Nanoparticle displacement assay with electrochemical nanopore-based sensors

Abstract

Please cite this article as: Gergely Lautner, Mónika Plesz, Gyula Jágerszki, Péter Fürjes, Róbert E. Gyurcsányi, Nanoparticle displacement assay with electrochemical nanopore-based sensors, Electrochemistry Communications (2016Communications ( ), doi: 10.1016Communications ( /j.elecom.2016 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT Abstract:The proof of concept of a nanoparticle displacement assay that enables the use of large diameter nanopores for the detection of targets of smaller molecular dimensions is presented. We hypothesized that an inherent signal amplification should arise from the selective displacement of nanoparticles preloaded in a nanopore by a much smaller molecular target. The method is demonstrated using peptide nucleic acid (PNA)-functionalized gold nanopore arrays in which short DNA-modified gold nanoparticles are anchored by weak interaction. Complementary microRNAs are detected via the resistance change caused by competitive displacement of nanoparticles from the PNA-functionalized nanopores.

Potentiometric sensing of nucleic acids using chemically modified nanopores

“…To overcome this problem the thiol-terminated PNA strands were immobilized in a prehybridized form with a short 7-mer complimentary DNA strand that we found earlier by surface plasmon resonance (SPR) to self-regulate the surface density of immobilized strands for optimal hybridization efficiency without the need of lengthy optimization experiments 37 and also to reduce the extent of non-specific adsorption. 34 After immobilization of the prehybridized strand we observed a cationic response that could be turned into an anionic response by removing the weakly bound short DNA strand with 0.1 M NaOH (Fig. 2).…”

“…S1, ESI †). 34,36 The gold surfaces were modified using thiol terminated 18-mer PNA (N′-Lys-GCTTTTTGCTCGTCTTAT-AEEA-C6-SH-C′), where C6 is (CH 2 ) 6 spacer and AEEA 2-(2-aminoethoxy) ethoxyacetic acid (Eurogentec Seraing, Belgium). 22-mer microRNA (5′-AUAAGACGAGCAAAAAGCUUGU-3′) or its DNA analog was used as the target while a similar length random Scheme 1 Schematics of the measurement setup (A) and the idealized sensing concept (B) for the potentiometric detection of nucleic acids.…”

“…For the proof of principle we used gold nanopore arrays consisting of 7 hexagonally arranged pores (Fig. S1, ESI †) 34 modified with positively charged thiol labeled PNA strands for the detection of a 22-mer microRNA (miRNA) as a model for conserved short length single stranded nucleic acids. 35 Our hypothesis was that such membranes exhibit anionic permselectivity which upon selective binding of the negatively charged complementary microRNA strands is switched to cationic permselectivity in a concentration dependent manner and the associated potential increase can be used for quantitation (Scheme 1).…”

PNA-Based MicroRNA Detection Methodologies

“…Using solid-state nanopore arrays, Gyurcsányi's group presented a biosensor based on the displacement of DNA-AuNPs conjugates for miR-208a detection in serum [68] (see Figure 3C). Gold nanopore arrays were functionalized with 18-mer PNAs complementary to the target miR.…”

“…Examples of biosensors exploiting AuNPs were already reported above. In the aforementioned cases, the use of nanoparticles was justified as a means to increase the number of capturing probes on the biosensor surface (see [59]) or, in the case of nanopore-based detection, as a way to block the pore (see [68]). AuNPs, however, can additionally be employed in order to amplify the signal, enhancing the system sensitivity and generating lower background, thus increasing the LOD of the systems.…”