A simple and universal enzyme-free approach for the detection of multiple microRNAs using a single nanostructured enhancer of surface plasmon resonance imaging

Andrea Sguassero, Álvaro Artiga, Carlo Morasso, Rafael Ramirez Jimenez, Rafael Martín Rapún, Roberta Mancuso, Simone Agostini, Ambra Hernis, Arturs Abols, Aija Linē, Alice Gualerzi, Silvia Picciolini, Marzia Bedoni, Marco Rovaris, Furio Gramatica, Jesus M. de la Fuente, Renzo Vanna

Research output: Contribution to journalArticle

Abstract

Here we describe a simple approach for the simultaneous detection of multiple microRNAs (miRNAs) using a single nanostructured reagent as surface plasmon resonance imaging (SPRi) enhancer and without using enzymatic reactions, sequence specific enhancers or multiple enhancing steps as normally reported in similar studies. The strategy involves the preparation and optimisation of neutravidin-coated gold nanospheres (nGNSs) functionalised with a previously biotinylated antibody (Ab) against DNA/RNA hybrids. The Ab guarantees the recognition of any miRNA sequence adsorbed on a surface properly functionalised with different DNA probes; at the same time, gold nanoparticles permit to detect this interaction, thus producing enough SPRi signal even at a low ligand concentration. After a careful optimisation of the nanoenhancer and after its characterisation, the final assay allowed the simultaneous detection of four miRNAs with a limit of detection (LOD) of up to 0.5 pM (equal to 275 attomoles in 500 μL) by performing a single enhancing injection. The proposed strategy shows good signal specificity and permits to discriminate wild-type, single- and triple-mutated sequences much better than non-enhanced SPRi. Finally, the method works properly in complex samples (total RNA extracted from blood) as demonstrated by the detection of four miRNAs potentially related to multiple sclerosis used as case study. This proof-of-concept study confirms that the approach provides the possibility to detect a theoretically unlimited number of miRNAs using a simple protocol and an easily prepared enhancing reagent, and may further facilitate the development of affordable multiplexing miRNA screening for clinical purposes.

Original languageEnglish
JournalAnalytical and Bioanalytical Chemistry
Early online dateAug 24 2018
DOIs
Publication statusE-pub ahead of print - Aug 24 2018

    Fingerprint

Keywords

  • Enhancement
  • miRNA
  • Multiplexing
  • Nanobiosensor
  • Nanoparticles
  • SPR

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry

Cite this