Date of Award
Honors College Thesis
Biological Sciences BS
Alex Flynt, Ph.D.
Fluorescence microscopy is a bioimaging technology that utilizes the excitation and emission of fluorophores to identify cellular structures, processes, and interactive events. Natural and synthetic organic dyes are frequently used in fluorescence microscopy techniques for imaging, therapeutic, and biomedical applications. It is also employed in the development of novel organic dyes for innovative methods of drug delivery and labelling. In this study, we investigate the behavior of an organic dye consisting of the post-polymerization modification of poly(2-vinyl-4,4-dimethyl azlactone) (PVDMA) with tetramethyl rhodamine cadaverine (TMR) and a coumarin-based molecule (DBAC). Using cultured HEK293 cells and fluorescence confocal microscopy, we show the dye preferentially localizes in lysosomal compartments within 10 minutes of incubation in cell culture. By comparing incubation times, we demonstrate how the dye experiences cleavage of its coumarin group and release into the cytosol with retention of the rhodamine group. Additionally, we co-stained the cells with LysoTracker Green DND-26 to quantify the degree of colocalization of the dye in lysosomes and cytosol. We show the rhodamine group has the highest colocalization with LysoTracker, and the colocalization of the coumarin group with LysoTracker and rhodamine is significantly reduced. This research elucidates the intracellular trafficking of the DBAC-TMR-PVDMA dye and provides necessary knowledge to further understand its behavior and develop an effective model for potential biomedical applications.
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Tassin, Garrett, "Uptake and Localization of Poly(2-vinyl-4,4-dimethyl azlactone) Modified with Rhodamine- and Coumarin-Based Molecules in Human Embryonic Kidney (HEK293) Cells" (2023). Honors Theses. 899.