High cell density, high product titer mammalian cell culture is the new paradigm for production of recombinant proteins. While the typical motivation is to get a high product titer, additional undesirable outcomes often include an increase in percentage solids in the cell culture fluid (cellular debris and sub-micron colloids), thereby offering new challenges to downstream processing. This article focuses on scouting and comparison of different approaches used for clarification of cell culture fluid. The approaches include centrifugation followed by depth filtration, direct depth filtration without centrifugation and feed pretreatment with use of specially designed density gradient filtration to improve efficiency of clarification and removal of process contaminants from feed stream. The work also evaluates impact of three different pretreatment approaches, namely pH adjustment to acidic condition, metal cation (calcium phosphate) flocculation, and polycationic polymer flocculation (using polymer-I and polymer-II). The results obtained indicate that the use of pretreatment significantly improves the clarification efficiency of depth filtration. Pretreatment options like polycationic polymer-I based flocculation resulted in a >5 fold reduction in filter area requirement as well as >6 fold reduction in HCDNA while retaining acceptable recovery of the IgG (>98%). Thus, pretreatment offers a significant reduction in the depth filtration footprint (~5-6 fold decrease in filter area requirement). However, one must take into consideration the process development time required, capital cost, consumable cost, cost of the pretreatment chemical, cost of testing to demonstrate clearance of treatment agent, ease of scale-ability, and process robustness when finalizing the optimal clarification approach.
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