A real-time PCR-based method for determining the surface coverage of thiol-capped oligonucleotides bound onto gold nanoparticles.

Nucleic Acids Research
Eun-Young KimSteven M Wolinsky

Abstract

Here we report a real-time PCR-based method for determining the surface coverage of dithiol-capped oligonucleotides bound onto gold nanoparticles alone and in tandem with antibody. The detection of gold nanoparticle-bound DNA is accomplished by targeting the oligonucleotide with primer and probe binding sites, amplification of the oligonucleotide by PCR, and real-time measurement of the fluorescence emitted during the reaction. This method offers a wide dynamic range and is not dependant on the dissociation of the oligonucleotide strands from the gold nanoparticle surface; the fluorophore is not highly quenched by the gold nanoparticles in solution during fluorescence measurements. We show that this method and a fluorescence-based method give equivalent results for determining the surface coverage of oligonucleotides bound onto 13 or 30 nm gold nanoparticles alone and in tandem with antibody. Quantifying the surface coverage of immobilized oligonucleotides on metallic nanoparticle surfaces is important for optimizing the sensitivity of gold nanoparticle-based detection methods and for better understanding the interactions between thiol-functionalized oligonucleotides and gold nanoparticles.

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Citations

Jan 3, 2013·Advanced Drug Delivery Reviews·Jose-Maria MontenegroWolfgang J Parak
Oct 20, 2011·Cellular and Molecular Life Sciences : CMLS·Tetiana Kurkina, Kannan Balasubramanian
May 27, 2009·Journal of Immunological Methods·Longyan ChenXian-En Zhang
Nov 4, 2015·ACS Applied Materials & Interfaces·Ying WanXiaolei Zuo
Nov 27, 2020·Nature·Benjamin S MillerRachel A McKendry
Jun 12, 2010·Analytical Chemistry·Dongmao Zhang, Siyam M Ansar
Oct 9, 2021·Scientific Reports·Cassandra M StawickiParag Mallick

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Methods Mentioned

BETA
surface plasmon resonance
fluorophore
transmission electron microscopy
PCR
electrophoresis
fluorescence resonance

Software Mentioned

ImageTool
BLAST

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