Skip to main content
eScholarship
Open Access Publications from the University of California

UC San Diego

UC San Diego Electronic Theses and Dissertations bannerUC San Diego

Solid Substrate for High Throughput Protein Chromatography

  • Author(s): Alvarez, Adrian
  • Advisor(s): Kummel, Andrew C
  • et al.
Abstract

Protein A chromatography is the industry standard method for the purification of monoclonal antibodies. In this work, an emulsion templated sol-gel reaction is used to synthesize rigid, millimeter-scale SiO2 particles with hierarchical porosity, presenting an alternative to existing protein A chromatography resins for high throughput purification. An average particle diameter of 0.61mm allows for increased packed bed length/width ratios and high mobile phase flow rates to be used with minimal pressure drop. FITC conjugated BSA and fluorescence microscopy are used to show that micron-scale, spherical pores (average diameter = 4.69µm) allow for unhindered diffusion of protein throughout the particles. High temperature calcination (1000°C) is used to induce SiO2 reflow and increase intraparticle wall density (decreasing wall thickness from 1.48µm to 1.04µm, p = 0.014), significantly increasing the overall particle strength. A BCA assay and fluorescence spectroscopy are used to show that treating the particles with concentrated H2O2 increases the immobilized protein ligand density from 6.8 to 24.0 mg protein/mL particles due to an increase in SiO2 surface hydroxyl density. Initial protein separation testing shows poor column efficiency with early protein breakthrough, suggesting mobile phase channeling between particles and a need to sieve the particles to decrease the size distribution.

Main Content
For improved accessibility of PDF content, download the file to your device.
Current View