Abstract

DNA aptamers are important tools for molecular recognition, particularly for a new generation of tools for biomedicine based on nucleic acid nanostructures. Here, we investigated the relative abilities of different shapes and sizes of DNA polyhedra to display an aptamer which binds to the malaria biomarker Plasmodium falciparum lactate dehydrogenase (PfLDH). The aptamer was shown to perform an Aptamer-Tethered Enzyme Capture (APTEC) assay with the hypothesis that the display of the aptamer above the surface through the use of a polyhedron may lead to better sensitivity than use of the aptamer alone. We compared different numbers of points of contact, different shapes, including tetrahedron, square, and pentagon-based pyramids, as well as prisms. We also investigated the optimal height of display of the structure. Our results demonstrated that the display of an aptamer on an optimized nanostructure improved sensitivity up to 6-fold relative to the aptamer alone in the APTEC assay. Other important factors included multiple basal points of contact with the surface, a tetrahedron proved superior to the more complex shaped structures, and height above the surface only made minor differences to efficacy. The display of an aptamer on a nanostructure may be beneficial for higher sensitivity aptamer-mediated malaria diagnosis. Aptamer displays using DNA nanostructure polyhedron supports could be a useful approach in a variety of applications.

Highlights

  • Aptamers are single-stranded oligonucleotides that can fold into specific secondary structures to facilitate molecular recognition events in diagnostics and therapeutics [1,2]

  • We previously identified a DNA aptamer against the malaria biomarker Plasmodium falciparum lactate dehydrogenase (PfLDH), which is useful in malaria point-of-care diagnostics [3,4]

  • Before generating the sequence for the random region, a constant sequence was assigned to the structure to ensure the generation of specific duplex sequences for the assembly, including the sequence of PfLDH aptamer, the thymidine residues at each turning point of the nanostructure and the spacer at each base vertex

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Summary

Introduction

Aptamers are single-stranded oligonucleotides that can fold into specific secondary structures to facilitate molecular recognition events in diagnostics and therapeutics [1,2]. We previously identified a DNA aptamer against the malaria biomarker Plasmodium falciparum lactate dehydrogenase (PfLDH), which is useful in malaria point-of-care diagnostics [3,4]. We have integrated the aptamer into a range of diagnostic strategies, including colorimetric assay [6,7], electrochemical sensing [8], and even incorporating it into. Enzyme Capture (APTEC) assay uses the intrinsic enzymatic activity of PfLDH to trigger a color change of nitrotetrazolium blue from colorless to blue as an observable signal [6]. While the APTEC assay is sensitive enough to diagnose malaria in most clinical blood samples of lower than 0.01% parasitaemia with 90% accuracy, it was not able to detect the disease in samples at Molecules 2018, 23, 1695; doi:10.3390/molecules23071695 www.mdpi.com/journal/molecules

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