The objective of harvest and clarification unit operations is the removal of cells and cell debris to enable product capture
on a chromatographic column. The first step in the harvest of monoclonal antibodies from mammalian cell culture is cell removal,
followed by filtration unit operations for additional clarification. Centrifugation and microfiltration have been the primary
harvest techniques adopted industrially. Although depth filtration can be used as a primary harvest method, it is more common
to see this unit operation follow the primary harvest step to provide additional clarification. Flocculants are sometimes
added before harvest to improve harvest and clarification operations. Filtration through absolute pore size membranes is typically
the final clarification step before capture chromatography. Expanded bed chromatography has been developed as an integrated
unit operation that combines harvest with product capture, but to date, practical limitations have kept this technique from
being adopted in commercial-scale operations.
Several types of unit operations have been used for harvest and clarification of extracellularly expressed products from mammalian
cell culture. Key techniques include centrifugation, microfiltration, depth filtration, filtration through absolute pore size
membranes, the addition of flocculants to improve other harvest steps, and expanded bed chromatography. This article provides
a basic review of the fundamentals of these unit operations and practical considerations during process development. Further
details on these unit operations have been reviewed elsewhere.1 The final section describes how these unit operations fit together to form the harvest and clarification process for monoclonal
antibody (MAb) products.
Centrifugation uses the density difference between solids and the surrounding fluid. The centrifugal force accelerates the
settling that would normally occur during sedimentation. Most industrial applications use disk stack centrifuges to remove
cells and cell debris.2 Disk stack centrifuges offer continuous operation, making their throughput consistent with the desire to limit the time for
harvest operations. Figure 1 shows the schematic for a disk stack centrifuge.
Editors note: Figure 1 for this article is not available online. To obtain a copy of the figure by fax, please contact firstname.lastname@example.org
The basic principles of centrifugation involve a balance between the buoyant force acting on solid particles and Stokes's
law, which expresses the drag force. The ratio of flow rate to the effective settling area (termed Σ) is held constant during
scale-up of centrifugal operations. For a disk stack centrifuge, Σ is expressed as:
in which n is the number of disks, r
are the outer and inner radii of the disk, and θ is the angle of the disks from the vertical.
Operating conditions are often first screened at laboratory scale in a tubular bowl centrifuge and then translated to a disk
stack centrifuge. For the two centrifuges: