The conventional light microscope is an indispensable tool for many physical and life science applications, but is of limited usefulness for three-dimensional imaging due to its increasingly narrow depth of field at high magnifications. Focused regions may be obscured by defocused neighbors or noise from extraneous light sources and subsurface scattering. By rejecting light originating from outside the depth of focus it is possible to minimize these problems. When a contiguous series of such focused slices, or optical sections, are obtained along an axis of an extended object they may be combined to form a complete, focused three-dimensional surface image. Here, a variety of methods to obtain optical sections in a reflective setup are presented. The first employs an optical feedback loop through a spatial light modulator (SLM) to selectively illuminate focused regions. The SLM is a flexible electro-optical device that also allows (non-feedback) experiments of an intensity modulated light source resulting in illumination with a linear structure. This structured illumination microscopy is an established sectioning technique, which requires three frame captures per axial position. By developing a color grid and exploiting the red, green, and blue channels of a CCD camera, the three frames have been reduced to one. The speed increase comes at a cost and the limiting effects of chromatic aberration are discussed. Digital holography offers an alternative to axial scanning by allowing the surface to be reconstructed from a single exposure. Use of multiple wavelength illumination with this extended focus imaging is proposed and preliminary results are shown.
| Identifer | oai:union.ndltd.org:USF/oai:scholarcommons.usf.edu:etd-3592 |
| Date | 01 June 2006 |
| Creators | Krzewina, Leo G |
| Publisher | Scholar Commons |
| Source Sets | University of South Flordia |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Graduate Theses and Dissertations |
| Rights | default |
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