Date of Award:
Doctor of Philosophy (PhD)
Timothy A. Gilbertson
Ronald C. Sims
Roger A. Coulombe, Jr.
The overall goal of this dissertation is to develop noninvasive imaging techniques that allow us not only to detect diseased cells but also to study the molecular mechanisms underlying these diseases.
Atomic force microscopy and Raman spectroscopy are applied to measure cellular mechanical properties (e.g. Young’s Modulus, adhesion force) and biochemical composition of living cancerous vs. healthy (A549 vs. SAEC) human lung epithelial cells. These biomechanical and biochemical properties can be utilized to differentiate between cancerous A549 and healthy SAEC human lung epithelial cells. Furthermore, different cellular responses to anticancer drug doxorubicin (DOX) treatment are also observed. Using AFM and Raman spectroscopy, we can quantitatively measure biophysical properties of different cells, as complementary parameters to other properties (e.g. gene and protein expression), helping identify the states of diseased cells.
Another major task of this dissertation is to develop noninvasive imaging techniques to detect cancer biomarker epidermal growth factor receptor (EGFR) at single cell level using advanced instrumentation. We first synthesized a gold nanorod (AuNR)-based nanoprobe for single-cell imaging of EGFR using surface-enhance Raman spectroscopy (SERS). SERS is able to quantitatively measure the EGFR expression level in different breast cancer cell lines and map the cellular distribution of EGFR in single cells. Moreover, SERS, as a noninvasive imaging technique, is able to monitor the process of nanoparticle uptake by single cell. Due to the diffraction limit of optical microscopy, SERS is unable to provide nanoscale imaging resolution. We then applied an AFM-based simultaneous Topography and RECognition (TREC) imaging technique to image EGFR with nanoscale resolution. TREC is first validated on mica surface and then successfully utilized to map the EGFR distribution in fixed and living breast cancer cells at single molecule level. In addition, we have explored the potential of a gadolinium-gold (Gd-Au) composite nanomaterial as a dual functional (MRI-SERS) imaging probe. Using this previous reported MRI contrast agent, we successfully apply SERS function in the detection of EGFR in three cancer cell lines.
The last part of the dissertation is to study fat-responsive G protein-coupled receptor 120 (GPR120), and its interaction with linoleic acid (LA). We have synthesized a dual functional composite nanoparticle for SERS-fluorescence bimodal imaging of GRP120 in living HEK293 cells. By SERS-fluorescence imaging, we are able to locate GPR120 distribution in single cells. Moreover, we have observed a dose-dependent GPR120 response to LA treatments using SERS. This work demonstrates the potential to use SERS-fluorescence bimodal imaging technique for real-time detection of the interaction between fatty acids and their receptors (e.g. GPR120, CD36).
Xiao, Lifu, "Molecular Sensing and Imaging of Human Disease Cells and Their Responses to Biochemical Stimuli" (2015). All Graduate Theses and Dissertations. 4278.
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