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Title: Raman imaging to assess the risk of disease using a targeted nanoparticle-DNA reporter system
Author: Callan, Kirsty
ISNI:       0000 0004 9357 2541
Awarding Body: University of Strathclyde
Current Institution: University of Strathclyde
Date of Award: 2020
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The aim of this work was to develop a novel method for detecting mRNA in cells using gold nanoparticles and surface enhanced Raman spectroscopy (SERS), with the ultimate goal of early detection of cardiovascular disease. Three different methods to detect mRNA in cells using Raman spectroscopy were investigated. The first approach used a method analogous to fluorescence in situ hybridisation (FISH), named Raman in situ hybridisation. An oligonucleotide complementary to the mRNA target was attached to gold nanoparticles and labelled with a Raman reporter. When incubated with cells it was hoped that the probe would hybridise to the target mRNA and allow visualisation using Raman mapping. Despite utilisation of various probe designs, no significant discrimination could be seen between the target oligonucleotide and a random control oligonucleotide. The second approach made use of a molecular beacon. Gold nanoparticles were functionalised with a dye and a dye-labelled hairpin loop oligonucleotide. In the closed position SERS signal was obtained from both dyes due to close proximity with the nanoparticles. In the presence of the target oligonucleotide, a change in the spectra was observed due to an increase in distance between the oligonucleotide dye and the nanoparticle. This allowed ratiometric detection of the target. For the final approach nanoparticles were coated with a Raman active dye and anoligonucleotide. Two batches of nanoparticles were prepared, each with oligonucleotides complementary to half of the target sequence. When combined with the target sequence, the oligonucleotides should both hybridise, bringing the nanoparticles closer together and enhancing the Raman signal. These probes could be detected in cells, however it was not possible to determine whether or not they were hybridised to the target mRNA meaning further work is required. Overall, each method had its own limitations and further development would be required to create a feasible assay.
Supervisor: Graham, Duncan ; Mullins, John ; Faulds, Karen Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral