Establishment of New Micro-Computed Tomography Methodologies : The use of Histology as a Complementary Tool
Laguna-Castro, Santiago (2020)
2020
Master's Programme in Biomedical Sciences and Engineering
Lääketieteen ja terveysteknologian tiedekunta - Faculty of Medicine and Health Technology
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Hyväksymispäivämäärä
2020-12-11
Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:tuni-202010287652
https://urn.fi/URN:NBN:fi:tuni-202010287652
Tiivistelmä
Introduction: During the last decade, the study of biological structures in 3D has gained great importance due to its potential benefits in medical diagnosis and research. Micro-computed tomography (µCT) makes this approach possible when small samples like biopsies are studied. Moreover, it is considered a high-resolution and non-destructive method, but it is exempt from inconveniences. Imaging soft tissue requires the use of contrast agents that are typically toxic and can induce morpho-structural alteration on samples. This may hinder the interpretation of results and even make samples useless for later histological analysis, which is vital for validating new µCT methodologies and further research. On the other hand, human biopsies in research are essential for developing new human diagnostical methods. Nevertheless, their availability is low, and most of the time, they are donated included in paraffin (FFPE samples), which difficulties the intake of contrast agents solutions by the sample.
Objective: This thesis aimed to develop a new µCT method that could be applied to FFPE samples while using a minimally toxic contrast agent (eosin) that facilitates further histological analysis.
Experimental design: The study consisted of the development of a paraffin removal method suitable for µCT imaging; the optimization of the eosin staining protocol and µCT imaging; the evaluation of the quality of the resulting images and possible tissular damage; the descriptive analysis of the tissue; to assess the viability of doing histology after the µCT; and the comparison of this new approach with traditional IE2 - an iodine contrast agent- µCT imaging. FFPE musculoskeletal chicken muscle samples were used for those purposes. Once the eosin protocol was optimized, it was further applied in a FFPE human intestine sample.
Results: The developed paraffin removal method, combined with the eosin-based µCT protocol, generated acceptable µCT images. However, the contrasting capacities of IE2 remained higher than the eosin. Descriptive analysis and histology of muscular samples were limited due to their physical alteration; induced by their particular sensitivity to acetic acid and ethanol. In contrast, the µCT imaging and histology and descriptive analysis of the small intestine biopsy presented no significant difficulties.
Conclusion: The obtained results and bibliography points towards one major direction. This type of strategy strongly depends on the tissue's biological nature and structure of the study, which means that protocols should be adjusted on a sample-case basis.
Objective: This thesis aimed to develop a new µCT method that could be applied to FFPE samples while using a minimally toxic contrast agent (eosin) that facilitates further histological analysis.
Experimental design: The study consisted of the development of a paraffin removal method suitable for µCT imaging; the optimization of the eosin staining protocol and µCT imaging; the evaluation of the quality of the resulting images and possible tissular damage; the descriptive analysis of the tissue; to assess the viability of doing histology after the µCT; and the comparison of this new approach with traditional IE2 - an iodine contrast agent- µCT imaging. FFPE musculoskeletal chicken muscle samples were used for those purposes. Once the eosin protocol was optimized, it was further applied in a FFPE human intestine sample.
Results: The developed paraffin removal method, combined with the eosin-based µCT protocol, generated acceptable µCT images. However, the contrasting capacities of IE2 remained higher than the eosin. Descriptive analysis and histology of muscular samples were limited due to their physical alteration; induced by their particular sensitivity to acetic acid and ethanol. In contrast, the µCT imaging and histology and descriptive analysis of the small intestine biopsy presented no significant difficulties.
Conclusion: The obtained results and bibliography points towards one major direction. This type of strategy strongly depends on the tissue's biological nature and structure of the study, which means that protocols should be adjusted on a sample-case basis.