Landes, Christy F2016-01-152016-01-152015-052015-04-20May 2015Kisley, Lydia. "Single Molecule Studies of Ion-Exchange Chromatography." (2015) Diss., Rice University. <a href="https://hdl.handle.net/1911/87860">https://hdl.handle.net/1911/87860</a>.https://hdl.handle.net/1911/87860As the pharmeceutical industry moves away from traditional small organic molecules towards biologically-based treatments, ion-exchange separation methods must be investigated to improve the cost and time required for protein purification. Several new single molecule, super-resolution techniques are presented to offer a mechanistic experimental understanding of chromatography unachievable through traditional ensemble-averaged methods. Super-resolution analysis visualizes single protein adsorption kinetics to single, super-resolved ligands, allowing for the first experimental validation of the statistical mechanical stochastic theory of chromatography. Imperative results on the spatial charge-distribution of ligands, reduction of heterogeneity by ionic strength, and tuning of protein/stationary phase interfacial interactions by pH are observed. A common finding that the sterics of the agarose support induces separation heterogeneity leads to super-resolution imaging of the agarose structure and diffusion properties. Finally, the single molecule techniques are applied to several applications beyond protein chromatography to demonstrate the potential for future materials research. Overall, we have shown that single molecule spectroscopy can aid in the mechanistic experimental and theoretical understanding of the ion-exchange chromatographic separation of proteins.application/pdfengCopyright is held by the author, unless otherwise indicated. Permission to reuse, publish, or reproduce the work beyond the bounds of fair use or other exemptions to copyright law must be obtained from the copyright holder.super-resolution spectroscopystochastic theorybioseparationsagaroseThesis2016-01-15