibility at all levels (0.05, 0.12, 0.43, 0.82 μmol/L). The linearity equation of modified assay was y = 0.9913x + 0.0046, in which y was the mean of measured concentration and x was the target concentration (R2 = 0.9994). In the range of 0.05–10.00 μmol/L, correlation between the Modified assay and Liquid Chromatography-tandem Mass Spectrometry assay was significant (r = 0.9968). In the range of 0.30–10.00 μmol/L, the correlation between modified and commercial assays was significant (r = 0.9987) as well.Conclusion sThe modified assay enhanced the sensitivity of Siemens VIVA-E to 0.05 μmol/L. In addition, the test number of a reagent Kit increased from 140 to 210. This means the cost of detection was reduced about 30%.
研究领域:
医学检验
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药理学
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临床化学
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DOI
PubMed
高影响因子
ANALYST
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2019
·Vol. 144
·期 16
·4995-5002
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IF 4.019
Rufeng Zhang,Yu Wang,Xiaonan Qu,Shasha Li,Yihan Zhao,Fenfen Zhang,Su Liu,Jiadong Huang,Jinghua Yu
Herein, a split G-quadruplex DNAzyme as a signal reporter was integrated into an electrochemical sensing platform for the detection of antibiotics with specificity and sensitivity. To improve the signal-to-noise ratio, two G-rich oligonucleotide sequences (G1 and G2) were blocked into two different hairpin probes, preventing the two segments from assembling into a spilt G-quadruplex structure. Moreover, we designed a double-arch probe, consisting of an aptamer as the recognition element and two-step enzymatic signal amplification. Concretely, the first is the Nt.BbvCI-assisted nicking cyclic reaction activated by target-aptamer binding, and the second is exonuclease III-aided cyclic amplification for generating abundant G1 and G2. The modified capture probe on the electrode was used to combine G1 and G2 to form the spilt G-quadruplex/hemin when K+ and hemin were present. This complex plays the role of DNAzyme with superior horseradish peroxidase activity in catalyzing the decomposition of H2O2. Under optimal conditions, this biosensor showed an excellent performance for sensing kanamycin with a detection limit of 83 fM for kanamycin concentrations ranging from 100 fM to 1 nM. Hence, the proposed strategy has potential as an efficient and actual platform for small molecule analysis.
研究领域:
生物化学
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分析化学
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生物技术
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DOI
PubMed
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
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2017
·
IF 5.401
Yali Yuan,Guangpeng Liu,Yaqin Chai,Ruo Yuan
In this study, an electrochemical proximity assay (ECPA)-coupled highly nonenzymatic amplifying strategy with electron mediator collected by a host-guest interaction was proposed for total protein of N. bombycis (TP N.b) detection. Antibody-dsDNA-gold nanoparticles functional Fe 3 O 4 probe (termed as [email protected] 3 O 4 NPs), in which the Ab-dsDNA composed by adenine rich ssDNA (A 1 ) and Ab labeled capture ssDNA 1 (Ab-S 1 ), was first incubated with target TP N.b and another Ab labeled capture ssDNA 2 (Ab-S 2 ) to perform the proximity immunoreaction. Satisfyingly, the immunoreaction could not only produce sandwich immunocomplex with the proximity hybridization of S 1 and S 2 , but also exposed the trigger ssDNA A 1 to in situ form substantial methylene blue (MB) intercalated DNA concatamers via hybrid chain reaction (HCR). To further realize the nonenzymatic amplification, the MB intercalated in DNA concatemers was released into solution by duplex specific nuclease (DSN) and subsequently collected by prepared cucurbit(7)-uril (CB[7])/ [email protected] 3 O 4 NPs/GCE via host-guest interaction, which thus produced a significantly amplified signal for quantitative detection of target. Since the catalyst Fe 3 O 4 NPs on electrode surface could directly electrocatalyze the reduction of coimmobilized MB, the proposed system possessed superior advantages compared with that of traditionally enzymatic systems, which not only simplified the operation process, decreased measurement error but also improved catalytic efficiency and stability.
