![]() ![]() Pressure-flow and DMA characterisation were used to examine the mechanical properties of the cycled resins to provide the first systematic study of these issues. The results indicate that the interaction of CIP reagents and foulants (as opposed to CIP reagents alone) cause the greatest impact on the structural integrity of the resins. In the case of Capto Adhere (highly cross-linked multimodal anion exchange resin) and MabSelect Xtra (highly cross-linked protein A resin), the mechanism of aging appeared to be associated foulants coating the surface fibres. ![]() Their matrices showed agarose fibre breakage with increased exposure to process conditions. ![]() The results indicated that MabSelect (highly cross-linked protein A resin) and Q-Sepharose High Performance (anion exchange cross-linked resin) appeared to show similar mechanisms of aging. The first set of experiments exposed the resins to the cleaning cycle only, whilst in the second set of experiments, the resins had been used for lifetime studies in the production of monoclonal antibodies (termed ‘aged’ resins). These studies expose the resins to repeated cycles of use to understand how they age in a particular bioprocess, enabling decision making about their use. The structural imaging and mechanical testing tools developed in this study were then applied to measure changes in resins that had undergone lifetime studies. These results indicate that DMA can be used as a small volume, high-throughput technique, relative to pressure-flow analysis, for the mechanical characterisation of chromatography media. The same trends were observed – The Capto family showed the highest resistance to deformation (Capto Adhere- 2.7, Capto Q- 1.92 1/%min-1) through to Sepharose CL-6B, Sepharose 4 Fast Flow and Sepharose CL-4B which exhibited the lowest slurry resistances (0.59, 0.4, 0.3 1/%min-1 respectively). The technique was applied to the nine studied resins and correlated with the results obtained using the pressure-flow technique. Dynamic mechanical analysis (DMA) was therefore developed as a novel technique in this field to address these limitations and allowed for further mechanical characterisation based on the viscoelastic properties using 1ml of resin. There were practical limitations in using the pressure-flow technique alone to for mechanical characterisation, including the large quantity of chromatography resin and buffers and the stringent criteria required to pack a column. The results showed that the Capto family had the highest critical velocities (Capto Adhere- 492, Capto Q- 477 cm/hr), whilst Sepharose CL-6B, Sepharose 4 Fast Flow and Sepharose CL-4B had the lowest critical velocity values (283, 204, 149 cm/hr respectively). Scanning electron microscopy (SEM) was used to image the structural properties of nine widely used agarose-based chromatography resins before use while pressure-flow analysis was used to characterise the mechanical properties of the same fresh resins. By understanding this, there is significant potential for facilitating timely and improved decisions in large-scale chromatographic operations, maximising resin lifetime whist maintaining acceptable column performance. This thesis, completed in collaboration with Eli Lilly & Co., aims to understand and assess the structural and mechanical changes that occur as agarose-based chromatography resins are exposed to different bioprocessing conditions in an attempt to explore the mechanisms by which different resins age. ![]()
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