Date of Award

Spring 2025

Document Type

Thesis - SCU Access Only

Publisher

Santa Clara : Santa Clara University, 2025

Department

Bioengineering

First Advisor

Bill Lu

Abstract

Current protein quantification methods including Western Blotting, ELISA, and fluorescence tagging, suffer from a variety of limitations such as spatial resolution, lack of real-time monitoring, and only end-point assays. In order to address such concerns, we have developed genetic sensors containing Gaussia luciferase (gLuc) and green fluorescent protein (GFP) in order to quantify protein storage. Each of our sensors contains different signaling peptides to target a variety of subcellular compartments. These sensors are transfected into human 293T with polyethylenimine (PEI), followed by a confocal microscopy and luciferase assays. In order to verify the results of our imaging analysis in human 293T cells, we tested the same genetic sensors in other human HepG2 cells, ensuring that protein localization is constant throughout the study. Using the Gluc marker as part of the luciferase assay, allows us to see protein localization and to determine how much protein is in each of the compartments. Our dual reporter system aims to solve the weaknesses of traditional quantification methods by providing a precise, sensitive, and real time method for quantifying protein storage that have implications for protein therapeutics and overall drug development in the biotechnology industry.

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