Electropolymerization of Bilayer with Phosphonic Acid Tethers for Immobilization of Biomolecules

Hartung, J. B.; Kowalik, Janusz; Kranz, Christine; Janata, Jiřı́; Josowicz, Mira; Sinha, A. K.; McCoy, Kendra

doi:10.1149/1.2041868

Cited by 15 publications

(14 citation statements)

References 42 publications

(44 reference statements)

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“…The probes are anchored to the polypyrrole bilayer through the available free phosphonic acid functional groups at the polymer surface. Divalent metal ions bound to phosphonic acid tethers of the poly[2,5-dithienyl- N -(phosphorylalkyl)pyrrole] (pTPT) film provide electrostatic anchor sites through the negatively charged phosphate groups of the probes. , Voltammetric transduction neither requires labeling of probes nor uses redox mediators or redox intercalators. It is based on the fact that a polypyrrole (PPy) film acts as an electrochemically driven ion exchanger and that large charged molecules attached near the surface of this polymer exchanger change the ability of the ions to move into or out of the film .…”

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confidence: 99%

Label-Free Voltammetric Detection Using Individually Addressable Oligonucleotide Microelectrode Arrays

et al. 2010

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The utility and performance of label-free, oligonucleotide probes for reagentless detection of dilute target analytes was examined using a voltammetric transduction principle in an array format. Multistep, solid-state fabrication yielded preproduction arrays of 16 individually addressable, 30 μm diameter microelectrodes in a 30 mm × 6.5 mm × 0.5 mm dipstick disposable device. The specificity of 16 nucleotide (nt) 2'-O-methylribonucleic acid and 22 nt DNA backbone probes bound through Mg(2+)-phosphonate bridges to polypyrrole films on the microelectrodes were studied using microbial target RNAs of various lengths. Probe-specific interactions with Escherichia coli O157 H7 23S rRNA (2907 nt) and Candida albicans 18S rRNA (1788 nt) were detected at 65 and 58 fmol/mL, respectively, in volumes as low as 0.5 mL. Specificity studies showed that, for a given probe, "nontarget" transcripts can contribute to changes in the voltammetric detection signal, though with responses that never exceed 70% of the detection signal acquired for specifically designed complementary targets. These results statistically validate the use of the voltammetric microelectrode array for obtaining a "yes-no" answer on complementary specific binding. The study also identifies challenges and pitfalls for the selection strategies of oligonucleotide probes.

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confidence: 99%

Label-Free Voltammetric Detection Using Individually Addressable Oligonucleotide Microelectrode Arrays

et al. 2010

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“…As mentioned in the introduction, the specificity of the bio-recognition of our electrochemical DNA hybridization method was studied with synthetic 27-mer strands of oligonucleotide of the sequence, which relates to a hypermethylated region of the GSTP1 gene (Hartung et al 2005). The correlation between GSTP1 hypermethylation and the occurrence of prostate cancer enables the practical use of these sequences for rapid detection of hypermethylation and by implication, prostate cancer.…”

Section: Resultsmentioning

confidence: 99%

“…The monomer of 2,5-dithienyl-(N-3-phosphorylpropyl) pyrrole, pTPTC3PO 3 H 2 where C3- in this tether group defines the number of methylated groups shortly abbreviated as (pTPT) was synthesized according to Hartung et al (Hartung et al 2005). Pyrrole (98%), Magnesium chloride hexahydrate, potassium chloride, tetrabutylammonium perchlorate (Bu 4 NClO 4 ), acetonitrile, and Tris–HCl buffer (0.1 M, pH 7.2) were all of reagent grade and were used as received.…”

Section: Methodsmentioning

confidence: 99%

Voltammetric Application of Polypyrrole-Modified Microelectrode Array for the Characterization of DNA Methylation in Glutathione S-Transferase Pi 1

2018

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Direct and efficient label-free voltammetric detection of Glutathione S-Transferase Pi 1 (GSTP1) hypermethylation is reported using a custom developed 16-channel Microelectrode Array chip. The microelectrode array chip is used in a dipstick configuration allowing detection of DNA hybridization in a solution volume of only 0.35 mL. Platinum microelectrode disks (n = 16) 30 µm in diameter have been modified with a polypyrrole bilayer before any contact with the oligonucleotides. The attachment of the 15-mer Probe DNA to the bilayer is random but controlled by the presence of aliphatic tether groups allowing it to form a bidentate complex with the probe DNA. The voltammetric detection procedure of methylated GSTP1-specific target DNA is combined with bisulfite treatment of target DNA. Changes at the interface of the modified microelectrodes in an array configuration are used to record simultaneously cyclic voltammetry on all of the devices. The detection of the hybridization is evaluated statistically for a event by comparing the changes in recorded cyclic voltammograms before and after exposure to the Target DNA. All cyclic voltammograms of the methylated target show a greater percentage change than those with the non-methylated target exposure and show a greater change in cyclic voltammogram area after methylated target exposure. We observe an average percentage difference of 25.6% ± 4.9 with a variation of 19.1%. These results demonstrate that the fast sensing strategy possesses sensitivity and good specificity. Furthermore, this technology can potentially support rapid, accurate diagnosis and risk assessment of patients with prostate cancer.

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“…Before use, pyrrole was purified by passing it through a neutral alumina column. pTPTC3−PO 3 H 2 ) is not commercially available and was synthesized according to Hartung et al Magnesium chloride was obtained from Fluka. Tetra- n -buthylammonium perchlorate was obtained from Alfa Aesar (Ward Hill, MA).…”

Section: Methodsmentioning

confidence: 99%

Label-Free DNA Detection Based on Modified Conducting Polypyrrole Films at Microelectrodes

et al. 2006

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A label-free electrochemical detection method for DNA hybridization based on electrostatic modulation of the ion-exchange kinetics of a polypyrrole film deposited at microelectrodes is reported. Synthetic single-stranded 27-mer oligonucleotides (probe) have been immobilized at 2,5-bis(2-thienyl)-N-(3-phosphorylpropyl)pyrrole film formed by electropolymerization on the previously formed polypyrrole layer. The 27- or 18-mer target oligonucleotides were monitored via the electrochemically driven anion exchange of the inner polypyrrole film. The performance of the miniaturized DNA biosensor system was studied in respect to selectivity, sensitivity, reproducibility, and regeneration of the sensor. Control experiments were performed with a noncomplementary target of 27-mer DNA and 12 base-pair mismatched 18-mer sequences, respectively, and did not show any unspecific binding. Under optimized experimental conditions, the label-free electrochemical biosensor enabled the detection limits of 0.16 and 3.5 fmol for the 18- and 27-mer DNA strand, respectively. Furthermore, we demonstrate reusability of the electrochemical DNA biosensor after successful recovery of up to 100% of the original signal by regenerating the DNA "label-free" electrode with 50 mM HCl at room temperature.

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Electropolymerization of Bilayer with Phosphonic Acid Tethers for Immobilization of Biomolecules

Cited by 15 publications

References 42 publications

Label-Free Voltammetric Detection Using Individually Addressable Oligonucleotide Microelectrode Arrays

Label-Free Voltammetric Detection Using Individually Addressable Oligonucleotide Microelectrode Arrays

Voltammetric Application of Polypyrrole-Modified Microelectrode Array for the Characterization of DNA Methylation in Glutathione S-Transferase Pi 1

Label-Free DNA Detection Based on Modified Conducting Polypyrrole Films at Microelectrodes

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