Label free electrical detection of DNA hybridisation for the example of influenza virus gene sequences

A. Kukol, P. Li, P. Estrela, P. Ko-Ferrigno, P. Migliorato

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51 Citations (Scopus)
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Abstract

Microarrays based on DNA–DNA hybridization are potentially useful for detecting and subtyping viruses but require fluorescence labeling and imaging equipment. We investigated a label-free electrical detection system using electrochemical impedance spectroscopy that is able to detect hybridization of DNA target sequences derived from avian H5N1 influenza virus to gold surface-attached singlestranded DNA oligonucleotide probes. A 23-nt probe is able to detect a 120-nt base fragment of the influenza A hemagglutinin gene
sequence. We describe a novel method of data analysis that is compatible with automatic measurement without operator input, contrary to curve fitting used in conventional electrochemical impedance spectroscopy (EIS) data analysis. A systematic investigation of the detection signal for various spacer molecules between the oligonucleotide probe and the gold surface revealed that the signal/background ratio improves as the length of the spacer increases, with a 12- to 18-atom spacer element being optimal. The optimal spacer molecule allows a detection limit between 30 and 100 fmol DNA with a macroscopic gold disc electrode of 1 mm radius. The dependence of the detection
signal on the concentration of a 23-nt target follows a binding curve with an approximate 1:1 stoichiometry and a dissociation constant
of Kp = 13 ± 4 nM at 295 K.
Original languageEnglish
Pages (from-to)143-153
JournalAnalytical Biochemistry
Volume374
Issue number1
DOIs
Publication statusPublished - 2008

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