| This dissertation describes the development of semiconductor colloidal nanocrystals (specifically, ZnS-capped CdSe) for biological labeling. Prior to my graduate studies, the main focuses of research in the area of colloidal nanoclusters were on understanding their optical properties and improving their synthetic procedures, as indicated by large numbers of published papers (well over 2,000). In the literature, numerous researchers demonstrated unique luminescence properties of these nanocrystals such as narrow spectral linewidths, sloping absorbance profiles, and size-tunable emission. All of these properties would be advantageous for probe-based applications, for example, biological and chemical sensing. Not until recently (ca. 1996) have high quantum yield (30–50%) quantum dots been successfully synthesized. For these quantum dots to be used for biological detection, several challenges had to be overcome. First, these nanocrystals were synthesized in an organic solvent and therefore, they were only soluble in non-polar solvents, such as chloroform and hexane. Second, biomolecules such as oligonucletides and proteins must be conjugated onto their surface.; Modification of the colloid's surface chemistry was used to produce water-soluble quantum dots, in which bifunctional molecules (e.g., mercaptoacetic acid) displaced the tri-n-ocytylphosphine oxide (TOPO) stabilizing ligand. The TOPO resided on the nanocrystal surface and was responsible for their hydrophobic characteristics. Biomolecules containing primary amines (three examples were demonstrated in this thesis) were conjugated onto the nanoparticle, using carbodiimide catalysis (commonly used for labeling fluorescent ligands to proteins and oligonucleotides). These conjugated biomolecules were still functional, as demonstrated by the endocystosis of transferrin-coated quantum dots or the binding of anti-IgG labeled nanocrystals to IgG in a latex agglutination test.; Furthermore, the effect of parameters (e.g., pH and temperature) that are important in biological experiments on the optical properties of semiconductor quantum dots was investigated. It was demonstrated that the fluorescence quantum efficiency and photobleaching rates were related to the pH, with the highest obtained values at basic pHs. Temperature also influenced the quantum efficiency, as determined by a linear decrease in fluorescence intensity as the temperature increased. |
