Fluorescence anisotropy-based structure-switching aptamer assay using a peptide nucleic acid (PNA) probe

Methods. 2016 Mar 15:97:69-74. doi: 10.1016/j.ymeth.2015.09.018. Epub 2015 Oct 9.

Abstract

This study describes for the first time the feasibility of using peptide nucleic acids (PNAs) as an alternative to the DNA probes in structure-switching aptamer fluorescence polarisation assays. The effects of experimental parameters such as the length of the PNA strand, the nature of dye and the buffer conditions on the assay performances are first explored using two different methodologies based on the competition between the PNA/aptamer hydribridisation and the target/aptamer complexation. D-ATP can be detected from 1 to 25 μM in a linear range and a detection limit (LOD) of 3 μM can be reached. For this target, this lowers by a factor >5 the LOD reported with conventional DNA-based fluorescent structure switching aptamer-based assays and by a factor 3 the LOD observed with non-competitive fluorescent sensing platform indicating the usefulness of the PNA-based approach.

Keywords: Aptamer-based assay; Fluorescence anisotropy; Peptide nucleic acid; Structure-switching.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / analysis*
  • Aptamers, Nucleotide / chemistry
  • Biosensing Techniques*
  • Fluorescence Polarization
  • Hydrogen-Ion Concentration
  • Limit of Detection
  • Peptide Nucleic Acids / chemistry*

Substances

  • Aptamers, Nucleotide
  • Peptide Nucleic Acids
  • Adenosine Triphosphate