This article discusses the evaluation of a novel single-use fluidized bed centrifuge (FBC) for harvesting of antibodies. An
FBC that contains four single-use 100-mL chambers was used to harvest Chinese Hamster Ovary (CHO) cell cultures. Optimal operating
parameters were defined by performing preliminary studies to determine the maximum chamber capacity and the feed flow rates
into the centrifuge. Results of the preliminary studies showed the maximum capacity was approximately 10x10
cells/chamber, and the initial and process feed flow rates used for the studies were 80 and 140 mL/min/chamber, respectively.
Five simulated cell-harvesting runs followed the preliminary studies. Three of the five simulated runs utilized healthy cell
cultures with viabilities > 90%, and the remaining two runs were with cell cultures with viabilities < 50%. Results showed
that the FBC was efficient in separating cells from the product, with low cell density and turbidity detected in the centrate.
Average clarification efficiencies were between 88–93% without the use of depth filtration post-clarification. There was no
increase in lactate dehydrogenase (LDH) and residual DNA levels indicating that minimal amount of shear stress was induced
by the centrifugation. The results, therefore, suggest that the FBC is a promising alternative for cell-harvesting applications.
The first step in the recovery of a secreted product from a mammalian cell culture is to separate the cells and cell debris
from the product in the supernatant of the cell culture. A disc-stack centrifuge or tangential flow filtration and microfiltration
(TFF–MF) is, generally, used to separate cells as a primary recovery step, followed by depth filtration as a secondary step
to remove remaining cell debris and other impurities before the product in the clarified liquid (centrate) can be loaded into
a chromatography column (1–3). More information on these primary technologies can be found in a review paper (4).
Image courtesy of kSep Systems
Most conventional cell harvesting technologies are not single-use systems and, therefore, require extensive cleaning and sterilization
between batches during production. Single-use technologies have been widely used in the biotechnology industry due to their
distinct advantages. Within the past decade, a variety of cell-culture processes has adapted single-use systems, ranging from
the use of WAVE technology for processes up to the 500 L scale as well as single-use stirred-tank bioreactor (SUBs) technology
up to the 2000 L scale as the production bioreactor vessel for therapeutic protein production (5, 6). Several single-use cell
harvesting technologies have also been utilized, including but not limited to the Pod Filter system from Millipore, TFF bioprocess
systems from SciLog, and Sartoclear depth filter systems from Sartorius Stedim.
In the past two years, single-use centrifuge technologies have also emerged as a potential alternative for primary recovery
processes. The Unifuge, made by Pneumatic Scale Angelus, is a fully automated centrifuge system that uses a single-use insert
inside the centrifuge bowl. The kSep technology, a FBC made by kSep Systems Inc., uses balancing centrifugal and fluid flow
forces to capture cells in up to four single-use chambers. Both technologies involve retaining cells inside the centrifuge
while the centrate that contains the product is continuously separated and discharged. Because of the FBC's advantages (e.g.,
washing capability, low shear, and continuous mode of operation), the authors selected the FBC for the evaluation of mammalian
cell harvesting. The FBC and its single-use set are shown in Figure 1.
Figure 1: FBC (left) and single-use set (right) loaded into the centrifuge rotor (Image courtesy of kSep Systems). (FIGURE
1 COURTESY OF kSEP SYSTEMS.)