Label-Free Voltammetric Detection Using Individually Addressable Oligonucleotide Microelectrode Arrays

Abstract

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.

An electrochemical DNA sensor based on polyaniline/graphene: high sensitivity to DNA sequences in a wide range

“…8a) decreased while the peak of P/G 1:100 sensor (inset of Fig. Kalantari et al reported that the target length (38, 107, 1788 and 2907 base pairs) can contribute to changes in the hybridization signal, 46 which was in good agreement with the P/G sensor. In Fig.…”

Voltammetric Label‐free Detection of DNA Hypermethylation Using Polypyrrole‐modified Microelectrode Array

“…All experiments were conducted using a chip consisting of an array of 16 platinum microdisk electrodes, soldered to the printed circuit board and connected to 16 channel multipotentiostat .…”

“…The procedures for assembly of MEA with polypyrrole bilayer were already reported . Briefly, the polypyrrole bilayer was chronocoulometrically deposited in a two‐step procedure using 0.1 M pyrrole and then 0.1 M poly[2,5‐dithienyl‐(N‐3‐phosphorylpropyl)pyrrole] (TPT−C3−PO 3 H 2 ), respectively.…”

“…It revealed excellent reproducibility and selectivity of DNA hybridization event. In order to further elucidate the impact of ODN probes randomly anchored to the PPy-layer on the reagent less voltammetric transduction principle dilute ODN targets, as a function of their length, were investigated in a microelectrode array format [25]. Probespecific interactions with Escherchia coli (2907nt) and Candida albicans (1788nt) were detected at 65 and 58 fmol/l, respectively, in volume as low as 0.5 ml.…”

Robust Functionalization of Large Microelectrode Arrays by Using Pulsed Potentiostatic Deposition

“…Microelectrode arrays (MEAs) constitute a widely used platform that enables spatially highly resolved parallel sensing [ 1 , 2 , 3 , 4 , 5 , 6 ]. Owing to the advances in microfabrication technology, established methods for manufacturing microelectrodes from a large variety of materials over a wide range of geometries and spatial arrangements are available [ 7 , 8 , 9 , 10 ].…”