Wenger, M.D.; (2010) Micro-tip chromatography; a route to an integrated strategy for high throughput bioprocess development. Doctoral thesis , UCL (University College London).

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Abstract

Bioprocessing groups must keep pace with the many biologics and vaccines entering development, while ensuring process robustness, controlling costs, and accelerating project timelines. Microscale techniques provide a means to cope with these challenges by enabling high-throughput investigations to identify problems early, reduce requirements for costly large-scale experiments, and promote quality-bydesign approaches for process optimisation. Micro-tip columns (packed sorbent in a pipette tip) for chromatography and Adaptive Focused Acoustics (AFA) for cell disruption are two such techniques with potential to deliver high-throughput process development. This thesis characterises these platforms and integrates them as elements of the development workflow. Firstly, the key parameters are defined for robust, automated micro-tip chromatography. Finite-bath methods for isotherms and kinetic measurements are demonstrated, with sorbent contact time found to be critical for uptake of proteins on porous adsorbents, consistent with pore diffusion being rate-determining. Based upon these micro-tip data, two data-driven models are applied to predict dynamic binding capacity, one employing a shrinking-core model, and the other, a stagedreaction model. Both show satisfactory agreement with experimental laboratory column results. Micro-tip chromatography is then illustrated as an accelerated process development strategy for a mixed-mode chromatography step, with the results found to be predictive of laboratory column-scale yield, purity and capacity. In a second application, micro-tip chromatography is used to evaluate the interaction of upstream fermentation changes upon the downstream chromatography. The microscale chromatography is predictive of laboratory-scale yield and purity, despite being 1000-times smaller, while increasing productivity by over ten-fold. The miniaturisation of the chromatography, however, necessitates the development of a microscale cell disruption method to fully realise the gains in throughput and volume reduction. The AFA technique meets this goal, providing representative feed material for chromatographic study. Together, micro-tip chromatography and AFA form the basis for a next-generation bioprocess development platform.

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