Louise Aveyard, Jenny
(2008)
Gold Nanoparticles For Biomolecular Assays.
PhD thesis, University of Liverpool.
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Abstract
The amalgamation of nanotechnology and biology has led to the development ofnew types of hybrid materials that are expected to produce major advances in areas such as materials science, therapeutics and diagnostics. One of the most promising developments is the use ofnanoparticles (NPs) as labels for the detection of analytes in biological assays. The aim of this research project was to prepare gold nanoparticle (GNP) labels for use in such assays. In chapter 1, the optical properties and the use of GNPs in homogeneous and heterogeneous colorimetric assays are reviewed. In chapter 2 a simple conjugation method is introduced that not only allows almost any biological molecule or hapten to be attached to GNPs but also allows the user to control or vary the mean number of molecules per particle. In this method a high molecular weight aminodextran polymer is functionalized with the molecule of choice and chemical attachment groups that are used to covalently anchor the polymer to the GNPs. This method was used to conjugate biotin and 1125 functionalized dextrans to GNPs. These functionalized dextrans were then used to investigate the conjugation procedure in more_detail. Results from GNP titrations and microbead assays demonstrate that the minimum amount of functionalized dextran required to prevent salt-induced flocculation ofthe GNPs (equivalence point) is the amount required to coat all of the GNPs and at this point there is no free functionalized dextran in solution. In chapter 3 the described method was used to conjugate different numbers DNP haptens to GNPs and then these labels were used in non-traditional reagent-limited lateral flow immunoassays. The number of molecules per GNP is varied by simply adjusting the stoichiometry of reagents in the dextran functionalization reaction. Controlling the number of molecules per particle can have important consequences on the sensitivity of a biological assay. Results showed that when the number of DNP molecules per particle decreased, there was an increase in the sensitivity of the assay. Furthermore when the results from these immunoassays were compared to those obtained from traditional reagent-limited lateral flow immunoassays, the nontraditional format proved to be over 50 % more sensitive. In chapter 4 the conjugation method was used to attach oligonucleotides to GNPs for use in a nucleic acids lateral flow (NALF) device. Although NALF devices are available commercially, detection is usually achieved with the use of antibodies or haptens which can be both problematic and expensive. In addition, many of these devices have issues with sensitivity and are often interfaced with complicated target amplification / purification protocols. In chapter 4 an antibody / hapten independent NALF device is described that can be used to detect the un-purified products from a simple polymerase chain reaction (PCR) amplification protocol. Using the developed NALF device it was possible to detect specific amplification products corresponding to ~1 attomole oftemplate molecules with the unaided eye.
| Item Type: | Thesis (PhD) |
|---|---|
| Depositing User: | Symplectic Admin |
| Date Deposited: | 20 Oct 2023 09:24 |
| Last Modified: | 20 Oct 2023 09:45 |
| DOI: | 10.17638/03174401 |
| Copyright Statement: | Copyright © and Moral Rights for this thesis and any accompanying data (where applicable) are retained by the author and/or other copyright owners. A copy can be downloaded for personal non-commercial research or study, without prior permission or charge |
| URI: | https://livrepository.liverpool.ac.uk/id/eprint/3174401 |
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