Multivalent linkers for improved covalent binding of oligonucleotides to dye-doped silica nanoparticles

Abstract

This paper describes the fabrication of oligonucleotide-coated Cy5-doped silica nanoparticles using a combination of multivalent linkers and their use in surface-based DNA sandwich hybridization assays. Dipodal silane is introduced as a means to fabricate amine-coated silica nanoparticles and its advantages compared to monopodal silanes are discussed. The use of dipodal silane in conjunction with three different polymer linkers (oxidized dextran, linear and 8-arm polyethylene glycol (PEG)) to immobilize single-stranded DNA to Cy5-doped nanoparticles is investigated and dynamic light scattering measurements and Fourier transform infrared spectroscopy are used to follow the progression of the functionalization of the nanoparticles. We observe a significant improvement in the binding stability of the single-stranded DNA when the dipodal silane and 8-arm PEG are used in combination, when compared to alternative conjugation strategies. Both 8mer and 22mer oligonucleotides are securely conjugated to the high-brightness nanoparticles and their availability to hybridize with a complementary strand is confirmed using solution-based DNA hybridization experiments. In addition, a full surface-based sandwich assay demonstrates the potential these nanoparticles have in the detection of less than 500 femtomolar of a DNA analogue of micro RNA, miR-451.

Synthesis and characterisation of far-red fluorescent cyanine dye doped silica nanoparticles using a modified microemulsion method for application in bioassays

“…Antibody functionalisation of Cy5 NPs, sample 2E plus silica shell, was achieved using a step-by-step approach with multivalent linkers (see supporting information) [29].…”

Dye-doped silica nanoparticles: synthesis, surface chemistry and bioapplications

“…A notable study by Bagwe et al (2006) showed how a combination of negatively charged 'inert' phosphonate groups and 'reactive' amine groups can be combined on the surface of RuBpy-doped silica NPs to reduce particle aggregation, while also still providing the reactive sites needed for later functionalisation. 'Bipodal' bis amine [3-(triethoxysilyl) propyl] has also been proposed as an alternative to traditional 'monopodal' APTMS as the former produces more colloidally stable amine-coated silica NPs (Kelleher et al 2015). Wang et al demonstrated that silica NPs can also be used as a platform for catalysis and was facilitated by the use of amino-functionalised colloids.…”

Ultrasensitive microarray bioassays using cyanine5 dye-doped silica nanoparticles

“…Cyanine5 NPs used in this study were prepared using a microemulsion method with an intramicellar nucleation pathway (Nooney et al 2015a). NP PEGylation was achieved using a step-by-step biconjugation strategy with multivalent linkers (Kelleher et al 2015). As part of the NPs synthesis the Cy5 dye is encapsulated inside a silica matrix protected by a tetraethyl orthosilicate shell.…”

“…The use of silica NPs also provides a chemically stable matrix with low photon absorption at visible and near infra-red wavelengths, and the silanol surface can be easily functionalised with proteins and biomolecules (Kumar et al 2008, Liu et al 2010. The potential use of dye-doped silica NPs has been reported (Bae et al 2012, Chinen et al 2015 and such NPs are already utilised in flow-cytometry (Carmen et al 2009), DNA assays (Kelleher et al 2015), and immunosorbent assays (Lian et al 2004, Wu et al 2008. In this study, NPs were prepared using the microemulsion method (Osseo-Asare and Arriagada 1990), which produces highly monodisperse NPs with efficient dye loading (Gubala et al 2011).…”