The increased cell density and product titer in current bioreactors have led to the increasing use of depth filters for clarification of cell culture fluid (CCF), either as a stand-alone unit operation or after centrifugation in the production of monoclonal antibody (mAb) products. Several recent studies have shown that depth filters can also reduce the concentration of smaller impurities like host cell proteins and DNA, decreasing the burden on subsequent chromatographic operations. The objective of this study was to evaluate the adsorptive properties of the Pall PDH4 depth filter, which contains diatomaceous earth, cellulose fibers, and a binder. Experiments were performed with both DNA and a series of model proteins with defined pI, molecular weight, and hydrophobicity chosen to match the range of host cell proteins in typical CCF. Breakthrough curves were determined by tracking the permeate concentrations in constant flux experiments using individual proteins / DNA and multi-component mixtures. The exact location of the adsorbed proteins and DNA within the depth filters were determined using confocal scanning laser microscopy (with fluorescently-labeled species) and Electron Dispersive Spectroscopy (SEM-EDS). Protein binding was greater for proteins that were positively-charged and had higher hydrophobicity, with the latter determined from the GRAVY value calculated from the known amino acid sequence. For example, the binding capacity of chymotrypsin (GRAVY = 0.064) was eight times larger than that for Ribonuclease A (GRAVY = -0.663), even though the proteins have similar pI. These filters also showed very high levels of DNA removal, despite the strong negative charge on the DNA. These results provide new insights into the factors governing the filter capacity and performance characteristics of the PDH4 depth filter that is widely used in the clarification of CCF.

Investigation of the adsorptive properties of depth filters in bioprocessing

2019 Spring National Meeting

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