Design of a filter train for precipitate removal in monoclonal antibody downstream processing

Abstract

Protein A chromatography has become widely established for the preparative purification of mAbs (monoclonal antibodies). Low pH elution from Protein A columns followed by neutralization can often lead to precipitation of impurities in the product stream, leading to a visually turbid solution. Pretreatment of the cell culture harvest stream with an increased surface area of the depth filter was found to reduce the magnitude of this problem through exploitation of the adsorptive properties of harvest depth filters. However, this was not a complete solution. Clarification of this turbid product stream prior to the polishing chromatographic steps in the downstream process posed significant filtration challenges. Development of a staged filtration process with the use of low plugging glass fibre depth filters as the first stage prior to membrane filtration through an absolute pore size membrane is described. Finally, a cost calculation is used to drive the selection of the final filter train for this application. The results presented here are expected to have wide applicability in mAb downstream processing as well as for other turbid solutions encountered in the downstream processing of other biomolecules.

Improved HCP Reduction Using a New, All‐Synthetic Depth Filtration Media Within an Antibody Purification Process

“…The harvest clarification step removes insoluble matter and protects the subsequent sterile filters and chromatography columns from plugging. Further downstream, depth filtration steps are also employed for secondary clarification and haze removal applications, especially after the Protein A capture and low pH viral inactivation (LpHVI) steps of a typical downstream purification process . This study focused on evaluating a novel depth filtration media used for secondary clarification within downstream purification processing of mAbs.…”

“…Further downstream, depth filtration steps are also employed for secondary clarification and haze removal applications, especially after the Protein A capture and low pH viral inactivation (LpHVI) steps of a typical downstream purification process. [4,5] This study focused on evaluating a novel depth filtration media used for secondary clarification within downstream purification processing of mAbs. The depth filters that are currently employed in commercial biopharmaceutical production processes are typically comprised of mixtures of cellulose pulps, diatomaceous earth (DE) based-filter aid, and a charged binder resin.…”

Enhancing Protein A performance in mAb processing: A method to reduce and rapidly evaluate host cell DNA levels during primary clarification

“…Protein A affinity chromatography commonly serves as the capture step in mAb production. The Fc region of a mAb exhibits a strong affinity and high selectivity toward Protein A, binding the target protein to the chromatography ligand or resin . During loading, the mAb is bound to the chromatography resin, whereas other impurities such as host cell proteins (HCPs) generally interact to a lesser extent with Protein A ligands and flow through the column to be discarded.…”

“…As a result, downstream unit operations are often scaled to accommodate a high level and range of impurities, reducing their effectiveness. For instance, additional unit operations, such as the inclusion of ancillary depth filters and chromatography columns, must often be incorporated into downstream unit operations to achieve the target throughput and purity …”

“…When HCPs and DNA are not completely removed during Protein A chromatography, the performance of downstream unit operations is also impacted. Failure to remove DNA can potentially lead to increased turbidity during neutralization after a low pH hold viral inactivation procedure, thus requiring the addition of a filtration step …”