Measuring physical and chemical properties of biological nanoparticles in solution with interferometric scattering in an active feedback trap
Abstract/Contents
- Abstract
- Single-particle studies can unveil the heterogeneity and dynamics of not only single biomolecules and proteins, but also larger protein assemblies and biological nanoparticles. Previous work has developed the anti-Brownian electrokinetic (ABEL) trap that uses fluorescence to track the position of a single diffusing particle, and applies feedback in the form of electric fields to suppress the Brownian motion and allow for long observation times for mobile particles. This dissertation presents my work on developing an ABEL trap that uses interferometric scattering for tracking single nanoparticles, and on making physical and chemical measurements on those nanoparticles. I begin in Chapter 1 by describing the benefits and limitations of the ABEL trap, and some recent developments in interferometric scattering that motivate us to use it for tracking particles. I then sketch out the concept of an interferometric scattering ABEL trap (or ISABEL trap), including some of the new studies which it should enable. I also describe biological nanoparticles from autotrophic bacteria called carboxysomes, which we study with the ISABEL trap. These self-assembled particles create a distinct chemical environment for the efficient action of the enzyme rubsico to fix organic carbon and are responsible for 10-25% of CO2 fixation on Earth. I discuss the theoretical aspects of interferometric scattering as relevant to our experiments in Chapter 2, describing the image formation process and the sources of noise. I then utilize this information in Chapter 3, where I describe the experimental implementation of the ISABEL trap. I describe three different setups that iteratively improved the sensitivity and capabilities of the instrument, broken down into the different components of the experimental setup. I then describe our experiments on carboxysomes in the ISABEL trap, and the physical and chemical information we can now access. Chapter 4 describes our work on measuring the redox environment inside individual carboxysomes with a redox sensitive fluorescent protein using ratiometric readout. This work takes advantage of the independent fluorescence channels opened up by interferometric scattering detection. Chapter 5 describes our work on measuring the mass of single carboxysomes, including measurements of single-particle scattering cross-sections, and the use of a simultaneous fluorescence channel to infer the mass distribution inside carboxysomes. This work takes advantage of the quantitative aspects of the scattering measurement, and its label-free nature. Finally, in Chapter 6, I describe the outlook for the ISABEL trap, including comparing the current and possible capabilities to other experiments, looking at other samples of interest, and mentioning future capabilities that show promise if developed.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2023; ©2023 |
| Publication date | 2023; 2023 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Lavania, Abhijit |
|---|---|
| Degree supervisor | Fejer, Martin M. (Martin Michael) |
| Degree supervisor | Moerner, W. E. (William Esco), 1953- |
| Thesis advisor | Fejer, Martin M. (Martin Michael) |
| Thesis advisor | Moerner, W. E. (William Esco), 1953- |
| Thesis advisor | Boxer, Steven G. (Steven George), 1947- |
| Degree committee member | Boxer, Steven G. (Steven George), 1947- |
| Associated with | Stanford University, School of Humanities and Sciences |
| Associated with | Stanford University, Department of Applied Physics |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Abhijit A. Lavania. |
|---|---|
| Note | Submitted to the Department of Applied Physics. |
| Thesis | Thesis Ph.D. Stanford University 2023. |
| Location | https://purl.stanford.edu/nx809sw9031 |
Access conditions
- Copyright
- © 2023 by Abhijit Lavania
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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