Phoebe J. Welch

BioE PhD Defense Presentation

Friday, April 14th, 2023, at 2:30 pm

Location: IBB 1128

Zoom link: https://gatech.zoom.us/j/97986483276?pwd=bmdHK1NYSEVrNlBJbGl1MkVCamRaQT09

Advisors: Prof. Chengzhi Shi (Mechanical Engineering, Georgia Institute of Technology)

Prof. Craig R. Forest (Mechanical Engineering, Georgia Institute of Technology)

Committee:

Prof. Costas Arvanitis (Mechanical Engineering, Georgia Institute of Technology)

Prof. Brooks Lindsey (Biomedical Engineering, Georgia Institute of Technology)

Prof. Lilo D. Pozzo (Chemical and Biological Engineering, Univ. of Washington)

Prof. Levi Wood (Mechanical Engineering, Georgia Institute of Technology)

Ultrasound imaging of cells using gas vesicles and perfluorocarbon nanodroplets

Ultrasound imaging greatly benefits from the use of contrast agents to highlight regions of the body that typically exhibit low contrast. While gas microbubbles are the primary ultrasound contrast agent for certain medical imaging applications, they cannot be used for intracellular imaging because of their limited stability in physiological conditions. This thesis investigates two newer ultrasound contrast agents, gas vesicles and perfluorocarbon nanodroplets, that can be generated by cells or delivered intracellularly for localization of a specific cell type or a particular cell, respectively. We first explore mammalian acoustic reporter genes (mARGs), which enable gas vesicle expression in mammalian cells for localization in deep tissue. In Aim 1, we modify the original mARG construct to increase the proportion of cells that express gas vesicles by making the genes drug selectable, simplifying the process of creating a gas vesicle-expressing cell line with high ultrasound contrast. While mARGs are useful for imaging cell populations, the resulting gas vesicles do not produce sufficient ultrasound contrast to identify individual cells.  To achieve ultrasonic single cell localization, we shift our focus to PFCnDs. In Aim 2, we examine the lipid shell composition of PFCnDs to find an optimal ratio of lipid components that enable PFCnDs to generate ultrasound contrast with reduced risk of cell damage. In Aim 3, we microinject HEK293T cells with PFCnDs using patch clamp, noting the time and pressure parameters that result in successful nanodroplet injection, and demonstrate ultrasonic single-cell localization using this technique. These dissertation studies advance the use of both gas vesicles and perfluorocarbon nanodroplets as intracellular ultrasound contrast agents for various applications of cellular imaging.