研究领域:
生物技术
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医学检测
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分子诊断
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DOI
高影响因子
TALANTA
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2021
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IF 6.057
Ting Huang,Guixun Chen,Birong Liu,Zizhong Yang,Yuanwei Huang,Baoping Xie,Min-Min Li,Jin-Xiang Chen,Jun Chen,Zong Dai
Sensitive, specific and rapid methods for detecting microRNAs (miRNAs) play critical roles in disease diagnosis and therapy. Enzyme-free amplification techniques based on DNAzyme assembly have recently been developed for the highly specific miRNA analysis. However, traditional DNAzyme-based assembly (free DNAzyme) amplifiers is mainly dependent on the target-induced split DNAzyme fragments to assemble into activated DNAzyme structures, which have made a compromise between the sensitivity and specificity due to the random diffusion of dissociative probes in a bulk solution with poor kinetics. Herein, based on a rationally designed DNA probe , we developed an intramolecular DNAzyme assembly (intra-DNAzyme) method to overcome these challenges. The miR-373 is used as model analyte for our current proof-of-concept experiments. Compared with the free-DNAzyme method, our method showed significantly improved analytical performance in terms of dynamic range, assay sensitivity and speed. This method can detect miR-373 specifically with a detection limit as low as 4.3 fM, which is about 83.7 times lower than the previous free-DNAzyme method. This intra-DNAzyme strategy would be of great value in both basic research and clinical diagnosis.
研究领域:
分子生物学
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生物技术
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纳米技术
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诊断学
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DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
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2015
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IF 6.409
Bingchen Li,Fei Liu,Yuanyuan Peng,Yunlei Zhou,Wenxuan Fan,Huanshun Yin,Shiyun Ai,Xiansheng Zhang
MicroRNA (miRNA) is riveting nowadays due to its close relevance to human malignancies. Exploiting a fast and convenient biosensor for sensitive and specific detecting miRNA is necessary and it has significant meaning for oncology. In this work, to understand the relationship between miRNA-21 and gastric cancer, we employ T4 RNA ligase 2 to initiate specific ligation reaction depending on the sequence of target RNA, and T7 exonuclease to catalyze the first stage cyclic enzymatic amplification method (CEAM) specifically, which also resting on the target RNA sequence , and the second stage CEAM to further amplify the response resulting from the initial target RNA. Our two-stage CEAM strategy can detect miRNA-21 as low as 0.36 fM with remarkable specificity, and most importantly, it can be used to inspect the expression level of miRNA-21 in blood serum of gastric cancer patients . This will offer new perspective for figuring out the pathways of miRNA-21 involving in cancer.
研究领域:
生物化学
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肿瘤学
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分析化学
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分子生物学
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产品关联
DOI
PubMed
高影响因子
TALANTA
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2020
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IF 5.339
Wenyue Zhang,Wenhui Hao,Xuetao Liu,Xiru Sun,Jingli Yan,Yucong Wang
A visualized assay for miRNAs detection has been developed in this work. The presented method is based on a combination of enzyme-free amplification cascades of catalyzed hairpin assembly (CHA) and hybridization chain reaction (HCR) and fluorescence quenching of dual-emission ratiometric fluorescent probes (RF probes). MiRNAs can efficiently initiate enzyme-free amplification reactions (CHA and HCR) and produce the long nicked dsDNAs with a lot of glucose oxidases (GOD) on the surface of dynabeads bridged by the GOD-labeled hairpin DNA probes . Hydrogen peroxide (H 2 O 2 ) is generated by oxidation of glucose catalyzed by GOD, which can quench the outer green fluorescence without affecting the internal red fluorescence of RF probes. Therefore, increased miRNA amount can result in change of the two fluorescence intensity ratios of RF probes with continuous color changes from green to red under a UV lamp, which can be easily recognized by naked eye. The proposed assay exhibits high sensitivity toward let-7a with dynamic range from 10 −13 M to 10 −8 M, and which is applied successfully to detecting let-7a in the small RNA samples.
