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Title: Antibody stability in bioprocessing focusing on shear effects at solid-liquid interfaces
Author: Biddlecombe, J. G.
Awarding Body: University College London (University of London)
Current Institution: University College London (University of London)
Date of Award: 2009
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Exposure to high shear during bioprocessing operations has been reported to be associated with losses in protein stability and lead to aggregation, rendering a product unfit for use. High shear forces occur in many process operations, therefore it is important to understand the influence of shear and to control its effects during bioprocessing. The purpose of this work was to assess the stability of human monoclonal antibodies of IgG4 isotype using a device designed to generate defined, quantifiable levels of shear in the presence of a solid-liquid interface. The device, based on a rotating disk, produced shear strain rates of up to 3.4x10^4 s^-1 and was designed to exclude air-liquid interfaces. Computational fluid dynamics (CFD) was used to study the fluid flow patterns within the devices and to determine the shear strain rate (s^-1) at a range of disk speeds. The structural integrity of the IgG4 after exposure to interfacial shear effects was studies by SDS-PAGE, IEF, dynamic light scattering, and peptide mapping by LC-MS. This analysis revealed that the main denaturation pathway of IgG4 exposed to these effects was the formation of large insoluble aggregates. Soluble aggregation, breakdown in primary structure, and chemical modifications were not detected. Factor associated with the solution conditions (pH, ionic strength, surfactant concentration, temperature), and the interface (surface roughness, and hydrophobicity) were studies for their effect on the rate of IgG4 monomer loss under high shear conditions. The dominant factors found to affect the rate of monomer loss under interfacial shear conditions were found to be pH and the nanometre-scale surface roughness associated with the solid-liquid interface. The addition of surfactant was found to have a significant stabilising effect at concentrations up to 0.02%(w/v). These studies highlight that the interaction of shear and protein-surface interactions are important factors to consider when designing stability studies for protein production.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available