Use of scanning electron microscopy and energy dispersive X‐ray spectroscopy to identify key fouling species during alternating tangential filtration

Abstract

Alternating tangential flow filtration (ATF) has become one of the primary methods for cell retention and clarification in perfusion bioreactors. However, membrane fouling can cause product sieving losses that limit the performance of these systems. This study used scanning electron microscopy and energy dispersive X‐ray spectroscopy to identify the nature and location of foulants on 0.2 μm polyethersulfone hollow fiber membranes after use in industrial Chinese hamster ovary cell perfusion bioreactors for monoclonal antibody production. Membrane fouling was dominated by proteinaceous material, primarily host cell proteins along with some monoclonal antibody. Fouling occurred primarily on the lumen surface with much less protein trapped within the depth of the fiber. Protein deposition was also most pronounced near the inlet/exit of the hollow fibers, which are the regions with the greatest flux (and transmembrane pressure) during the cyclical operation of the ATF. These results provide important insights into the underlying phenomena governing the fouling behavior of ATF systems for continuous bioprocessing.

Contributions of Chinese hamster ovary cell derived extracellular vesicles and other cellular materials to hollow fiber filter fouling during perfusion manufacturing of monoclonal antibodies

“…Antifoam micelle was detected on fouled hollow fiber membrane surface by Kelly et al (Kelly et al, 2014), but we did not find silicon, a major element of antifoam, enriched in foulant patches. Silicon was also not observed by Sundar et al (Sundar et al, 2023). It's worth noting that Kelly et al (Kelly et al, 2014) imaged the frozen membrane without washing, but Sundar et al (Sundar et al, 2023) and we washed the membrane before imaging.…”

“…Silicon was also not observed by Sundar et al (Sundar et al, 2023). It's worth noting that Kelly et al (Kelly et al, 2014) imaged the frozen membrane without washing, but Sundar et al (Sundar et al, 2023) and we washed the membrane before imaging.…”

“…Filter fouling during perfusion production can lead to loss of product sieving and early run termination (Kim & Zydney, 2006;Zydney, 2021). Despite decades of studies, the culprit of fouling remains elusive (Kelly et al, 2014;Sundar et al, 2023;Wang et al, 2017;Wang et al, 2019;Xu et al, 2017).…”

Co‐current filtrate flow in TFF perfusion processes: Decoupling transmembrane pressure from crossflow to improve product sieving

“…However, this study does not allow us to distinguish between the beneficial contributions of backflushing at both the inlet and outlet, utilization of the entire membrane surface, and/or relaxation of the fouling deposit when the crossflow direction changes. The rTFF_2 run with higher amount of cell culture debris showed lower product sieving than rTFF_1 (Figure 5d), highlighting that rTFF is still prone to product retention which can likely be attributed to pronounced fouling at the inlet or exit of the hollow fiber modules at elevated debris levels (Sundar et al, 2023).…”

“…This causes an increase in the drag forces that push particles into the membrane (Ripperger & Altmann, 2002), which can lead to a denser deposit and greater particle penetration into the membrane, both detectable as an increase of irreversible fouling resistance (Weinberger & Kulozik, 2022). Sundar et al (2023) demonstrated that the greatest fouling in ATF systems occurs at the ends of the hollow fiber module, that is, where the filtrate flux is greatest. This might explain studies observing significant product sieving losses despite using properly sized ATF modules (Kim et al, 2016; Wang et al, 2017).…”