![]() ![]() In some embodiments, the AEX and CEX have no adjustments made to the pH or the conductivity of the in-process pool between the AEX and CEX, or CEX and AEX. Specifically, the approach uses cation exchange chromatography (CEX) and anion exchange chromatography (CEX) in tandem, either in AEX-CEX or CEX-AEX order. ![]() One aspect of the present disclosure is directed to purification of a protein of interest using an AEX-CEX or CEX-AEX purification approach wherein there is no in-line adjustment to any of the purification conditions, including pH or conductivity. The present disclosure is related to a combined chromatography process for the continuous purification of a protein of interest wherein an AEX and CEX are operated in flow-through mode, without any in-line adjustments to chromatography conditions including pH and/or conductivity. Nonetheless, the in-line adjustments of the buffer condition can be cumbersome and increase the operation cost and time and skid footprint. Automation and in-line adjustment of eluates have been reported to improve the purification steps. Necessary buffer adjustment (e.g., pH or conductivity adjustment) is often needed for each unit operation. Cation-exchange (CEX), anion-exchange (AEX), hydrophobic interaction (HIC) and mixed mode chromatography have all been used as polishing steps.Ĭurrent downstream purifications are often performed as batch processes with hold tanks between different unit operations (chromatography and filters). Downstream purification processes typically use Protein A chromatography for capture, followed by one or two polishing steps as described in Shukla et al., Downstream Processing Of Monoclonal Antibodies-Application Of Platform Approaches, J Chromatogr B Analyt Technol Biomed Life Sci, 848 (2007) 28-39. The clarified bulk is then subjected to downstream purification (DSP). The cell culture broth is then harvested through direct depth filtration or centrifugation. The manufacturing of mAbs begins with the protein expression in recombinant mammalian cultures. A specific subset of proteins, monoclonal antibodies (mAbs) and their derivative products (e.g., Fc-fusion proteins, bispecific antibodies), play an important role in treating some of the most challenging human diseases owing to the safety, efficacy and high quality of these types of biologics. Various upstream and downstream techniques can be found, for example, in Biopharmaceutical Processing: Development, Design, and Implementation of Manufacturing Processes, Jagschies et al., 2017. After production, “downstream” purification processes are employed to separate and isolate the protein of interest from any undesirable contaminants that may also be present in the mixture produced in the upstream process. Generally, the production and purification occurs first with various “upstream” cell culture processes to produce a protein of interest. Large-scale, efficient, and economic purification of proteins represents one of the major hurdles to overcome in the current biotechnology and pharmaceutical industries. ![]()
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