Preparation, theory, and biological applications of highly luminescent cadmium selenide/zinc sulfide quantum dots in optical and electron microscopy

This dissertation describes the preparation, theory, and applications of ZnS overcoated CdSe (core) quantum dots for applications as fluorescent probes in optical microscopy and as electron energy loss spectroscopy (EELS) probes in electron microscopy, with applications to the biological sciences. The dissertation begins with a brief overview of quantum dots and their history. Next, a brief overview of the necessary semiconductor theory is discussed including the origin of the band gap, the origin of holes, the concepts of phonons, and trap states. Then, the role of the confinement potential in the quantum dot fluorescent spectrum is discussed in the context of the 3-dimensional spherical well. Included in this discussion is the role of excitonic electron-hole bound states. To provide a complete document useful to anyone who wishes to continue work along these lines, included is a methods section which describes the complete process of synthesis of the CdSe cores, overcoating the cores with ZnS, size selection of nanocrystals, water solubilization, and protein conjugation. The methods used in live cell labeling are included as well. In the section that follows, a discussion of the mathematical methods of image correlation spectroscopy (ICS) for extracting dynamic constants such as flow rates and diffusion constants from time lapse optical image data is discussed in the context of quantum dot fluorescent probes. Dynamic constants were obtained using live NIH3T3 mouse fibroblast cells labeled with IgG-anti-EGF conjugated quantum dots. These same cells were then fixed, imbedded in resin, sectioned to 100nm thick sections and imaged under the electron microscope. The electron dense cadmium selinide provides the contrast necessary to perform direct imaging of EGF receptor sites. In order to improve the data and move toward multi-channel imaging in the electron microscope, EELS spectroscopy and elemental mapping of quantum dots was performed. The theory along with a discussion of the instruments needed to perform EELS is presented. This data was intended to provide new applications of quantum dots as biological probes in EELS spectroscopy. This dissertation should provide the reader with a thorough understanding of the nature of quantum dots and give a number of biological applications.