Table 2 summarizes the scalability of the proposed process for two other human monoclonal antibodies. The largest variability
was observed with the DNA measurements. All remaining parameters were very comparable. Final purified material retained the
quality obtained at bench-scale. Recovery values tended to be better with scale-up.
Table 2. Comparison of in-process contaminant clearance using the cation exchange (CEX) resin for capture and a hydrophobic
charge induction (HCI) resin (MEP HyperCel) for polish as the process is scaled-up. The overall yield for HuMAb-6 and HuMAb-7
for large-scale process was about 80% for both antibodies.
Regulatory agencies require at least two separate steps for viral inactivation and clearance during the production of therapeutic
proteins. These steps must be based on different modes of action, typically, a low pH hold and a viral filtration step.3 Viral inactivation was conveniently positioned between the chromatography steps, but this step can also be positioned at
the end of both sequences. Similarly, an additional specific step, such as viral clearance filtration, is performed after
the second column.
Table 3 shows the viral clearance of HuMAb-6 and HuMAb-7 following scheme A. Quantification of virus-like particles in the
cell culture supernatant yielded up to 10 logs of infectious and non-infectious viral particles. The minimum safety factor
for clearance of A-MuLV for the process ranged between 7 and 10 logs, for HuMAb-6 and HuMAb-7, respectively. Therefore, the
final antibody product (antibody and residual process contaminants) had sufficient viral clearance and safety viral factor,
allowing its use as a therapeutic.
Table 3. A-MuLV clearance capacity of the purification process using a cation exchange resin for capture and hydrophobic charge
induction resin for polish. Results were estimated from bench-scale runs.
With increasing titers in antibody manufacturing, purification processes must increase process efficiency and productivity.
By minimizing the number of purification process steps, the number of buffers to be prepared and process components are also
reduced. The two-column purification technology design described here can be used with various types of proteins with only
minor process adjustments. The scheme is a starting point for subsequent optimization and iteration steps because chromatography
parameters often can be optimized, for example, for higher binding capacity ranges. One additional advantage of using this
scheme is the flexibility of choosing the direction of the process steps that is most convenient for a specific product or
Gisela M. Ferreira, PhD, is a process engineer, and Jill Dembecki and Krina Patel are associate scientists. Alahari Arunakumari, PhD, is the director of process development, all at Medarex, Inc., 908.479.2451, firstname.lastname@example.org
1. Rathin D and Morrow, KJ Jr. Progress in antibody therapeutics. Am Biotechnol Lab. 2006; 24(8):8–10.
2. Thompson R. Antibody therapeutics: product development, market trends, and strategic issues. 2006 Oct. Available from:
3. Zoon KC. Points to consider in the manufacturing and testing of monoclonal antibody products for human use. US FDA, CBER.
Rockville, MD; 1997 Feb 28.