Computed tomography based spectral imaging for fluorescence microscopy

Persistent Link:
http://hdl.handle.net/10150/280122
Title:
Computed tomography based spectral imaging for fluorescence microscopy
Author:
Ford, Bridget K.
Issue Date:
2002
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
Multispectral imaging has been used for decades in remote sensing to enhance the classification, discrimination and characterization of materials. Only recently has this same technology been similarly applied to fixed biological samples in cytogenetics, pathology and medicine. A further extension to in vivo studies is often limited by the low levels of associated fluorescence as well as the increased temporal resolution required to analyze physiological changes. In addition, the cellular response to a specific agonist is often heterogeneous across the cellular field requiring a combination of sufficient spatial and temporal resolutions. A computed tomography imaging spectrometer (CTIS) has been developed which overcomes these limitations by simultaneously collecting extended range spectral information (470-740 nm, 5 nm sampling) across a 2-D field of view (200 μm x 200 μm, 0.96 μm sampling). The CTIS uses a computer generated hologram to produce a 5 x 5 array of images with differing amounts and directions of dispersion. This set of images allows the 3-D signal (x, y, λ) from a fluorescent sample to be mapped onto a 2-D detector array. In this way, the full spectral and spatial information is acquired for a 2-D cellular field during a single integration time (presently 2 sec for biological specimens). The CTIS's design, calibration, and underlying theory are described in detail. In addition, the capability of the CTIS to simultaneously collect the fluorescence emission of multiple fluorophores across a 2-D cellular field is demonstrated. Specifically, the combined spectral variations of seminapthorhodafluor-I and enhanced green fluorescent protein were followed in rat insulinoma cells in order to extend the linear range of intracellular pH detection.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Engineering, Biomedical.; Physics, Optics.
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Graduate College; Optical Sciences
Degree Grantor:
University of Arizona
Advisor:
Descour, Michael R.

Full metadata record

DC FieldValue Language
dc.language.isoen_USen_US
dc.titleComputed tomography based spectral imaging for fluorescence microscopyen_US
dc.creatorFord, Bridget K.en_US
dc.contributor.authorFord, Bridget K.en_US
dc.date.issued2002en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractMultispectral imaging has been used for decades in remote sensing to enhance the classification, discrimination and characterization of materials. Only recently has this same technology been similarly applied to fixed biological samples in cytogenetics, pathology and medicine. A further extension to in vivo studies is often limited by the low levels of associated fluorescence as well as the increased temporal resolution required to analyze physiological changes. In addition, the cellular response to a specific agonist is often heterogeneous across the cellular field requiring a combination of sufficient spatial and temporal resolutions. A computed tomography imaging spectrometer (CTIS) has been developed which overcomes these limitations by simultaneously collecting extended range spectral information (470-740 nm, 5 nm sampling) across a 2-D field of view (200 μm x 200 μm, 0.96 μm sampling). The CTIS uses a computer generated hologram to produce a 5 x 5 array of images with differing amounts and directions of dispersion. This set of images allows the 3-D signal (x, y, λ) from a fluorescent sample to be mapped onto a 2-D detector array. In this way, the full spectral and spatial information is acquired for a 2-D cellular field during a single integration time (presently 2 sec for biological specimens). The CTIS's design, calibration, and underlying theory are described in detail. In addition, the capability of the CTIS to simultaneously collect the fluorescence emission of multiple fluorophores across a 2-D cellular field is demonstrated. Specifically, the combined spectral variations of seminapthorhodafluor-I and enhanced green fluorescent protein were followed in rat insulinoma cells in order to extend the linear range of intracellular pH detection.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectEngineering, Biomedical.en_US
dc.subjectPhysics, Optics.en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.disciplineOptical Sciencesen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.advisorDescour, Michael R.en_US
dc.identifier.proquest3061006en_US
dc.identifier.bibrecord.b43042338en_US
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