Contributions of depth filter components to protein adsorption in bioprocessing

Abstract

Depth filtration is widely used in downstream bioprocessing to remove particulate contaminants via depth straining and is therefore applied to harvest clarification and other processing steps. However, depth filtration also removes proteins via adsorption, which can contribute variously to impurity clearance and to reduction in product yield. The adsorption may occur on the different components of the depth filter, that is, filter aid, binder, and cellulose filter. We measured adsorption of several model proteins and therapeutic proteins onto filter aids, cellulose, and commercial depth filters at pH 5-8 and ionic strengths <50 mM and correlated the adsorption data to bulk measured properties such as surface area, morphology, surface charge density, and composition. We also explored the role of each depth filter component in the adsorption of proteins with different net charges, using confocal microscopy. Our findings show that a complete depth filter's maximum adsorptive capacity for proteins can be estimated by its protein monolayer coverage values, which are of order mg/m , depending on the protein size. Furthermore, the extent of adsorption of different proteins appears to depend on the nature of the resin binder and its extent of coating over the depth filter surface, particularly in masking the cation-exchanger-like capacity of the siliceous filter aids. In addition to guiding improved depth filter selection, the findings can be leveraged in inspiring a more intentional selection of components and design of depth filter construction for particular impurity removal targets.

Moving from the bench towards a large scale, industrial platform process for adeno‐associated viral vector purification

“…Ultipor™ VF Grade DV50 (Pall Corp.) mentioned as preferred filters with preferred clearance of 5 LRV and at least 85% recovery of AAV (Hermens & Smith, 2013). In another patent, Genzyme references the use of various viral retentive filters for the removal of viruses from an AAV1 feed stream, such as Asahi KASEI and Pall virus filters, and the use of Millipore's Viresolve® NFR filter (Paulene, Gagnon, Nichols, & Thorne, 2010).…”

Effectiveness of host cell protein removal using depth filtration with a filter containing diatomaceous earth

“…Cell/debris removal occurs on both the external surface and throughout the depth of the filter due to sieving/straining by the porous matrix, while impurity removal occurs by a combination of electrostatic, hydrophobic, hydrogen bonding, or potentially mixed‐mode interactions. Khanal et al 14 showed that adsorption of small impurities (like lysozyme or β‐lactoglobulin) in depth filters containing either diatomaceous earth (Celite) or perlite is primarily driven by electrostatic interactions arising in large part from the positively charged polymeric binder, leading to greater removal of negatively charged proteins. In contrast, the removal of DNA was found to be a strong function of the DNA size 15 .…”

“…Typical commercial depth filters contain cellulose or polypropylene fibers, which provide the matrix structure of the filter; a high surface area filter aid like diatomaceous earth (DE), perlite, or activated carbon; and a binder, which insures the overall physical stability of the complex structure. 13 Cell/debris removal occurs on both the external surface and throughout the depth of the filter due to sieving/straining by the porous matrix, while impurity removal occurs by a combination of electrostatic, hydrophobic, hydrogen bonding, or potentially mixedmode interactions. Khanal et al 14 showed that adsorption of small impurities (like lysozyme or β-lactoglobulin) in depth filters containing either diatomaceous earth (Celite) or perlite is primarily driven by electrostatic interactions arising in large part from the positively charged polymeric binder, leading to greater removal of negatively charged proteins.…”

Lentiviral Vector Bioprocessing

“…Depth filters are a class of filters of a sponge-like texture, wherein particles are retained throughout the entire filter bed that is typically composed of polymer, binder and a filtration aid such as diatomaceous earth. For some depth filters, the media bears a charge for electrostatic retention of host cell proteins and DNA [184,185], with diatomaceous earth showing improved feed capacity although at high concentration it can lower LV titre [186]. In some small-scale studies, the use of multiple depth filter media, including cellulose-based and synthetic media (such as polyacrylic fibre with silica filter aids), has shown 90% turbidity reductions whilst maintaining high recovery of non-enveloped vectors, such as simian adenovirus based vectors [187].…”