The Study of Protein and Polymer Phase Separation Kinetics and Thermodynamics Using Temperature Gradient Microfluidics
Restricted (Penn State Only)
- Author:
- Graff, Erica
- Graduate Program:
- Chemistry
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- June 05, 2023
- Committee Members:
- Paul Cremer, Chair & Dissertation Advisor
Philip Bevilacqua, Major Field Member
Alexey Silakov, Major Field Member
Darrell Velegol, Outside Unit & Field Member
Philip Bevilacqua, Program Head/Chair - Keywords:
- phase separation
monoclonal antibodies
temperature gradient microfluidics
apparent activation energies - Abstract:
- This dissertation consists of works concerned with the liquid-liquid phase separation of proteins, protein models, and polymers. The projects involving protein use a monoclonal antibody as the model system for studying phase separation, and how phase separation can be altered based on solution conditions such as pH and the concentration or identity of salt. The methods used to investigate these systems include temperature gradient microfluidics, temperature ramp experiments, UV/Vis, fluorescence microscopy, differential scanning calorimetry, high accuracy particle counting, long-term aggregation studies, and microscale nephelometry. Temperature gradient microfluidics (TGM) was the primary method in most of this work due to the unique parameters that it is able to collect in a high-throughput manner. Both kinetic and thermodynamic data can be acquired from these experiments and have been used to characterize phase separating systems. Poly(ethylene glycol) was used as a crowder in order to induce phase separation in antibody solutions. Through this work, we found that two different excipients have distinct effects on the phase separation data produced by TGM experiments, which opens the door for future work in exploring how cosolute interactions alter phase separation behavior. We also found that parameters from temperature gradient experiments correlated well with diffusion interaction parameters and long term stability studies, and poly(ethylene glycol) will phase separate in the presence of salt and partition certain food dyes.