Digital Protein Analysis: Technologies for Protein Diagnostics and Proteomics through Single-Molecule Detection

Protein quantification tools are valuable in biological and biomedical research and in clinical applications because they enable measurements that elucidate physiological states, including disease states. Protein biomarker detection is likely to play a major role in patient health and aid in the personalization of medicine. However, protein detection has lagged behind other bio-analytical methods due to intrinsic properties of proteins as well as the complexity of biological fluids, such as blood. This thesis describes the design, development, and testing of several technologies for the advancement of protein detection in clinical and research settings. A common thread through these technologies is the isolation of millions of protein molecules on a solid surface, the interrogation of those molecules by fluorescently-labeled antibodies, and the identification and quantification of those molecules by single molecule resolution imaging. By systematically investigating the parameters that have prevented single-molecule quantification of surface-immobilized proteins, we achieved technologies that can digitally quantify biomarkers in single-plex and multi-plex formats and be used to detect protein-protein interactions. Additionally we characterized a surface coating method that will be of benefit to a wide array of biophysical studies. Finally, we conducted proof-of-principle experiments and computer simulations for the development of a high-throughput proteomics technology that relies on only a small set of probes: 8 to 50) to analyze each of the approximately 23,000 human proteins.

Name thesis:degree_name

Doctor of Philosophy (PhD)

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biology and Biomedical Sciences: Computational and Systems Biology

Year dc:date.available

2011

Author dc:creator

• Tessler, Lee

Contributors dc:contributor

• Robi Mitra

• Molecular Biology
• Biochemistry
• Biophysics
• biomarkers
• diagnostics
• immunoassay
• microscopy
• proteomics
• sequencing

Language dc:language

English (en)

DOI

https://doi.org/10.7936/K70G3H46

OAI identifier oai:identifier

oai:openscholarship.wustl.edu:etd-1345