Doctor of Philosophy
Dr. Zhifeng Ding
Scanning electrochemical microscopy (SECM) scans a biased ultramicroelectrode (≤ 25 µm) probe over a sample to characterize topography, physical properties and chemical reactivity. In this dissertation, SECM was used to investigate the metal-induced changes in membrane response of single live human bladder cancer cells (T24). SECM imaging was coupled to 3D finite element method (FEM) simulations which were the first of their kind, providing advanced quantification of sample traits under conditions not previously usable.
The effects of Cd2+ on T24 cell membrane permeability were examined. Experimental depth-scan imaging was coupled with full 3D FEM simulations, eliminating many limitations of previous 2D-axially symmetric models. Hundreds of probe approach curves (PACs) can now be extracted from depth-images and theoretically fit to quantify membrane permeability at any location across the cell surface (Chapter 2).
SECM was utilized to examine the membrane response of T24 cells following exposure to toxic dichromate (Cr(VI)). Two electrochemical mediators were examined, the membrane permeable ferrocenemethanol (FcCH2OH) and impermeable ferrocenecarboxylate (FcCOO‑). Cr(VI) induced permeability change was observed with both mediators and compared (Chapter 3). Chronic Cr(VI) induced cell stress, was then examined. Similar permeability curve shape was observed, with shifts in response time based on concentration of Cr(VI) stressor (Chapter 4).
Trace essential metals such as Cr(III) are essential in low concentrations but toxic in high concentration. Membrane-response was investigated by SECM, using both FcCH2OH and FcCOO- redox mediators. Theoretical SECM depth-scans were produced using 3D FEM simulations, and used to quantify cell membrane permeability (Chapter 5).
Complex close-proximity cell clusters were experimentally imaged by SECM 3D scanning mode. Tailored 3D model geometries were created, generating complimentary theoretical maps of the experimental cell clusters. The simulations were capable of providing a strong theoretical fit to the experimental results. Limits of cell proximity for SECM characterization were determined based on the probe size (Chapter 6).
Nanoscale SECM imaging of single live cells were performed using a laser-pulled 130 nm radius Pt disk electrode. A tailored 3D model was created, from which cell topography was accurately characterized using membrane-impermeable Ru(NH3)63+, and cell membrane permeability was quantified with FcCH2OH (Chapter 7).
Filice, Fraser, "Coupling Scanning Electrochemical Microscopy and 3D Modelling to Probe Membrane Permeability of Human Bladder Cancer (T24) Cells" (2018). Electronic Thesis and Dissertation Repository. 5781.
Available for download on Wednesday, May 01, 2019