Fast enzyme-linked electrochemical sensing of DNA hybridization at pencil graphite electrodes. Application to detect gene deletion in a human cell culture

In this paper we present a rapid electrochemical enzyme-linked DNA hybridization assay using disposable pencil graphite electrodes (PeGE) to detect target DNA (tDNA) sequences in DNA fragments amplified by polymerase chain reaction. The procedure consists of several short (1–2 min) incubation steps,...

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Bibliographic Details
Published inJournal of electroanalytical chemistry (Lausanne, Switzerland) Vol. 862; p. 113951
Main Authors Špaček, Jan, Eksin, Ece, Havran, Luděk, Erdem, Arzum, Fojta, Miroslav
Format Journal Article
LanguageEnglish
Published Elsevier B.V 01.04.2020
Subjects
DNA hybridization
Do-it-yourself devices
Enzyme-linked electrochemical assay
Gene deletion
Pencil graphite electrode
Ultralow cost
Do-it-yourself devices
Enzyme-linked electrochemical assay
DNA hybridization
Gene deletion
Pencil graphite electrode
Ultralow cost
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Summary:In this paper we present a rapid electrochemical enzyme-linked DNA hybridization assay using disposable pencil graphite electrodes (PeGE) to detect target DNA (tDNA) sequences in DNA fragments amplified by polymerase chain reaction. The procedure consists of several short (1–2 min) incubation steps, including adsorption of the tDNA at unpretreated PeGE from denaturing medium, surface blocking with milk proteins, hybridization with a biotinylated oligonucleotide probe and binding of streptavidin-alkaline phosphatase conjugate to the biotin tags. Then the PeGE is transferred into background electrolyte solution containing 1-naphthyl phosphate, which is enzymatically dephosphorylated to give electrochemically oxidizable indicator 1-naphthol. The assay, which can be performed within 7–8 min, offers a perfect discrimination between specific and nonspecific DNA amplicons and easy detection of about ~40 femtomoles of tDNA in large excesses of non-complementary DNA. An application on the detection of p53 gene deletion in a human cell culture, featuring a real biological model, is presented. The designed setup has a potential to be applied as one of simple, fast, robust ultralow cost “do-it-yourself “instruments recently introduced as diagnostic tools for third world countries.
ISSN:1572-6657
1873-2569
DOI:10.1016/j.jelechem.2020.113951