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Four-wave mixing heterodyne interferometric imaging of gold nanoparticles in cells

Giannakopoulou, Panagiota 2018. Four-wave mixing heterodyne interferometric imaging of gold nanoparticles in cells. PhD Thesis, Cardiff University.
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Abstract

Gold nanoparticles (NPs) are widely utilised for bio-imaging applications owing to their facile synthesis, ease of surface functionalisation and bio-conjugation, as well as bio-compatibility. However, when embedded in highly heterogeneous and fluo- rescing environments such as biological cells and tissues, these NPs have to be large (typically >50 nm diameter) to be distinguished optically against backgrounds via their linear absorption and scattering at the surface plasmon resonance (SPR). As a result, cell imaging protocols often adopt the use of fluorophore tags attached onto the NP, and assume that the fluorophore is a reliable reporter. These constructs are routinely used in correlative microscopy techniques such as Correlative Light Elec- tron microscopy (CLEM) and presume that the fluorophore colocalises well with the electron dense NP. In this thesis, I present the application of our recently developed four-wave-mixing (FWM) imaging modality to investigate the spatial correlation be- tween a gold NP and a fluorescently labeled biomolecule attached onto it on a variety of gold NPs-fluorophore conjugate constructs inside biological cells. Owing to the specific nature of the FWM process, which is a non-linear light matter interaction with the free-electron gas in the gold metal, I was able to detect single gold NPs down to 10 nm diameter background-free and with high 3D spatial resolution even in highly heterogeneous and fluorescing cellular environments. The FWM signal from gold was spatially correlated with the fluorescence signal of the fluorophore at- tached onto the biomolecule of the construct. Surprisingly, even covalently attached conjugate constructs exhibited low colocalization highlighting the limitations of flu- orescence tagging as a means to address fundamental questions about intracellular pathways of high biological interest. This study opens new prospects to the use of FWM microscopy for imaging small gold NPs potentially even in live cells and to the understanding of the cellular uptake of bio-conjugated gold NPs.

Item Type: Thesis (PhD)
Date Type: Submission
Status: Unpublished
Schools: Biosciences
Subjects: Q Science > Q Science (General)
Q Science > QC Physics
Uncontrolled Keywords: Gold nanoparticles, four-wave mixing, nonlinear microscopy, correlative microscopy
Date of First Compliant Deposit: 18 March 2019
Last Modified: 19 Mar 2019 10:15
URI: http://orca-mwe.cf.ac.uk/id/eprint/120821

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