Interaction of cell culture with downstream purification: a case study

Abstract

Separation of product from secreting mammalian cells in the culture both means the transition from product generation to product isolation. This interface within a biotech production process has to perform a proper solid/liquid phase separation of the cell suspension to make the product containing fluid amenable for further purification. These subsequent steps require fluid with low occurrence of contaminants in order to function properly. The goal of this study was to evaluate some economic and fast cell separation methods for the preparation of a product fluid ready for use in further ultrafiltration and chromatographic processes. We have performed experiments to test the usefulness of disc stack centrifuges and tangential flow microfiltration units at large scale. Both systems revealed outstanding prospects with regard to throughput and scale up properties. However, the centrificgation did not lead to a fluid sufficiently free of particles for direct ultrafiltration or chromatography. Thus, an additional filtration step was necessary. On the other hand microfiltration led to an acceptable quality of process fluid directly. By optimisation of process parameters an effective, reproducible and robust cell separation can be obtained. However, our experience has been that such optimal conditions are somewhat specific for a narrow range. Thus, even the equipment functioning well with one type of cell would possibly not perform as well with another cell or even with the same cell under conditions slightly different to the usual situation.

Effect of pore size, shear rate, and harvest time during the constant permeate flux microfiltration of CHO cell culture supernatant

“…During constant permeate flux operation, the degree of TMP increase provides an indication of fouling. Berthold and Kempken17 compared filtrations of hybridoma cell suspension at different constant permeate flux. A rapid and sharp drop of permeate pressure (and therefore TMP increase) was observed at the higher permeate flux, whereas a constant permeate pressure and TMP were observed for the filtration operated at the lower permeate flux.…”

DNA Depletion by Precipitation in the Purification of Cell Culture‐Derived Influenza Vaccines

“…But apart from economical considerations that may speak against this, membrane microfiltration could still be required for the protection of chromatography columns [47,48]. For this reason, a membrane microfiltration step was incorporated immediately after the selective precipitation of DNA.…”

“…Generally, continuous centrifugation outperforms filtration as it does not show problems related to fouling or aging of membranes [48]. Therefore, it allows for the fast clarification of large-scale cell cultures, which would be particularly beneficial in case of an influenza outbreak when hundred thousands of liters of purified virus harvest have to be generated for vaccine production.…”

Comparison of different options for harvest of a therapeutic protein product from high cell density yeast fermentation broth

“…Continuous centrifugation has been widely used for harvest of large-scale cell culture (Berthold and Kempken, 1994;Winter, 2004) and microbial fermentation (Clarkson et al, 1996;Varga et al, 2001) processes in the biopharmaceutical industry. For extracelluar protein product, secondary clarification using depth filter after centrifugation is normally required prior to further downstream processing.…”