Guest Column | January 11, 2018

Renovating An Existing Biotech Plant To Accommodate A Higher-Titer Product

By Erich H. Bozenhardt and Herman F. Bozenhardt

Over the last few years, we have discussed various design approaches and compliance trends. In this article, we will tackle a design problem that typically emerges in a mature production organization. Most biotechnology organizations have designed their monoclonal antibody (mAb) production operations to handle 1 to 4 g/L titers. The typical design features a cluster of large-scale (≥10,000 L) batch bioreactors or multiple trains of 1,000-liter reactors. The bioreactor turnover is generally such that a harvest happens every few days, and, when sequenced properly, a single downstream purification train can handle the output of the several bioreactors. In the world of perfusion, the downstream columns are small and dedicated to the perfusion “train.”

Through a better understanding of media needs and improved clonal selection, processes are reaching the next level of titers in the 5 to 15 g/L range. As a result of these increased titers and the strategies employed to produce higher titers, the next-generation biotech facility is needed. While some product portfolios might be a good fit for 2,000 L single-use scale, high titers for high-demand products can reap a significant reduction in cost of goods using large-scale trains and can minimize capital by using existing assets. Existing facilities will need to adapt.

This is where we are today, with a wide spectrum of plants moving into their next generation of production of mAbs and other therapeutic expressions. This new higher titer changes the way we look at both the process and the facility. On the process side, we will need less bioreactor volume, but does that translate into smaller vessels (e.g., using seed vessels as the production bioreactor) or more product changeovers? That depends on the product demand, operational capabilities, and economics of the facility. The downstream concentration may require an entire re-think in terms of the column size, capacity, spatial velocity, and turnover to accommodate the required capture.

Design Considerations

The ideal approach is to execute a complete capacity analysis using the existing process history and pilot plant data against a specific process/platform. Even when this is done correctly, there will be challenges to the design case, as the capital project planning will typically be in parallel with the final stages of process development. Here are general guidelines and considerations for proactive development and capital planning:


Higher titers are resulting due to a combination of improvements in the cell line expression and overall increase in cell density. Upstream changes are needed to enable high-titer cultures, improving nutrient and gas feeds.

  • Increased gas demands may be higher than the currently installed mass flow controller (MFC) range. For traditional stainless steel (SS) systems, consider installing higher-range MFCs in parallel or multiple lower-ranged units to maintain flow accuracy. This can also be accommodated for the single-use bag/MFC combo depending on the size of the bioreactor.
  • High fed-batch requirements may require additional media hold capacity. This, in fact, may force the size of the upstream part of the plant to increase.
  • Process analytical technology (PAT), or a closed loop control: In general, commercial manufacturing is still in the data collection phase, but next-generation processes are being developed with feedback control focused on fine-tuning nutrient additions and gas concentration control. The use of online mass spec or atomic absorption to understand the respiratory quotient and correlate it with time and cell growth becomes more critical in this application. The precision of the control and the timing of the batch termination could be the difference between achieving the optimal yield or a serious cell line degradation.
  • An increased cell mass could require 2,000 L scale processes that previously used depth filtration for harvest to switch to using centrifugation. These larger-scale processes could see a slowdown due to the centrifugation process. Centrifugation processes typically require longer processing time than filtration and thus might warrant the bioreactor to have a dual temperature cycle with a cool-down period due to long hold time. Due to this change in processing, companies need to examine the complexity of the centrifuge, its cleaning (and validation), and its cost and increased footprint.


As we discussed earlier, the results of the upstream improvements have shifted the bottlenecks in many existing plants to downstream operations by feeding them a higher concentration stream and requiring them to process and retain the material. This alone has increased the risk for a significant loss or process interruption. Downstream units with connections, tubing systems, filtration, and skids have a greater opportunity for contamination, leaks, and material loss.

