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Electrochemical Impedance Spectroscopy Study of Concanavalin A Binding to Self-Assembled Monolayers of Mannosides on Gold Wire Electrodes.

ABSTRACT: The interactions of the lectin Concanavalin A (Con A) with self-assembled monolayers (SAMs) of thiolated mono-, di-, and tri-mannosides were studied on the surface of gold wires using electrochemical impedance spectroscopy (EIS). The SAMs of mannosides were prepared either pure or along with thiolated triethylene glycol (TEG) at different molar ratios (1:1, 1:2, 1:4, 1:9, and 1:19) to better understand and optimize the interaction conditions. The charge-transfer resistance of the [Fe(CN)6]3-/4- redox probe was compared before and after the interaction at different concentrations of Con A to determine the equilibrium dissociation constant (Kd) and limit of detection (LOD). Values of Kd were found in the nanomolar range showing multivalent interactions between mannosides and Con A, and LOD was found ranging from 4-13 nM depending on the type of mannoside SAM used. Analysis using the Hill equation suggests negative cooperativity in the binding behavior. Peanut agglutinin was used as a negative control, and cyclic voltammetry was used to further support the experiments. We have found that neither the pure nor the widely dispersed monolayers of mannosides provide the conditions for optimal binding of Con A. The binding of Con A to these SAMs is sensitive to the molar ratio of the mannoside used to prepare the SAM and to the structure of the mannoside. A simple cleaning method has also been shown to regenerate the used gold wire electrodes. The results from these experiments contribute to the development of simple, cheap, selective, and sensitive EIS-based bioassays, especially for lectin-carbohydrate interactions.

SUBMITTER: Bhattarai JK

PROVIDER: S-EPMC5388449 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Electrochemical Impedance Spectroscopy Study of Concanavalin A Binding to Self-Assembled Monolayers of Mannosides on Gold Wire Electrodes.

Bhattarai Jay K JK Tan Yih Horng YH Pandey Binod B Fujikawa Kohki K Demchenko Alexei V AV Stine Keith J KJ

Journal of electroanalytical chemistry (Lausanne, Switzerland) 20160928

The interactions of the lectin Concanavalin A (Con A) with self-assembled monolayers (SAMs) of thiolated mono-, di-, and tri-mannosides were studied on the surface of gold wires using electrochemical impedance spectroscopy (EIS). The SAMs of mannosides were prepared either pure or along with thiolated triethylene glycol (TEG) at different molar ratios (1:1, 1:2, 1:4, 1:9, and 1:19) to better understand and optimize the interaction conditions. The charge-transfer resistance of the [Fe(CN)<sub>6</ ...[more]

PMID: 28413372

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Project description:The background: The monolayers self-assembled on the gold electrode incorporated transition metal complexes can act both as receptor ("host" molecules) immobilization sites, as well as transducer for interface recognitions of "guest" molecules present in the aqueous solutions. Their electrochemical parameters influencing the sensing properties strongly depend on the transition metal complex structures. The objectives: The electrochemical characterization of the symmetric terpyridine-M2+-terpyridine and asymmetric dipyrromethene-M2+-terpyridine complexes modified with ssDNA probe covalently attached to the gold electrodes and exploring their ssDNA sensing ability were the main aims of the research presented. The methods: Two transition metal cations have been selected: Cu2+ and Co2+ for creation of redox-active monolayers. The electron transfer coefficients indicating the reversibility and electron transfer rate constant measuring kinetic of redox reactions have been determined for all SAMs studied using: Cyclic Voltammetry, Osteryoung Square-Wave Voltammetry, and Differential Pulse Voltammetry. All redox-active platforms have been applied for immobilization of ssDNA probe. Next, their sensing properties towards complementary DNA target have been explored electrochemically. The results: All SAMs studied were stable displaying quasi-reversible redox activity. The linear relationships between cathodic and anodic current vs. san rate were obtained for both symmetric and asymmetric SAMs incorporating Co2+ and Cu2+, indicating that oxidized and reduced redox sites are adsorbed on the electrode surface. The ssDNA sensing ability were observed in the fM concentration range. The low responses towards non-complementary ssDNA sequences provided evidences for sensors good selectivity. The conclusions: All redox-active SAMs modified with a ssDNA probe were suitable for sensing of ssDNA target, with very good sensitivity in fM range and very good selectivity. The detection limits obtained for SAMs incorporating Cu2+, both symmetric and asymmetric, were better in comparison to SAMs incorporating Co2+. Thus, selection of the right transition metal cation has stronger influence on ssDNA sensing ability, than complex structures.

Project description:BackgroundDNA hybridization is at the basis of most current technologies for genotyping and sequencing, due to the unique properties of DNA base-pairing that guarantee a high grade of selectivity. Nonetheless the presence of single base mismatches or not perfectly matched sequences can affect the response of the devices and the major challenge is, nowadays, to distinguish a mismatch of a single base and, at the same time, unequivocally differentiate devices read-out of fully and partially matching sequences.ResultsWe present here two platforms based on different sensing strategies, to detect mismatched and/or perfectly matched complementary DNA strands hybridization into ssDNA oligonucleotide monolayers. The first platform exploits atomic force microscopy-based nanolithography to create ssDNA nano-arrays on gold surfaces. AFM topography measurements then monitor the variation of height of the nanostructures upon biorecognition and then follow annealing at different temperatures. This strategy allowed us to clearly detect the presence of mismatches. The second strategy exploits the change in capacitance at the interface between an ssDNA-functionalized gold electrode and the solution due to the hybridization process in a miniaturized electrochemical cell. Through electrochemical impedance spectroscopy measurements on extended ssDNA self-assembled monolayers we followed in real-time the variation of capacitance, being able to distinguish, through the difference in hybridization kinetics, not only the presence of single, double or triple mismatches in the complementary sequence, but also the position of the mismatched base pair with respect to the electrode surface.ConclusionWe demonstrate here two platforms based on different sensing strategies as sensitive and selective tools to discriminate mismatches. Our assays are ready for parallelization and can be used in the detection and quantification of single nucleotide mismatches in microRNAs or in genomic DNA.

Dataset Information

Electrochemical Impedance Spectroscopy Study of Concanavalin A Binding to Self-Assembled Monolayers of Mannosides on Gold Wire Electrodes.

Publications

Electrochemical Impedance Spectroscopy Study of Concanavalin A Binding to Self-Assembled Monolayers of Mannosides on Gold Wire Electrodes.

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