Fluorescence correlation spectroscopy has become a standard technique for modern biophysics and single molecule spectroscopy research. Here is presented a novel widefield extension of the established single-point technique. Flow in microfluidic devices was used as a model system for microscopic motion and through widefield fluorescence correlation spectroscopy flow profiles were mapped in three dimensions. The technique presented is shown to be more tolerant to low signal strength, allowing image data with signal-to-noise values as low as 1.4 to produce accurate flow maps as well as utilizing dye-labeled single antibodies as flow tracers. With proper instrumentation flows along the axial direction can also be measured. Widefield fluorescence correlation spectroscopy has also been utilized to produce super-resolution confocal microscopic images relying on the single-molecule microsecond blinking dynamics of fluorescent silver clusters. A method for fluorescence modulation signal extraction as well as synthesis of several novel noble metal fluorophores is also presented.
| Identifer | oai:union.ndltd.org:GATECH/oai:smartech.gatech.edu:1853/33849 |
| Date | 26 March 2010 |
| Creators | Nicovich, Philip R. |
| Publisher | Georgia Institute of Technology |
| Source Sets | Georgia Tech Electronic Thesis and Dissertation Archive |
| Detected Language | English |
| Type | Dissertation |
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