Light-sheet microscopy used for tracking particles
type: CreativeWork
description: 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-she…
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-she…
author: Russell, Craig Terence
language: en
keywords: light-sheet, microscopy, pose-estimation, optical projection tomography, particle tracking, spt, fluorescence
license: https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/925b07b3-2c99-4f62-a9c4-d1fcfe7cfc7b/download
license: https://www.rioxx.net/licenses/all-rights-reserved/
date published: 2019-07-19
date modified: 2026-07-22T22:24:30Z
provider: Cambridge University
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Cambridge

PropertyValue
property ID: DOI
value: 10.17863/CAM.41677