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

Abstract

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.

An Overview of Biosensors Based on Glutathione Transferases and for the Detection of Glutathione

“…Indeed, under the prostatic carcinoma setting, GSTP1-1 expression is silenced due to aberrant hypermethylation in its gene promoter. Different sensitive biosensors were developed and used for the determination of GSTP1-1 hypermethylation as a prostate cancer biomarker [73][74][75].…”

“…The device was appropriate to detect methylation in GSTP1-1 with good sensitivity and high specificity. Furthermore, it is suitable for a yes or no response in order to obtain a rapid diagnostic test [75]. A fluorogenic substrate that allows direct and selective detection of GSTP1-1 activity in living cells was also developed [71].…”

“…This biosensor presented specificity and reproducibility, with low detection limits, as well as being disposable and regenerable. More recently, a label-free voltametric assay was also developed [75]. The biosensor used a 16-channel platinum microelectrode array chip.…”