| One of the most promising advances in the field of regenerative medicine has been the isolation of pluripotent stem cells, embryonic and adult, capable of differentiating into multiple tissue types. Yet a critical question remains: what is the fate of transplanted stem cells in vivo over the time of an experimental study? Presently, there are no effective noninvasive, non-genetic toolsets available to monitor the fate of delivered therapeutic stem cells. If an advanced level of understanding of stem cell location and fate in vivo can be attained, researchers may be able to cure diseases such as Parkinson's disease, diabetes, heart disease, multiple sclerosis, spinal cord injuries, and other degenerative diseases.;Due to their tunable physiochemical and fluorescent characteristics, QDs have potential as a tool for stem cell-tracking in vitro and in vivo. However, observations of QD labeled cells have been mainly qualitative in nature, limiting our ability to determine ideal loading conditions for long-term imaging in vivo. Currently, there are no standard methods for determining the number of particles taken per cell to make comparisons between experiments and literature. Without this value, optimizing a QD toolset for tracing stem cell fate will be challenging.;Therefore this thesis developed methods for quantitative loading of cells with QDs using commercially available materials and flow cytometry. Using these methods, the fate of 705 and 800 nm emitting QD-labeled cells longitudinally in cell culture was determined. In a muscle derived stem cell (MDSC) line, the effects that pArg- and CTB-QD labeling may have, if any, on osteogenic and myogenic stem cell function was evaluated. Lastly, the threshold and limits of detecting 705 and 800 nm emitting QD-labeled cells in vivo was quantified.;These novel quantitative methods have allowed us to make comparisons of QD uptake and retention by cells with respect to cell type, QD conjugate type, QD bioactivity, and experimental conditions. While there are physical, chemical, and biological questions in the field regarding QD technology for cell biology and biomedical engineering research applications (such as the mechanism of uptake of QDs by cells or subcellular location), I developed a quantitative measure of QD uptake by single cells, to allow for dose optimization studies for cell-tracking in biomedical in vitro and in vivo model systems, and to support quantitative studies of QD toxicity -- an area still under investigation. In the long term, the application of quantified methods developed here will help move cell-based therapies from the bench to the clinic. |
