A superstructure-based electrochemical assay for signal-amplified detection of DNA methyltransferase activity

Zhang, Hui; Yang, Yin; Dong, Huilei; Cai, Chenxin

doi:10.1016/j.bios.2016.07.103

Cited by 30 publications

(13 citation statements)

References 57 publications

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“…A LOD of 3.5 × 10 –3 U mL –1 is obtained based on the control group plus three times standard deviation. The sensitivity of the proposed ECL biosensor is much higher than those of the reported methods − (Table S1), with 7.1-fold higher than that of electrochemical assay, − 8.6-fold higher than that of electrochemiluminescence assay, 10-fold higher than that of photoelectrochemical assay, 40-fold higher than that of fluorescence assay, , and 714.3-fold higher than that of colorimetric assay . The high sensitivity of this biosensor can be ascribed to (1) the strong ECL emission of the Ag-MOG induced by target M.SssI MTase, (2) the high quenching efficiency of Fc in the absence of target M.SssI MTase, and (3) the efficient supramolecular host–guest recognition reaction.…”

Section: Resultsmentioning

confidence: 99%

A Host–Guest Interaction-Based and Metal–Organic Gel-Based Biosensor with Aggregation-Induced Electrochemiluminescence Enhancement for Methyltransferase Assay

Cui

Zhao

et al. 2021

Anal. Chem.

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Metal−organic gels (MOGs) are new soft materials with the characteristics of high colloidal stability, superb luminescence properties, and facile synthesis. Herein, we develop for the first time a host−guest interaction-based and MOG-based biosensor with aggregation-induced electrochemiluminescence (ECL) enhancement for M.SssI methyltransferase (M.SssI MTase) assay. This biosensor employs a MOG as the luminophor and potassium persulfate as the coreactant, and the formation of the Ag-MOG from the aggregation of silver nanoclusters can induce significant ECL enhancement. Two complementary single-stranded DNAs (ssDNAs, i.e., biotinylated DNA-1 and Fc-labeled DNA-2) that contain specific recognition sequence 5′-CCGG-3′ can form a double-stranded DNA (dsDNA) probe. In the absence of M.SssI MTase, the dsDNA probe will be digested by restriction endonuclease HpaII, leading to the release of Fc from magnetic beads (MBs). The β-CD can specifically recognize the released Fc through guest−host interaction, resulting in the quenching of an ECL signal. In contrast, the presence of M.SssI MTase enables the formation of fully methylated dsDNA, which cannot be cleaved by HpaII, making Fc remain on the MB surface and consequently generating an improved ECL signal. This biosensor can specifically detect M.SssI MTase with a linear range of 0.05−100 U mL −1 and a limit of detection of 3.5 × 10 −3 U mL −1 , and it enables accurate detection of M.SssI MTase in human serum. In addition, it can be used for inhibitor screening, with wide applications in drug discovery and disease diagnosis.

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Section: Resultsmentioning

confidence: 99%

A Host–Guest Interaction-Based and Metal–Organic Gel-Based Biosensor with Aggregation-Induced Electrochemiluminescence Enhancement for Methyltransferase Assay

Cui

Zhao

et al. 2021

Anal. Chem.

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“…PNAs are DNA mimics in which the bases are attached to a neutral N-(2-aminoethyl)-glycine pseudopeptide backbone, and LNAs are nucleic acid analogues in which the ribose ring is "locked" by a methylene bridge that shows increased thermal stability [4]. Other capture probes used are chemically modified at one of the oligonucleotide ends, and usually, biotin, thiol, or amine residues are aggregated, biotin and thiol being the most used chemical modifications [7][8][9][37][38][39][40]. Moreover, the surface chemistry used in this study enabled the immobilization of cDNA, which allowed developing a biosensor for gene expression determination from children with obesity.…”

Section: Real-time Quantitative Polymerase Chain Reactionmentioning

confidence: 99%

NLRP3, IL-1β, and Caspase-1 Gene Transcript Identification and Expression by QCM-D in Obese Children

et al. 2019

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Background. Obesity in children is highly prevalent in Mexican population. Adipose tissue has been related to specific pro- and anti-inflammatory cytokine and inflammasome gene and protein expression patterns. Actually, there is no existing biosensor for detecting gene expression patterns in children with obesity. The quartz crystal microbalance with dissipation monitoring (QCM-D) has been used as a transducer for DNA biosensor design. Results. In this study, the gene expression pattern of IL-1β, NLRP3, and CASPASE-1 in children with obesity was successfully determined by means of QCM-D. Gene expression patterns were validated with those obtained by quantitative polymerase chain reaction (qPCR), a validated molecular biology technique for gene expression quantification. QCM-D analysis of the detected mass corresponding results for each of the genes showed a major detected mass for IL-1β, followed by similar NLRP3 and constitutive gene 18S deposited mass and a smaller deposited mass for CASPASE-1. Surprisingly, when comparing mRNA gene expression results for NLRP3, IL-1β, and CASPASE-1 obtained with qPCR and QCM-D, similar patterns were found, revealing greatest expression of IL-1β, followed by NLRP3, with CASPASE-1 being the molecule of least expression in the group of children with obesity. AFM images illustrate the step-by-step changes that took place on the quartz surface. Conclusions. QCM-D proved successfully for determining the gene transcripts and expression of NLRP3, IL-1β, and CASPASE-1 in children with obesity, with similar results validated by qPCR. “QCM-D decreases detection costs compared with a validated molecular biology technique.” The QCM-D biosensor developed by our group was successful for gene expression determination; in the future, it can be used for molecular diagnosis.

