Experiments were conducted with a recombinant E. coli solution at a cell density of approximately 4% w/w for production of a plasmid used for gene therapy treatment. The primary
objective of the application was to concentrate the E. coli feed by a minimum concentration factor of 10-fold using a single-use technology. Initial trials were conducted with the small-scale
device and the filtrate flux rate for concentration was selected to be 22.5 LMH. It was estimated that six full-scale modules
(membrane area of 3 m2) will be required to process 500 L of feed at a feed flow rate of approximately 21 L/min (CFF of 7 L/min/m2). To demonstrate scalability to a series and parallel configuration, a trial was conducted to compare the performance of
a single full-scale module against the performance of three full-scale modules operated in series. The filtrate flux rate
was maintained at 22.5 LMH for each module throughout the trial. Pressure gauges were installed throughout the system to enable
accurate TMP determination across each module. The results shown in Figure 4 indicate that each of the three capsules manifolded
together in series provided a VT versus TMP curve very similar to a single module operated in isolation.
In summary, the Kleenpak TFF MF capsules' single-use TFF technology offers major benefits for cell harvesting and clarification.
The self-contained format eliminates the need to purchase and install holders or housings and also reduces exposure to biologically
active solutions. This is a critical consideration for manufacturers of vaccines and other products that present potential
workplace hazards. The optimized feed channel screen provides good performance at low flows and the technology is scalable
and reproducible.
HYDROPHOBIC INTERACTION MEMBRANE CHROMATOGRAPHY
Table 1
Sartobind Phenyl membrane adsorber from Sartorius Stedim Biotech (Goettingen, Germany) uses hydrophobic interaction chromatography
(HIC) principles and assembled into a 30-layer radial flow process capsule (Sartobind). It is based on hydrophilic regenerated
stabilized cellulose with the hydrophobic phenyl groups covalently attached to the cellulose matrix. The novel macroporous
membrane structure with 1–3 µm pores has been designed for high flow rates and high binding capacities. As seen in Table 1,
the binding capacity for a MAb on HIC membrane is found to be at par with conventional HIC resins even at significantly higher
flow rates. Two MAbs were loaded onto a membrane device and on a column at specified flow rates. The bed height for the column
was 30 mm and 8 mm for the membrane. The dynamic binding capacity as illustrated in Figure 5A was similar for the two formats.
Combining the advantages of membrane chromatography with virtually no diffusion limitation, shorter processing time because
of convective flow, and a ready-to-use disposable capsule format, the Sartobind HIC membrane adsorber represents a new membrane-based
tool applicable for flow-through as well as bind-and-elute applications. It provides a robust alternative capturing large
and unstable molecules that need to be processed rapidly.
Figure 5
At least two subsequent polishing steps are typically implemented in MAb downstream processes. The Sartobind anion exchange
membrane chromatography in combination with a hydrophobic interaction membrane chromatography offers different selectivities
and could reduce host cell proteins, high molecular weight aggregates, DNA, and leached protein A to acceptably low levels
that assure safety of the product. This may enable the downstream process to be just a one-column-step purification process.
Nathalie Frau, PhD, is a senior scientist in purification process development, biotechnology division, Sartorius Stedim North America.
Articles by Nathalie Frau, PhD
Anurag S. Rathore, PhD, is a consultant, Biotech CMC Issues, and a member of the faculty in the department of chemical engineering at the Indian Institute of Technology. Rathore is also a member of BioPharm International's Editorial Advisory Board.
Articles by Anurag S. Rathore, PhD
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