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.

中文翻译：

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用于提高单克隆抗体治疗剂生产过程中深层过滤澄清效率的预处理

高细胞密度、高滴度的哺乳动物细胞培养是生产重组蛋白的新范例。虽然典型的动机是获得高产品滴度，但额外的不良结果通常包括细胞培养液中固体百分比的增加（细胞碎片和亚微米胶体），从而给下游加工带来新的挑战。本文重点介绍用于澄清细胞培养液的不同方法的探索和比较。这些方法包括离心后深度过滤、不离心的直接深度过滤和使用专门设计的密度梯度过滤的进料预处理，以提高澄清和从进料流中去除工艺污染物的效率。该工作还评估了三种不同预处理方法的影响，即 pH 调节酸性条件、金属阳离子（磷酸钙）絮凝和聚阳离子聚合物絮凝（使用聚合物 I 和聚合物 II）。所得结果表明，预处理的使用显着提高了深层过滤的澄清效率。基于聚阳离子聚合物-I 的絮凝等预处理选项导致过滤面积需求减少 > 5 倍，HDNA 减少 > 6 倍，同时保持可接受的 IgG 回收率 (>98%)。因此，预处理显着减少了深度过滤足迹（过滤面积要求减少了约 5-6 倍）。但是，必须考虑所需的工艺开发时间、资金成本、消耗品成本、