Up and downstream – hand-in-hand for integrated process characterization of a cherry-tagged enzyme
Upstream processes are hard to predict and therefore complex to design. High-throughput cultivation screenings in a micro-scale format are performed to create an appropriate design space. But the highest titer of cultivation doesn’t necessarily lead to superior process performance overall.
Considering both: Conjoint up and downstream development
Cultivation conditions are of fundamental importance to downstream processing. Critical impurities, which elute close to the product, greatly influence the process performance. Therefore, the complexity of the protein purification procedure strongly depends on the cultivation conditions. However, up and downstream processes are mostly developed separately. For an altogether superior performance in upstream and downstream, both processes should be developed conjointly.
Combination of high-throughput cultivation screening and chromatography modeling
A combined process characterization was investigated for a cherry-tagged enzyme. High-throughput experiments were performed to screen a large range of upstream conditions. From these, five random experiments were chosen for consideration in combination with the downstream process. The latter was simulated using ChromX, for which four different gradient elution experiments were performed to calibrate the model and to determine isotherm parameters. Afterwards, both processes were optimized with respect to overall yield and purity. The results of the optimization could be validated successfully at a later stage.
Superior process performance upstream and downstream
The combined characterization procedure has proven to be an effective tool for integrated process characterization. The final result can be depicted as a Pareto front that outlines the global set of optimal system points. The optimal operation point can then be chosen based on product quality demands.