Figure
6. Product quality results from each sampling point across the duration
of the run. Black dashed line indicated the acceptable cut-off for the
final AEX product. Each datapoint is from a different day in the
process. AEX final product sample points are labeled with day of sample
collection. (a) HCP titers in ng/mL (acceptable cut-off below line). (b)
Percent monomer (acceptable cut-off above line). (c) DNA titers in ng/mL
(acceptable cut-off below line). Circled DNA datapoint on AEX step shows
out of spec sample due to intentional overloading of AEX membrane.
Advanced Automation Control
Strategies
A key objective of this work was to evaluate the ease of integrating
multiple unit operations, which requires complex automation strategies.
Unit operation integration was handled completely by the PAK system,
which removed the burden of in-house automation development and
implementation. With the PAK system, unit operation integration is
mediated by system-monitored break vessels. For this run, this included
the (1) harvest break vessel, (2) protein A capture product vessel, and
(3) VI product flow kit vessel. The harvest break vessel and the virus
inactivation neutralized product break vessel were controlled to a level
setpoint using PID feedback control over the vessel outlet and inlet
pump, respectively. The protein A product vessel used minimum, low,
high, and maximum level criteria to trigger downstream pause/restart
events. Overall, all vessels were controlled as intended. The
significance of this control strategy is that it eliminates the concern
of over/under flow of intermediate break vessels.
The protein A elution vessel changeover performed on day 9, as described
in the Bioburden Control section, resulted in a buildup of cell culture
material in the harvest vessel, as the system was paused during this
time while perfusion material continued to accumulate. The 10L capacity
of the harvest vessel coupled with its 50L overflow bag provided ample
time for an operator to swap vessels and restart the system without
further incident. The protein A product vessel triggered the downstream
operations to begin once reaching a target level of 7.5 liters. After
draining to a level of 1.5 liters, the downstream unit operations were
triggered to pause to allow the vessel to refill. These trigger levels
were manually changed through the HMI throughout the run to allow for
the trigger events to coincide with an operator onsite. This was due to
an abundance of caution and is not required for the process to operate.
A critical, yet often overlooked parameter in a continuous process is
the mass flow rate of the overall process. Rather than set each unit
operation to a specified flow rate and over-size break vessels to absorb
differences, the PAK system actively controls the flow rate of each
individual step and the system as a whole. This is achieved by
controlling the last product pump in series to a flow rate set point,
while regulating flow of upstream product pumps to maintain intermediate
break vessel level (the protein A capture step is an exception). As a
result of the vessel level control, any change in flow rate of the last
product pump results in a corresponding change in flow rate of the
upstream product pumps. This control strategy allows for low break
vessel volumes (150 ml for this 50 L cell culture per day process).
The mass flow rate set point (set for the last product pump in series)
is determined by the most critical process parameter. For this run, the
most critical process parameter was determined to be the virus
inactivation residence time. Therefore, the mass flow rate of the entire
process was controlled to ensure the VI residence time setpoint was met.
A schematic of this process is displayed in Figure 7.