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“…In order to avoid this complexity, in the current study, weexplored whether simply monitoring the total charge generated by the electrostatically-attached RuHex onto the adsorbed DNA could report on the level of DNA methylation present in samples, where generated total redox charge is the function of adsorbed DNA sequences on the electrode surface [26][27][28][29]. Since in this "signal-on" approach,the charge of the RuHex complex is quantitatively reflecting the amount of the adsorbed DNA at the electrode surface [30], the electrochemical signal generated by the chronocoulometric (CC) interrogation of DNAbound RuHex will give the level of methylation present in the amplified samples.It is also important to note that unlike RuHexbased conventional methods [30], the current method detects DNA methylation by simply monitoring the adsorbed target DNA on an unmodified SPE-Au electrode.…”

Section: Dnamentioning

confidence: 99%

“…faster. Fourth, the detection step of our proposed assay can take only ten min in total (excluding bisulfite treatment and asymmetric PCR steps) to achieve electrochemical readout, which is considerably faster than many recent electrochemical DNA methylation assays [13,29,40].…”

Section: Gene Specific Methylation Detection and Validation In Cell Lmentioning

confidence: 99%

“…In recent years, much attention has been focused on thedevelopment ofsensitive, specific, relatively simple and inexpensive method for the analysis ofDNA methylation using electrochemistry, colorimetry, surface plasmon resonance and Raman scattering readouts [17][18][19][20][21][22].While most of these readout methods have theirown merits and demerits, electrochemical readout offersadditional advantages in clinical diagnosticsapplications due to their relatively higher sensitivity and specificity, cost-effectiveness and compatibility with the miniaturization [23][24][25].In these assays, sensor requires a surface-attached capture probe to hybridize the complementary target sequence, and form duplex DNA that intercalatively bind with a redox-active transition-metal cations (e.g., [Ru(NH3)6] 3+ ) for the generation of electrochemical signals [26][27][28][29]. As described in many conventional electrochemical assays [30][31][32][33], the saturated amount of charge-compensation [Ru(NH3)6] 3+ complex (RuHex) on the electrode surface is electrochemically determined, which is directly proportional to the number of negatively charged phosphate residues and thereby the surface density of the target…”

Section: Introductionmentioning

confidence: 99%

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Quantification of gene-specific DNA methylation in oesophageal cancer via electrochemistry

Haque

Gopalan

Islam

et al. 2017

Analytica Chimica Acta

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Development of simple and inexpensive method for the analysis of gene-specific DNA methylation is important for the diagnosis and prognosis of patients with cancer. Herein, we report a relatively simple and inexpensive electrochemical method for the sensitive and selective detection of gene-specific DNA methylation in oesophageal cancer. The underlying principle of the method relies on the affinity interaction between DNA bases and unmodified gold electrode. Since the affinity trend of DNA bases towards the gold surface follows as adenine (A) > cytosine (C) > guanine (G)> thymine (T), a relatively larger amount of bisulfite-treated adenine-enriched unmethylated DNA adsorbs on the screen-printed gold electrodes (SPE-Au) in comparison to the guanine-enriched methylated sample. The methylation levels were (i.e., different level of surface attached DNA molecules due to the base dependent differential adsorption pattern) quantified by measuring saturated amount of charge-compensating [Ru(NH)] molecules in the surface-attached DNAs by chronocoulometry as redox charge of the [Ru(NH)] molecules quantitatively reflects the amount of the adsorbed DNA confined at the electrode surface. The assay could successfully distinguish methylated and unmethylated DNA sequences at single CpG resolution and as low as 10% differences in DNA methylation. In addition, the assay showed fairly good reproducibility (% RSD= <5%) with better sensitivity and specificity by analysing various levels of methylation in two cell lines and eight fresh tissues samples from patients with oesophageal squamous cell carcinoma. Finally, the method was validated with methylation specific-high resolution melting curve analysis and Sanger sequencing methods.

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A superstructure-based electrochemical assay for signal-amplified detection of DNA methyltransferase activity

Cited by 30 publications

References 57 publications

A Host–Guest Interaction-Based and Metal–Organic Gel-Based Biosensor with Aggregation-Induced Electrochemiluminescence Enhancement for Methyltransferase Assay

A Host–Guest Interaction-Based and Metal–Organic Gel-Based Biosensor with Aggregation-Induced Electrochemiluminescence Enhancement for Methyltransferase Assay

NLRP3, IL-1β, and Caspase-1 Gene Transcript Identification and Expression by QCM-D in Obese Children

Quantification of gene-specific DNA methylation in oesophageal cancer via electrochemistry

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