Development of FTIR tomography for thermal-source imaging of 3D biochemical distributions in micro-samples of cells and fibers
FTIR microspectroscopy is an established 2D hyperspectral imaging technique with which to measure distributions of biochemical functional groups (e.g. lipids, proteins, nucleic acids) within organic samples. The emergence of FTIR microtomography, first reported by Martin et al. (2013), extends image...
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| Other Authors: | , |
| Format: | Doctoral or Postdoctoral Thesis |
| Language: | English |
| Published: |
2018
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| Online Access: | http://hdl.handle.net/1993/33025 https://doi.org/10.5203/THESIS_FINDLAY_1 |
| Summary: | FTIR microspectroscopy is an established 2D hyperspectral imaging technique with which to measure distributions of biochemical functional groups (e.g. lipids, proteins, nucleic acids) within organic samples. The emergence of FTIR microtomography, first reported by Martin et al. (2013), extends image resolution to three spatial dimensions (3D). This thesis describes a generalized, laboratory-scale approach to 3D imaging with FTIR microtomography. A modified method to handle a variety of micro-sample morphologies, i.e. longer fibres (spider silk) and larger globular cells (diatoms and buccal cells), was required. Towards this end, a motorized sample holder with increased flexibility was designed at the University of Manitoba for the collection of thermal source FTIR microtomographic data sets. A tomography accessory for microscopes (US patent No. US15065379; June, 2017) was prototyped, and assessed via imaging of a custom built size-standard phantom. The tomography accessory was further used to collect data illustrating niche applications of FTIR microtomography. The 3D resolved FTIR spectra (voxel spectra) of a fiber and two cell types of interest, namely artificial spider silk, human buccal epithelial cells and Arctic sea ice diatoms, were characterized. The 3D distribution and abundance of compounds were reconstructed while maintaining a sub-cellular level of resolution in all three spatial dimensions. Specifically, the embedding of the silk fiber in a refractive-index matched transparent matrix reduced scatter increased the quality of 3D FTIR images. Additional fine details of these silk specimens not observable with 2D FTIR images, e.g. double-stranded morphological substructure, were captured with FTIR microtomography. The application of a coating was non-destructive and reversible. Further polarization contrast FTIR microtomographic imaging of the coated spider silk revealed sub-volumes within the fiber with differing responses to polarized IR light. October 2018 |
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