pumps with magnetic levitation (e.g., Puralev®, Levitronix). By use of such a

centrifugal pump, Coronel et al. demonstrated similar production yields of IAV in

orbital shaken bioreactors for both ATF and TFF systems, respectively [44].

Nevertheless, and despite of the partial cleaning of membranes by the tangential

flow, membrane fowling is common in TFF systems at HCD and during virus

production due to the accumulation of cell debris caused by cell lysis. During the

virus propagation phase, almost complete retention of virus particles in the mem-

brane was commonly observed [70]. Therefore, in particular for virus production,

material and structural properties of the hollow fiber membranes selected are critical

for efficient production. A systematic overview on different membrane materials,

and structural and physicochemical properties with respect to filter fouling and virus

harvesting is given by Nikolay et al. (2020). With new membranes/materials

available, it will certainly be worthwhile to re-evaluate this method.

Although many problems including fouling, shear stress and product retention

are not completely solved yet, simple scale-up, good reproducibility, and suitability

for various cell lines and viruses make TFF an attractive option for cell retention in

virus production.

6.6.3

ALTERNATING TANGENTIAL FLOW

First introduced in 2000 by Shevitz, the ATF systems have been since then applied in a

variety of research and industry projects [71]. The external, pressure-based ATF

system consists of a hollow fiber unit, which is positioned between the bioreactor

and a diaphragm pump. A simplified overview and description of the ATF setup is

shown in Figure 6.6. The diaphragm pump pulls and pushes the cell suspension in the

hollow-fiber unit in an alternating way. This process can be divided into two phases:

exhaust cycle and pressure cycle [19]. Active filtration occurs during the pressure

cycle. A vacuum causes the convex diaphragm to be pulled downwards into the

chamber, thereby increasing the volume of the liquid chamber and drawing cell

suspension into the filter unit. In the filter unit, tangential filtration of the cell broth

occurs. The following exhaust cycle creates the alternating nature of the ATF system

by pushing the diaphragm with pressurized air back into the liquid chamber. This

backflush transfers the filtrate back into the bioreactor without the need for an addi-

tional peristaltic pump thereby reducing the shear stress for cells. Moreover, the entire

filter length is backflushed reducing the risk of blocking of the membrane pores [72].

A peristaltic pump removes constantly the separated permeate and fresh medium

can be added directly into the bioreactor, while the cells are retained in the hollow

fiber unit. For cross-flow filtration, cake-formation (increasing thickness with in-

creasing axial distance) and tubular liquid flow profiles can occur that lead to oscil-

lating fluctuations in pressure and sporadically declining fluxes [72]. The cross-flow

velocity minimizes the tendency of membrane fouling, but even in ATF systems high

cell concentrations and long growth cycles might ultimately lead to filter clogging.

As mentioned before, the ATF system is very well-established and commercially

available. Hollow fiber units can be obtained from several suppliers. In animal cell

culture, excellent separation performance was demonstrated in HCD cultivations up

to 2E08 cells/mL, and currently most perfusion-based processes for recombinant

Process intensification

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