Development and optimisation of gold nanoparticle based bioassays

Abstract

Gold nanoparticles, in particular nanostars, are being utilised more regularly in the field of biosensing. Despite their useful attributes, there is still a need to optimise aspects of the synthesis and stability of the nanostars. The seedless, synthetic method comprised of 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) is a facile, rapid method, however, produces heteromorphic nanostars. A modification of Xie et. al’s method resulted in a silver-assisted, seedless gold nanostar synthesis method. The nanostars resulting from this method are monodispersed, multi-branched and approximately 37nm + 2nm in diameter. It proved to be a repeatable method that produced monodispersed and robust nanostars. Once functionalised with polyvinylpyrrolidone 10 000, the new nanostars were observed to be stable in various conditions such as salt, ionic strength and cell culture medium environments. Upon assessing the colorimetric ability of the new nanostars, it was observed that the gold nanostar colorimetric assay could be tailored for a specific application using either hydroxylamine or sodium hydroxide as colorimetric catalysts. The colours obtained for both catalysts were vivid and easily detectible by naked eye determination. It was also observed that the hydroxylamine hydrochloride catalyst was more suited in detecting the absence or presence of an analyte, whereas, the sodium hydroxide was suitable for concentration dependent detection assays. Choosing the sodium hydroxide base, the colorimetric ability of these nanostars showed to be more sensitive and more visually colourful than the HEPES gold nanostars synthesised without silver nitrate. They were then applied to a colorimetric assay based on the method by Liu and colleagues. Upon the addition of fructosyl oxidase and glycated valine, the biosensor reacted to the generated hydrogen peroxide resulting in a sufficient colour gradient based on substrate concentration. When the same colorimetric assay parameters were applied to an ELISA type assay, the colour variation between different concentrations of glucose oxidase enzyme was also vivid and detectible spectrophotometrically. These new nanostars show the potential of replacing expensive equipment, reagents and lengthy experiments to determine glycated haemoglobin and microalbuminuria concentrations used in diabetes diagnosis and hypertension