{"id":{"repo_id":"cambridge","oai_identifier":"oai:www.repository.cam.ac.uk:1810/294572"},"canonical_url":"https://search.dev.ndltd.org/etd/cambridge/oai:www.repository.cam.ac.uk:1810/294572","repository":{"repo_id":"cambridge","name":"Cambridge University","base_url":"https://api.repository.cam.ac.uk/server/oai/request"},"display":{"title":"Light-sheet microscopy used for tracking particles","abstract":"Fluorescence microscopy is one of the cornerstones of modern biology but has generally been limited to 2D culture dishes. Light-sheet microscopy, a recent advance which was awarded Nature Method of the Year in 2014, allows fast, non-invasive 3D imaging across an entire organism. This works by decoupling illumination and detection such that the microscope only illuminates a thin section of tissue at a time. By scanning this light-sheet through an organism we can image in 3D, more quickly and with less damage than other techniques such as confocal microscopy. In this work, a custom digitally scanned light-sheet microscope was built, for which the technology was applied and developed to enable two biological studies: the study of material properties of developing embryos and the tracking of virus particles in live cells. In addition to designing and constructing a light-sheet fluorescence microscope, several technological improvements were also investigated to better address these biological questions. The first was a three-dimensional region-of-interest technique which greatly simplifies volumetric imaging calibration whilst also being more robust, with an observed 42% improvement in light collection efficiency compared to current approaches. The projective mathematical theory, used in this technique, was then applied to optical projection tomography to produce a new triangulation-based reconstruction algorithm that is robust to affine sample motion, including mechanical jitter and systematic drift. The second improvement for light-sheet microscopy builds upon confocal slit scanning, a technique used to increase image contrast whilst doubling the acquisition time for a single image. By exploiting the acquisition procedure for confocal slit scanning, full speed imaging with the same increased contrast was realised. Finally an open-hardware solution for multi-scale sample mounting was produced. These improvements to speed, contrast and acquisition speed in the light-sheet microscope allowed us to address the biological questions of interest.","abstract_html":"Fluorescence microscopy is one of the cornerstones of modern biology but has generally been limited to 2D culture dishes. Light-sheet microscopy, a recent advance which was awarded Nature Method of the Year in 2014, allows fast, non-invasive 3D imaging across an entire organism. This works by decoupling illumination and detection such that the microscope only illuminates a thin section of tissue at a time. By scanning this light-sheet through an organism we can image in 3D, more quickly and with less damage than other techniques such as confocal microscopy. In this work, a custom digitally scanned light-sheet microscope was built, for which the technology was applied and developed to enable two biological studies: the study of material properties of developing embryos and the tracking of virus particles in live cells. In addition to designing and constructing a light-sheet fluorescence microscope, several technological improvements were also investigated to better address these biological questions. The first was a three-dimensional region-of-interest technique which greatly simplifies volumetric imaging calibration whilst also being more robust, with an observed 42% improvement in light collection efficiency compared to current approaches. The projective mathematical theory, used in this technique, was then applied to optical projection tomography to produce a new triangulation-based reconstruction algorithm that is robust to affine sample motion, including mechanical jitter and systematic drift. The second improvement for light-sheet microscopy builds upon confocal slit scanning, a technique used to increase image contrast whilst doubling the acquisition time for a single image. By exploiting the acquisition procedure for confocal slit scanning, full speed imaging with the same increased contrast was realised. Finally an open-hardware solution for multi-scale sample mounting was produced. These improvements to speed, contrast and acquisition speed in the light-sheet microscope allowed us to address the biological questions of interest.","abstract_has_math":false,"creators":["Russell, Craig Terence"],"institution":"University of Cambridge","degree_name":"Doctor of Philosophy (PhD)","degree_level":"Doctoral","degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":["Rees, Eric John"],"committee_chairs":[],"committee_members":[],"year":2019,"date_issued":"2019-07-19","date_published":"2019-07-19","updated_at":"2026-07-22T22:24:30Z","subjects":["light-sheet","microscopy","pose-estimation","optical projection tomography","particle tracking","spt","fluorescence"],"languages":["en"],"rights":[],"rights_urls":["https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/925b07b3-2c99-4f62-a9c4-d1fcfe7cfc7b/download","https://www.rioxx.net/licenses/all-rights-reserved/"],"identifier_entries":[]},"links":{"outbound_url":"https://doi.org/10.17863/CAM.41677","outbound_label":"DOI","outbound_source":"dc:identifier.doi"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.advisor","label":"Advisor","values":["Rees, Eric John"]},{"key":"dc:contributor.sponsor","label":"Sponsor","values":["EPSRC - Integrated Photonics and Electronics CDT"]},{"key":"dc:creator","label":"Author","values":["Russell, Craig Terence"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.issued","label":"Date","values":["2019-07-19"]},{"key":"dc:publisher.institution","label":"Dc Publisher Institution","values":["University of Cambridge"]},{"key":"dc:relation.isreferencedby.uri","label":"Dc Relation Isreferencedby URI","values":["https://www.repository.cam.ac.uk/handle/1810/294572"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"dc:type.qualificationlevel","label":"Dc Type Qualificationlevel","values":["Doctoral"]},{"key":"dc:type.qualificationname","label":"Dc Type Qualificationname","values":["Doctor of Philosophy (PhD)"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["light-sheet","microscopy","pose-estimation","optical projection tomography","particle tracking","spt","fluorescence"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["en"]},{"key":"dc:rights","label":"Dc Rights","values":["https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/925b07b3-2c99-4f62-a9c4-d1fcfe7cfc7b/download","https://www.rioxx.net/licenses/all-rights-reserved/"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.doi","label":"DOI","values":["10.17863/CAM.41677"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/0d4823fe-27c8-4128-ac0e-196932eb4f55/download"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Fluorescence microscopy is one of the cornerstones of modern biology but has generally been limited to 2D culture dishes. 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The first was a three-dimensional region-of-interest technique which greatly simplifies volumetric imaging calibration whilst also being more robust, with an observed 42% improvement in light collection efficiency compared to current approaches. The projective mathematical theory, used in this technique, was then applied to optical projection tomography to produce a new triangulation-based reconstruction algorithm that is robust to affine sample motion, including mechanical jitter and systematic drift. The second improvement for light-sheet microscopy builds upon confocal slit scanning, a technique used to increase image contrast whilst doubling the acquisition time for a single image. By exploiting the acquisition procedure for confocal slit scanning, full speed imaging with the same increased contrast was realised. Finally an open-hardware solution for multi-scale sample mounting was produced. 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