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Development of nanoparticle approaches for correlative imaging and tracking of biological sample
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Foisor, Mihaela Veronica (2022) Development of nanoparticle approaches for correlative imaging and tracking of biological sample. PhD thesis, University of Warwick.
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WRAP_Theses_Foisor_2022.pdf - Submitted Version - Requires a PDF viewer. Download (313Mb) | Preview |
Official URL: http://webcat.warwick.ac.uk/record=b3860832
Abstract
The main techniques used in cell biology and biomedical fields to visualize cellular events and better understand their dynamics are light microscopy and electron microscopy. These two techniques allow visualization of different aspects of the cellular environment, with light microscopy techniques tracking events in temporal resolution and electron microscopy showing an increase in spatial resolution. By combining these two techniques and performing correlative light microscopy and electron microscopy (CLEM), one can achieve both temporal and spatial resolution on the same sample. The main drawback with CLEM is the ability to produce probes that are capable of being imaged in both techniques in live cellular systems.
In this thesis, we introduce a new type of bifunctional probe that allows live CLEM tracking of GFP tagged proteins in in vitro cell lines. The new design is based on a small nanoparticle conjugated to a GFP binding nanobody, which is then transfected into GFP expressing cells via Pep-1 cell penetrating peptide.
Chapter 3 focuses on characterizing the new live CLEM probe by size, affinity to GFP proteins and binding to chimeric proteins in live cells whilst using gold nanoparticles as the nanometal material. In Chapter 4, the probe is improved by changing the gold nanoparticle to a quantum dot-based probe. This improvement allows a better characterization of the delivery of the system in live cells whilst also permitting tracking of probe intracellularly. Finally, Chapter 5 performs pre-embedding CLEM experiments using the quantum dot probe to track GFP tagged clathrin cages and ER bound proteins in HeLa and RPE-1 cells.
Item Type: | Thesis (PhD) | ||||
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Subjects: | Q Science > QC Physics Q Science > QH Natural history Q Science > QH Natural history > QH301 Biology Q Science > QP Physiology |
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Library of Congress Subject Headings (LCSH): | Microscopy, Electron microscopy, Green fluorescent protein, Molecular probes, Quantum dots | ||||
Official Date: | January 2022 | ||||
Dates: |
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Institution: | University of Warwick | ||||
Theses Department: | Department of Chemistry | ||||
Thesis Type: | PhD | ||||
Publication Status: | Unpublished | ||||
Supervisor(s)/Advisor: | Smith, Corinne J. ; Royle, Stephen J. | ||||
Sponsors: | Engineering and Physical Sciences Research Council ; Molecular Analytical Science Centre for Doctoral Training | ||||
Format of File: | |||||
Extent: | xviii, 214 pages : illustrations | ||||
Language: | eng |
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