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Hybridomas, Genetic Engineering of 441

changes which prevent undue gradients developing along
the fibers. The hollow fiber system is suitable for both
anchorage-dependent and independent cells. Continuous
operation allows a high rate of product recovery from a
stationary high-density culture held in the extracapillary
space over a long period of time.

XXI. THE CONTROL OF CULTURE
PARAMETERS

FIGURE 16 Airlift fermenter. There are several culture parameters that are important
to control for maximum cell growth and antibody pro-
duction. These include agitation, temperature control, pH
E. The Hollow Fiber Bioreactor control, and oxygen supply.
This consists of bundles of synthetic, semipermeable hol-
low fibers which offer a matrix for cell growth similar A. Agitation
to the vascular system in vivo. Liquid can flow through
Animal cells tend to be fragile compared to fungal or bac-
the fibers (the intracapillary space) or through the space
terial cells. Cells in suspension can be damaged by vari-
between the fibers (the extracapillary space). In the nor-
ous forces acting in a stirred culture, the major damaging
mal operation culture medium is pumped through the in-
tracapillary space and a hydrostatic pressure permits the force is from bubble bursting on the culture surface re-
exchange of nutrients and waste products across the cap- sulting from culture aeration. The hydrodynamic shear
illary wall. The cells and large molecular weight products force resulting from the motion of a stirrer is thought to
be of lesser importance; nevertheless the stirring speeds
are held in the extracapillary space (Fig. 17).
commonly adopted for animal cell cultures are consider-
A major limitation of this type of system is that the
ably lower than those for bacterial cultures of equivalent
pressure difference that may establish along the length of
volume.
fibers can cause nutrient gradients and uneven cell growth.
The simplest stirring operation involves the rotation of
Such pressure differences and gradients become an in-
a suspended bar by a magnetic stirrer. This is the system
creasing problem with scale-up. The design of some hol-
used in glass spinner bottles and is suitable for stirring
low fiber systems is intended to correct this problem. Here
cultures up to a volume of 1 liter. At larger volumes, such
the pressure differential between the intra- and extracap-
magnetic stirrers are not suitable because of the increased
illary space is continuously monitored by sensors which
energy required for rotation. Top-drive mechanical motors
serve to control the opening and closing of valves which
are normally used for stirred tank reactors from bench-top
in turn affect the capillary pressure.
models to the larger commercial fermenters. In the de-
The exchange of molecules through the fiber wall oc-
sign shown in Fig. 15 the stirring shaft fits through the
curs in phases governed by a cyclic mode of pressure
stainless-steel head-plate and into the sterile culture. The
stirring motor and outer part of the drive shaft can nor-
mally be disconnected from the head-plate to allow the
fermenter to be autoclaved. In early fermenter models,
the stirring shaft was connected through the head-plate
by replaceable rubber/silicone seals, which were vulner-
able to damage and provided entry points of contamina-
tion. Later models have sealed units which are far more
reliable.
Typically, maximum stirring rates of 100–150 rpm are
used for cells in suspension. In order to ensure ade-
quate mixing at low stirring speeds, the culture vessels
are designed with a round bottom, which distinguishes
themfromtheflat-bottomedbacterialfermenters.Impeller
blades which are fitted at the end of mechanical drives
FIGURE 17 Hollow fiber bioreactor. shafts are designed to allow vertical as well as horizontal