Enzyme-Amplified Amperometric Detection of 3000 Copies of DNA in a 10-μL Droplet at 0.5 fM Concentration

Abstract

We reported earlier the detection of a 38-base DNA strand at 20 pM concentration by an enzyme-amplified sandwich-type amperometric assay. The assay utilized a carbon electrode on which a redox polymer, comprising a DNA capture sequence, was electrodeposited. When present in the tested solution, part of the probed sequence hybridized with the capture probe. Hybridization of its remaining part with a horseradish peroxidase (HRP)-labeled sequence resulted in the flow of an H2O2 electroreduction current, the redox polymer wired HRP forming an electrocatalyst. Here we report a > 10(4)-fold improvement in the detection limit of the assay. DNA was detected at 0.5 fM concentration when the earlier used 3.6-mm-diameter carbon electrode was replaced by a 10-microm-diameter microelectrode. The radial diffusion of electrons through the film on the microelectrode allowed the electrodeposition of a thicker film of the redox polymer, an increase in the loading of the capture sequence, and increased the collection efficiency of the electron vacancies originating in the electroreduced H2O2. When the volume probed by the microelectrode was 10 microL, as few as 3000 copies of DNA were detected.

Rational Tuning of the Electrocatalytic Nanobiointerface for a “Turn‐Off” Biofuel‐Cell‐Based Self‐Powered Biosensor for p53 Protein

“…[2,30] However,t he reported aptamer-basedb iochemicalsignals for controlling BFCs' powerrelease can just realize aq ualitative determination of the analytes.T his can be ascribed to the little controlo ver the molecular orientation in the above systems, which resultsi ni nefficient electron transfer and electrical contact. In order to solvet he above issue related to electron transfer,r ecently researchers have focusedo nt he introduction of mediator (e.g.,f errocene [31,32] and [Os(2,2'-bipyridine) 2 Cl 2 ] 2+/3 + ) [33][34][35] into the DNA-modified electrode interface for promoting electron transfer.H owever,t hese studies have their inherentd isadvantages. Mediator-based electron transfer in biocatalysis processes is sophisticated, inconvenient and complex fort he development of miniaturized sensors, since the mediator leak is unavoidable from the modified film.…”

Highly Sensitive Indicator‐Free Impedance Sensing of DNA Hybridization Based on Poly(m‐aminobenzenesulfonic acid)/TiO2 Nanosheet Membranes with Pulse Potentiostatic Method Preparation

“…Most strategies for nucleic acid detection require a DNA hybridization event, which could be monitored by either labeled [1] or label-free [2,3] methods. Label assays identify the target analyte, which has been previously modified with a linked label such as enzymes, heterocyclic dyes, ferrocene derivatives, or organometallic complexes.…”

A Competitor‐switched Electrochemical Sensor for Detection of DNA

“…Among electrochemical methods for DNA detection above, sandwich assay utilizing DNA hybridization is widely used due to its facility in design and construction. [15][16][17][18] Recently, electrochemical detections of DNA by immobilizing hairpin and single-stranded DNA on electrode surface have been reported. [19][20][21][22][23] This kind of methods are based on an electrochemical response resulted from target-induced structure change of the probe strand containing a redox-active reporter group, which was immobilized on electrode surface.…”