Abstract

Traditional proteomics studies can measure many protein biomarkers simultaneously from a single patient-derived sample, promising the possibility of syndromic diagnoses of multiple diseases sharing common symptoms. However, precious cellular-level information is lost in conventional bulk-scale studies that measure tissues comprising many types of cells. As single cells are the building blocks of organisms and are easier to biopsy than traditional bulk samples, performing proteomics on a single-cell level would benefit clinicians and patients. Single-cell proteomics, combined with mass spectrometry imaging, can be used to analyze cells in their microenvironment, preserving spatial information. We have previously used laser-capture microdissection to isolate single motor neurons from tissue and analyze them in our single-cell proteomics platform. However, our sampled population of cells was necessarily limited by the low throughput of the measurement platform, and by the sensitivity of our liquid chromatography-mass spectrometry system to debris introduced in the laser-capture microdissection isolation workflow. In the work described in this dissertation, we dramatically improved the throughput of single-cell proteomics, created a method for removing insoluble debris that clogged our liquid chromatography-mass spectrometry system, and developed a high-performance, low-cost method for nanoflow gradient formation. Together, these methodologies will increase the depth of information and the number of biological replicates that can measured in single-cell proteomics. We hope that these technologies will be applied to future liquid chromatography systems to enable large scale single-cell proteomics studies of tissues. This will reveal the cellular origins of disease on a multimolecular level, while keeping important spatial information. Thus, we expect the technologies and ideas developed here to play a key role in understanding the cellular proteomics in biomedical and clinical settings.

Degree

PhD

College and Department

Physical and Mathematical Sciences; Chemistry and Biochemistry

Rights

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