Internal virus polarization model for virus retention by the Ultipor^®VF Grade DV20 membrane

Abstract

Several recent studies have reported a decline in virus retention during virus challenge filtration experiments, although the mechanism(s) governing this phenomenon for different filters remains uncertain. Experiments were performed to evaluate the retention of PP7 and PR772 bacteriophage through Ultipor VF Grade DV20 virus filters during constant pressure filtration. While the larger PR772 phage was fully retained under all conditions, a 2-log decline in retention of the small PP7 phage was observed at high throughputs, even under conditions where there was no decline in filtrate flux. In addition, prefouling the membrane with an immunoglobulin G solution had no effect on phage retention. An internal polarization model was developed to describe the decline in phage retention arising from the accumulation of phage in the upper (reservoir) layer within the filter which increases the challenge to the lower (rejection) layer. Independent support for this internal polarization phenomenon was provided by confocal microscopy of fluorescently labeled phage within the membrane. The model was in good agreement with phage retention data over a wide range of phage titers, confirming that virus retention is throughput dependent and supporting current recommendations for virus retention validation studies. These results provide important insights into the factors governing virus retention by membrane filters and their dependence on the underlying structure of the virus filter membrane.

Probing effects of pressure release on virus capture during virus filtration using confocal microscopy

“…These results suggest that previously captured virus within the DV20 and Viresolve ® NFP membranes are able to diffuse out of the pores (capture sites) when the filtration pressure is removed, with the released phage either recaptured deeper within the filter or transmitted through the membrane when the filtration is continued. This behavior is qualitatively consistent with predictions of the internal polarization model proposed by Woods and Zydney () and Jackson et al (), although the more complex capture pattern seen with the Viresolve ® NFP would likely require a much more sophisticated mathematical description.…”

“…In both experiments, the phages were captured near the filter inlet, consistent with the slight asymmetry in the pore size of the DV20 membrane (Bakhshayeshi et al, ). This upper region was described by Jackson et al () as the “reservoir zone” within the DV20 membrane. The results obtained after the pressure release experiment were quite different, with the red phage (in the challenge before the pressure release) captured in two distinct bands separated by approximately 5 μm within the depth of the filter.…”

“…Confocal microscopy using a green carboxyfluorescein‐labeled phage showed two separate green bands near the entrance of the filter after the pressure release. This behavior was described using a modified version of the internal polarization model developed by Jackson et al (), with some of the previously captured phage migrating deeper into the filter after the pressure release. However, it was difficult to validate some of the critical assumptions in this model since it was impossible to distinguish between phage that were captured before and after the pressure release.…”

Continuous downstream processing for high value biological products: A Review

“…Significant viral clearance can also be achieved in chromatographic processes (Miesegaes et al, ), although most manufacturers also employ a virus filtration step to provide a robust, primarily size‐based, method for virus removal. Although virus filtration cannot be directly converted to continuous operation due to issues of membrane fouling and LRV decline (Jackson et al, ), it would be possible to operate two virus filters in parallel with the feed switched to the second filter when the capacity of the first filter was achieved.…”

Effects of solution conditions on virus retention by the Viresolve®NFP filter

“…Woods and Zydney and Dishari et al have shown that confocal microscopy can provide important insights into the key phenomena governing virus capture during virus filtration. This includes both the decline in virus retention seen in some virus filtration experiments as well as the transient jump in virus transmission seen in response to a disruption in the filtration process . In particular, Dishari et al developed a novel two‐dye approach to visualize the capture of viruses before and after a disruption in the filtration pressure.…”