研究领域:
分子生物学
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生物化学
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医学诊断
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DOI
PubMed
高影响因子
MICROCHIMICA ACTA
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2019
·Vol. 186
·期 2
·1-9
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IF 5.479
Jiang Ningjing,Hu Yongjun,Wei Wei,Zhu Tingfeng,Yang Kang,Zhu Guichi,Yu Meng
A surface-enhanced Raman scattering (SERS) method is described for the determination of microRNA that is associated with various forms of cancer. The substrate consists of functionalized gold-silver bimetallic structure, and the sensitivity is strongly enhanced by making use of a re-circulated enzymatic amplification system (REAS). Poly-dopamine acts as both a reductant and a protective of the substrates. It was employed to link the gold core and silver satellite. The unique “hot spots” consisting of a Au@PDA@Ag nanocomposite improve the Raman signal and sensitivity. The reductive feature of PDA can prevent the susceptible oxidation of metallic silver to maintain the high Raman activity. To improve the sensitivity of the assays, a re-circulated enzymatic amplification system was developed in which the nicking endonuclease triggers the nucleic acid reaction system to enter an amplified cycle. By integrating the bimetallic nanosubstrate and magnetic separation into the REAS, microRNA can be detected by SERS (best at the Raman band of 1586 cm −1 ) with a limit of detection as low as 0.2 fM. In our perception, the assay provides an exciting new avenue to study the expression of tumor genes. Thus, it holds vast promise in cancer diagnosis.
Schematic presentation of the SERS method based on poly-dopamine mediated bimetallic SERS substrate and re-circulated enzymatic amplification.
研究领域:
分析化学
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分子生物学
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癌症研究
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DOI
PubMed
高影响因子
LUMINESCENCE
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2013
·Vol. 29
·期 4
·338-343
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IF 1.273
Yuan Zhou,Tao Zhou,Rui Zhou,Yonggang Hu
A rapid, simple, facile, sensitive and enzyme-amplified chemiluminescence immunoassay (CLIA) method to detect antibodies against porcine parvovirus has been developed. Horseradish peroxidase (HRP) and the detection antibody were simultaneously co-immobilized on the surface of gold nanoparticles using the electrostatic method to form gold nanoparticle-based nanoprobes. This nanoprobe was employed in a sandwich-type CLIA, which enables CL signal readout from enzymatic catalysis and results in signal amplification. The presence of porcine parvovirus infection was determined in porcine parvovirus antibodies by measuring the CL intensity caused by the reaction of HRP–luminol with H 2 O 2 . Under optimal conditions, the obtained calibration plot for the standard positive serum was approximately linear within the dilution range of 1:80 to 1:5120. The limit of detection for the assay was 1:10,240 ( S / N = 3), which is much lower than that typically achieved with an enzyme-linked immunosorbent assay (1:160; S / N = 3). A series of repeatability measurements using 1:320-fold diluted standard positive serum gave reproducible results with a relative standard deviation of 4.9% ( n = 11). The ability of the immunosensor to analyze clinical samples was tested on porcine sera. The immunosensor had an efficiency of 90%, a sensitivity of 93.3%, and a specificity of 87.5% relative to the enzyme-linked immunosorbent assay results. Copyright © 2013 John Wiley & Sons, Ltd.
研究领域:
兽医学
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纳米技术
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免疫学
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分析化学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ANALYTICAL CHEMISTRY
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2014
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IF 5.825
Mo Wang,Zhengliang Fu,Bingchen Li,Yunlei Zhou,Huanshun Yin,Shiyun Ai
Taking advantage of the special exodeoxyribonuclease activity of T7 exonuclease, a simple, sensitive, selective, and label-free microRNA biosensor based on the cyclic enzymatic amplification method (CEAM) has been proposed. First, thiol functionalized DNA probes were assembled onto a gold nanoparticles modified gold electrode surface through a Au–S bond, followed by hybridizing with target miRNA. Subsequently, DNA in RNA/DNA duplexes was digested by T7 exonuclease, which can release the microRNA molecules from the electrode surface and return into the buffer solution. Meanwhile, the released microRNA can further hybridize with the unhybridized DNA probes on the modified electrode surface. On the basis of it, an isothermal amplification cycle is realized. The T7 exonuclease-assisted CEAM achieved a low detection limit of 0.17 fM. Moreover, this assay presents excellent specificity with discriminating only a single-base mismatched microRNA sequence. Furthermore, this work can also be applied to detect avian leukemia based on the decreased expression level of microRNA-21.
研究领域:
生物化学
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分子生物学
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分析化学
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癌症研究
✓ 已验证
产品关联
DOI
PubMed
高影响因子
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