In the attempt to mitigate the ‘Downstream Bottleneck’, we have identified that target protein precipitation is an appealing alternative to protein A chromatography because it takes advantage of the enhanced supersaturation brought by increased titer. Unlike affinity chromatography processes whose performance suffers at high titer and requires either additional ligand or extended process operation time, target protein precipitation induced by the addition of reagents requires less reagents at higher titer and has many reactor configurations that can be accommodated by current manufacturing facilities. Among possible precipitants with different mechanisms, we have identified the combination of polyethylene glycol (PEG) and zinc chloride (ZnCl<sub>2</sub>) as a potentially generic precipitant system which has a synergistic rather than additive effect in reducing protein solubility. In this work, we investigated the solubility behavior of model proteins and monoclonal antibodies to obtain solubility maps to guide the design of a continuous precipitation-filtration system. A stopped flow device was used to spectroscopically monitor protein precipitation kinetics at different conditions to provide insight for the design of a tubular precipitation reactor. Furthermore, precipitate particles generated in batch and continuous formats were examined via light scattering technique to establish a correlation between precipitate particle morphology and subsequent microfiltration performance for precipitate dewatering and washing.  We processed harvested cell culture fluids containing a monoclonal antibody with this system and were able to obtain high antibody yields in the precipitate phases while the large majority of the host cell proteins remained soluble and could be removed by washing – this performance is predicated on favorable precipitate morphologies.  Based on the understanding of the protein solubility and precipitate morphology, integrated with the understanding of microfiltration of protein precipitates, a tubular reactor can be constructed and optimized to fulfill purity and yield requirements for the capture of recombinant proteins in biopharmaceutical manufacturing. <br/>

Process development of a precipitation-filtration unit for recombinant protein capture step. I: Characterizing protein solubility and precipitate morphology

2019 Spring National Meeting

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