Date of Award

2019

Document Type

Thesis

Degree Name

Bachelors

Department

Natural Sciences

First Advisor

Walstrom, Katherine

Area of Concentration

Biochemistry

Abstract

Cancer achieves its deadliest state when a solid tumor begins shedding cells into the bloodstream, known as metastatic disease. Given the mortality associated with this stage of disease progression, the detection of these circulating tumor cells (CTCs) has significant prognostic value. However, there does not yet exist a valid method to characterize these cells once isolated. The ability to functionally characterize CTCs would certainly facilitate patient care by informing clinicians of pertinent studies such as drug responses. We recently developed a technique that has the potential to address this gap: lipid tethering. By utilizing cytophobic polyelectrolyte multilayers paired with a terminal cytophilic coating, we have been able to immobilize and study several cell lines. This method is unique in that it preserves a cell’s freefloating phenotype while also restricting its movement, allowing for thorough imaging and longitudinal study. Given the nascence of this technology, the objective of this thesis is to contribute to optimization and validity. We sought to improve tethering efficiency by implementing a thermal cross-linking step after the deposition of the polyelectrolyte multilayer, conferring greater stability by imidizing the layers. This resulted in greatly improved cell retention on lipid tethers upwards of 95% when washed with chemically distinct reagents. Additionally, we tested another aspect of cell behavior, viability, to investigate if lipid tethering affected a cell’s ability to persist. We found that tethering MCF10A cells, a nontumorigenic mammary epithelial line, yielded a similar rate of cell death to suspended MCF10A cells as expected, supporting the hypothesis that lipid tethering mimics a free-floating environment. Unsuccessful experiments here included staining for cleaved PARP, an apoptotic marker, and performing a Western blot for cleaved PARP on lipid tethers. Future studies should also seek to replicate these studies in a wider variety of cell lines and, eventually, begin implementing robust functional characterization studies leading to clinical implementation.

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