  • High-capacity chromatography resin should be considered as part of the process development (>50 g/L Protein A, 100 g/L for anion/cation).
  • Membrane chromatography is a possible alternative or co-processing system. Membranes offer high binding capacity and can handle higher flow rates than traditional resins.
  • Even with high-capacity resin, buffer volumes can be too large for existing buffer trains. In-line dilution and in-line conditioning may be necessary. In-line dilution systems meter buffer concentrate and water to the target concentration. In-line conditioning meters concentrates the individual buffer components with water, and then pH adjusts the stream. These systems can work with existing tankage, assuming the existing tankage material can tolerate high chlorides (e.g., AL6XN, 904SS, Hastelloy).
  • For new systems, it is important to keep in mind the higher alloys will cost 1.6 to 4 times the typical cost of 316 L SS. Single-use systems (SUSs)/disposables lower the initial capital and are easier to install in an operating facility. However, the tubing systems and sheer volume must be considered as well as how to consistently and reliably build them for each batch, avoiding connector problems, losses, and spills.
  • Larger lots increase the risk for catastrophic loss, resulting in more testing and other supporting costs. On the other hand, lot splitting reduces the testing and documentation efficiencies gained by yielding more product from one bioreactor.
  • Larger intermediate volumes in the realm of three to four times the traditional volumes require much larger SUS bags and present handling/logistical issues. The bags require larger carriers (e.g., pallet tanks, possibly with jackets) and need more physical space and access to the next process step.
  • Larger intermediate volumes for large-scale stainless steel systems may not fit in existing tankage. For facilities that utilized gray space for tankage, it might be practical to swap/add tankage.
  • Single-pass tangential flow filtration (SPTFF) can be used to reduce volumes from chromatography elution in-line. This can be a great benefit if used properly to reduce the liquid volume. SPTFF systems may not have been used in the past and require careful planning, training, and execution, as the SPTFF operation has to sync with the elution steps on the chromatography skid to consistently meet a target concentration.
  • Depending on target molecule stability, it is possible to use a standard tangential flow filtration (TFF) unit to concentrate simultaneously with a chromatography elution in a fed-batch mode. The surge capacity of the TFF tank and permeate flow need to be considered against maintaining the appropriate velocity through the column during elution.
  • Smaller-scale TFF systems that currently use a peristaltic pump might need to consider higher flows. The answer may be found in multi-chamber diaphragm pump systems such as the Quattroflow pumps, which are still positive displacement pumps but have significantly improved flow and pressure output.
  • In the new higher titer process, the downstream bulk fill volume will go up, straining manual processes, logistics, bag carriers, and volumes.
  • Higher bulk concentrations are possible for newly developed processes and appear to increase efficiency. However, processes going into commercial operations in the near term are likely to have concentrations similar to current lower-titer processes and ultimately require a dilution step in the bulking process, which may not have been contemplated in the past. The alternative is to ship a higher concentration level to the fill site, requiring them to buy larger vessels and take longer to prep the final fill.

Supporting Systems

The utility systems, specifically the liquid supply and drainage, need to be expanded to accommodate the upstream and downstream.

  • Increased buffer needs will drive up water for injection (WFI) generation and usage. This usage will come at specific and defined times in the processing and may require a larger WFI tank for surge capacity. It may not require an expansion in the WFI system; however, a capacity analysis should be completed.
  • The liquid waste volumes will increase and require a capacity analysis and potential expansion of the process waste and decontamination system.
  • Finally, the larger bulk fill volumes will increase the need for refrigerator/freezer space and access capability (larger doors, wider corridors, higher capacity elevators, and potentially automated guided vehicles [AGVs]).

The Plant Site

The hope is the increased titer plant is more efficient and lower in cost. In terms of the facility, the possible need for additional media hold volume has the biggest space impact from upstream. From the above discussion, the downstream operational footprint may need to increase in size and complexity. The introduction of larger volumes and possibility new systems (e.g., single-pass tangential flow filtration [SPTFF], in-line dilution/in-line conditioning [ILD/ILC]) will require more logistical support and more operators.

The utility, cold storage, and other support services required will also increase the footprint and require some site reconfiguration and expansions, especially if the WFI holding tanks/generation need to be replaced with larger units.


Overall, the conversion from a low- to a higher-titer biotech production is feasible and will increase efficiency and production capacity. However, the operator and designer must consider the following:

  • The need to introduce newer technologies to accommodate the higher titers (high-capacity resins, membrane chromatography, SPTFF, online analyzers, etc.)
  • The overall increase in the size/footprint of the facility at about $700 per square foot
  • Increase in operation staff and retraining the existing staff
  • Increased operational and financial risk, as greater product value is handled in smaller batch sizes, thus increasing the chances of catastrophic loss from a failure or mistake

About The Authors:

Herman Bozenhardt has 41 years of experience in pharmaceutical, biotechnology, and medical device manufacturing, engineering, and compliance. He is a recognized expert in aseptic filling facilities and systems and has extensive experience in the manufacture of therapeutic biologicals and vaccines. His current consulting work focuses on aseptic systems, biological manufacturing, and automation/computer systems. He has a B.S. in chemical engineering and an M.S. in system engineering, both from the Polytechnic Institute of Brooklyn.

Erich Bozenhardt, PE, is the process manager for IPS-Integrated Project Services’ process group in Raleigh, NC. He has 11 years of experience in the biotechnology and aseptic processing business and has led several biological manufacturing projects, including cell therapies, mammalian cell culture, and novel delivery systems. He has a B.S. in chemical engineering and an MBA, both from the University of Delaware.