Author ORCID Identifier

https://orcid.org/0000-0001-9006-7345

Date of Award

Spring 6-13-2026

Document Type

Thesis (Ph.D.)

Department or Program

Engineering Sciences

First Advisor

Scott C. Davis

Abstract

Fluorescence imaging has become a powerful tool in biomedical research and clinical practice, offering molecular specificity and high sensitivity for visualizing biological processes in real time, However, the successful translation of novel fluorescent probes or medicines from preclinical development to clinical applications requires rigorous validation of their biodistribution and target specificity. Traditional methods for assessing performance, such as bulk excised organ analysis or two-dimensional histology, provide limited information as they cannot capture the complete, organism-wide distribution of these probes or medicines. This thesis aims to improve this analysis pipeline through the application and optimization of fluorescence cryotomography for quantitative, high-resolution imaging of fluorescence through the whole-body.

Our fluorescence cryotomography (FCT) approach combines an automated serial sectioning system with a hyperspectral fluorescence imaging system to generate volumetrics datasets that map spatial distribution of fluorescence throughout an entire specimen. The use of the platform is demonstrated through its application in gene therapy and for the advancement of novel fluorescent contrast agents. Specifically, we use whole-body FCT in combination with a Cre-inducible Ai14 reporter animal to map the functional transduction efficiency and potential off-target biodistributions of adeno-associated viral (AAV) vectors. Second, in fluorescence-guided surgery (FGS), FCT is applied to validate the targeting efficiency of novel tumor-targeting and nerve-targeting probes. We also demonstrate the application of these nerve-targeting probes for sub-surface depth estimation in tissue-simulating phantoms and in animal models.

We also report on the development of a high-grade glioma model using a Cre-inducible Oncopig platform using AAVs. This model offers a pre-clinical platform for assessment of new technologies that require a surgical setting, which we demonstrate with fluorescence-guided resection of the induced glioma in an animal.

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