Highly sensitive electrochemical detection of DNA hybridisation by coupling the chemical reduction of a redox label to the electrode reaction of a solution phase mediator

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Author list: Ngoensawat U., Rijiravanich P., Somasundrum M., Surareungchai W.

Publisher: Royal Society of Chemistry

Publication year: 2014

Journal: Analyst (0003-2654)

Volume number: 139

Issue number: 22

Start page: 5740

End page: 5746

Number of pages: 7

ISSN: 0003-2654

eISSN: 1364-5528

URL: https://www.scopus.com/inward/record.uri?eid=2-s2.0-84907974429&doi=10.1039%2fc4an01011f&partnerID=40&md5=48af89b1e6369abae88756ce9820c58b

Languages: English-Great Britain (EN-GB)

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Abstract

We have described a highly sensitive method for detecting DNA hybridisation using a redox-labeled stem loop probe. The redox labels were poly(styrene-co-acrylic) (PSA) spheres of 454 nm diameter, modified by methylene blue (MB) deposited alternatively with poly(sodium 4-styrene sulphonate) (PSS) in a layer-by-layer process. Each PSA sphere carried approx. 3.7 × 105 molecules of MB, as determined optically. DIG-tagged stem loop probes were immobilised on screen printed electrodes bearing anti-DIG antibodies. Binding with the target enabled straightening of the stem loop, which made attachment to the MB-coated PSA spheres possible. For measuring the current from the direct reduction of MB by differential pulse voltammetry, a 30 mer DNA target common to 70 strains of Escherichia coli was calibrated across the range 1.0 fM to 100 pM (gradient = 3.2 × 10−8 A (log fM)−1, r2 = 0.95, n = 60), with an LOD of ∼58 fM. By using Fe(CN)6 3−/4− as a solution phase mediator for the MB reduction, we were able to lower the LOD to ∼39 aM (gradient = 5.95 × 10−8 A (log aM)−1, r2 = 0.96, n = 30), which corresponds to the detection of 0.76 ag (∼50 molecules) in the 2 μL analyte sample. We hypothesise that the lowering of the LOD was due to the fact that not all the MB labels were able to contact the electrode surface. © 2014 the Partner Organisations.

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