DNA Tetrahedra Modules as Versatile Optical Sensing Platforms for Multiplexed Analysis of miRNAs, Endonucleases, and Aptamer–Ligand Complexes

Abstract

The sensing modules for analyzing miRNAs or the endonucleases consist of tetrahedra functionalized with three different fluorophore–quencher pairs in spatially quenched configurations and hairpin units acting as recognition elements for the analytes. Three different miRNAs (miRNA-21, miRNA-221, and miRNA-155) or three different endonucleases (Nt.BbvCI, Eco RI, and Hin dIII) uncage the respective hairpins, leading to the switched-on fluorescence of the respective fluorophores and to the multiplex detection of the respective analytes. In addition, a tetrahedron module for the multiplexed analysis of aptamer ligand complexes (ligands = ATP, thrombin, VEGF) is introduced. The module includes edges modified with three spatially separated fluorophore–quencher pairs that were stretched by the respective aptamer strands to yield a switched-on fluorescent state. Formation of the respective aptamer ligands reconfigures the edges into fluorophore-quenched caged-hairpin structures that enable the multiplexed analysis of the aptamer–ligand complexes. The facile permeation of the tetrahedra structures into cells is used for the imaging of MCF-7 and HepG2 cancer cells and their discrimination from normal epithelial MCF-10A breast cells.

Triggered Dimerization and Trimerization of DNA Tetrahedra for Multiplexed miRNA Detection and Imaging of Cancer Cells

“…In fact, we have recently reported on the use of fluorophore–quencher‐modified tetrahedra as functional nanostructures for the multiplexed analysis of miRNAs. [ 83 ] The use of donor–acceptor bifluorophores reveals, however, clear advantages over fluorophore/quencher pairs, due to the fact that the fluorophore/quencher pair results in a single fluorescence intensity signal, whereas the bifluorophore donor–acceptor configuration yields two complementary readout signals (the decay of the donor emission and the emergence of the acceptor emission). The later emission is, particularly, important for intracellular miRNA imaging since it provides a readout fluorescence channel to avoid false‐positive signals.…”

Static quenching upon adduct formation: a treatment without shortcuts and approximations

“…Three fluorophores were integrated as the transducers of the parallel and multiplexed analysis of the different analytes. 52 Another DNA tetrahedron based multicolour nanoprobe was used for simultaneous imaging of three tumour-related mRNAs in living cells. Here, the fluorescence quenching served as the initial step to guarantee a dark background and a sufficient contrast to observe fluorescence restoration caused by competitive chain replacement reaction in cancerous cells.…”

DNA nanostructure-based nucleic acid probes: construction and biological applications

“…DNA tetrahedron-linked hairpin probes can be used to construct a multifunctional optical sensing platform for multiple analyses of miRNAs, endonucleases, and small molecules. 72 The DNA tetrahedral framework can also promote folding dynamics and improve the thermodynamic stability of aptamers. The binding affinity of the aptamer is increased by ∼3 times after being modified on a DNA framework.…”