Optical Scatter Imaging Using Digital Fourier Microscopy

Katrina Seet (2008). Optical Scatter Imaging Using Digital Fourier Microscopy PhD Thesis, School of Physical Sciences, The University of Queensland.

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Author Katrina Seet
Thesis Title Optical Scatter Imaging Using Digital Fourier Microscopy
School, Centre or Institute School of Physical Sciences
Institution The University of Queensland
Publication date 2008-10-01
Thesis type PhD Thesis
Supervisor Dr. Andrei Zvyagin
Dr. Timo Nieminen
Dr. Tim McIntyre
Dr. Aleksandar Rakic
Total pages 203
Total colour pages 20
Total black and white pages 183
Subjects 240000 Physical Sciences
Formatted abstract
Although several optical microscopy techniques are available, many of these are not suitable
for biological materials. Biological materials are generally transparent, showing little image
contrast between internal structures. In addition to this limitation, the highly scattering nature
of cellular structures within biological tissues also obscures fine details. Many techniques
attempt to reduce the unwanted background noise of images. However such techniques can
also distort images such that the true geometry of the specimen may not be represented.
Staining can also aid in the enhancement of specimen contrast, although these can damage,
and sometimes, destroy the specimen. Existing microscopy methods are, therefore, unsuitable
for some biological specimens.
The digital Fourier microscopy (DFM) technique was developed for the enhancement of image
contrast and Fourier filtering at the post-acquisition stage. The technique is sensitive to
the smaller sized scatterers within the sample. This sensitivity is one advantage of recording
in the Fourier domain. In addition, the incorporation of digital holography also permits
Fourier filtering where the form-factor, related to the Fourier transform of scatterers, can be
extracted. This data can be used for object characterisation by modeling, i.e., it becomes
an inverse scattering problem. Recorded digital holograms have the additional flexibility of
digital filtering. This allows the design of custom filters where the scatterers themselves can
be used to create masks for further enhancement of the signal-to-background ratio, i.e., the
contrast, of specific scatterers within the specimen. The principle of DFM is demonstrated
through numerical simulations and application to biological phantoms.
Biological phantoms were fabricated and independently characterised to demonstrate the selective imaging and refractometry capability of the DFM technique. Phantoms were created to mimic cell nuclei embedded in a cytoplasm matrix. Polystyrene and organosilica
microspheres were embedded in thin films of polyvinyl alcohol. These were spun-coated
onto thin microscope coverslips. To verify accuracy in DFM measurements, laser tweezer
refractometry and scattering spectroscopy were also used to determine the refractive index
of polystyrene and organosilica microspheres. An adaptation to the scattering spectroscopy
technique had been developed for this purpose (Seet et al., 2005). Extinction spectra of
dilute suspensions of monodisperse spheres were recorded and their spectroscopic signatures
were used to determine the size and refractive index. To illustrate the capabilities of DFM,
the refractometry on melanocyte cells was also conducted.
We envisage many other biomedical applications of DFM. The DFM technique could also be
used for studying the onset and early stages of cell calcification in cell cultures as in coronary
artery disease (Campbell and Campbell, 2000). The progression of calcification can therefore
be monitored by refractive index measurements of the cell culture constituents. The use of
DFM for the determination of the refractive index and diameter of polymer beads may also
assist the refinement of existing DNA sequencing methods (Kae, 2003). Currently, this is
limited by the use of colloid dye-doped beads (Battersby et al., 2002; Finkel et al., 2004).
Additional Notes 8, 15, 24, 28, 37, 45, 58, 67, 74-75, 89, 94-95, 97, 100-101, 127, 136-137, 139

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Created: Fri, 19 Dec 2008, 13:56:28 EST by Miss Katrina Seet on behalf of Library - Information Access Service