Navigating in the single-use space – How to find the right cell culture bioreactor

A Guest Blog by Eva Lindskog, PhD Upstream Marketing Leader, GE Healthcare Life Sciences

The bioreactor is the cornerstone on which cell culture for biopharmaceutical production is based. The bioreactor should provide an efficient means to expand the cell culture and to repeatedly deliver a product with the desired quality attributes. Choosing the right bioreactor is vital, but how do you know which bioreactor is the most suitable for your applications?

From the early-day use of glass flasks in research and stainless steel vessels in manufacturing, the bioreactor has come a long way. A fundamental change in bioreactor technology is the shift to disposables. Although the use of stainless steel vessels still prevail at the very large manufacturing scales, single-use options are increasingly common in culture scales up to 2000 L. The benefits with disposables are now widely acknowledged for both upstream and downstream applications. Single-use equipment offers increased flexibility with quicker set-up and changeover between runs, and reduces the need for costly and time-consuming cleaning and cleaning validation operations.

Single-use bioreactor technologies can be classified based on the system design. The most common systems are based on stirred-tank and rocking bioreactor technologies, although alternatives do exist, such as bioreactors based on air-wheel mixing, bubble mixing, and orbital shaking. Stirred-tank bioreactors for single-use are based on the same principles as stainless steel vessels, with cylindrical support tanks including impellers for mixing and spargers for culture gassing. The culture contact material can be rigid or flexible plastic. The rocking motion technology takes a different approach, with a flexible, plastic, pillow-like bioreactor chamber. The chamber is placed on a tray with a back and forth movement that generates a continual mixing action and gas transfer within the culture fluid.

The first consideration when choosing a single-use bioreactor is to determine what capacity you require for your application, and hence what bioreactor size you will need. The smallest single-use bioreactors have capacities at the mL-scale and the largest options support culturing up to 2000 L. Further navigation within the single-use bioreactor space will depend on a number of factors, for example, where the bioreactor will be used in the biopharmaceutical process.

Research and preclinical cultivation

Key bioreactor requirements for research and preclinical development applications are ease of use and flexibility, small volume capacity, and accurate control of culture conditions. An easy-to-use and flexible system facilitates set-up and enables exploration of different process alternatives, from batch to fed-batch and perfusion culturing, for a variety of cell types including those of mammalian, insect, and plant origin. A small volume capacity minimizes the space required for the bioreactor and the medium volume required for the experiment can be kept low, a benefit of special importance when the culture medium or the additives needed are expensive. Accuracy in measurement and control of culture parameters, such as temperature, pH, and DO, is critical to achieve reliable and repeatable results that support conclusions for further process development.

For small volumes where limited culture surveillance is required, a simple shaker flask is an option. However, culture expansion will require combining several small shaker flasks into a larger vessel, a transfer process associated with a risk of contamination. When increased culture monitoring and improved control is required, the easy-to use rocking bioreactor is a good option.

Process development

When it comes to process development, the choice of bioreactor will depend on the purpose of the studies to be undertaken. Upstream process development activities include formulation of culture media and feeds as well as determination of bioreactor settings suitable for the cell clone to be studied. The end goal is to obtain a robust, high-yielding process suitable for the manufacturing scale.

When the purpose is to optimize process parameters for subsequent scale-up for manufacturing, it is of crucial importance to use a well-characterized bioreactor system that provides an accurate representation of the likely large-scale outcome. Unexpected scale-up effects are well-known but unwelcome. A poorly characterized process can be very costly if a problem suddenly appears at a later stage. There are many reasons for such large-scale failures and some of them relate to the basic dynamics of the bioreactor systems, for example, in terms of mixing and gas transfer. A common challenge is to keep the partial CO2 pressure sufficiently low in the reactor to prevent adverse effects on product quality. Some products are sensitive to high CO2 pressure as such, whereas indirect adverse osmolality effects might originate from the addition of base to maintain a constant pH.

A good general understanding of the culture parameters, in combination with a small, single-use bioreactor capable of mimicking the situation in the manufacturing vessel, is an excellent starting point to avoid unwanted surprises at a later scale. The stirred-tank bioreactor, in a multi-vessel set-up to facilitate design of experiments, is a commonly used reactor type for process development purposes. Glass bioreactors are still widely used, although, single-use options are increasingly popular and the associated benefits are recognized by many.

In the case of optimization of feed regimen as well as media and feed formulations, the speed and simplicity of a rocking-type of bioreactor might advocate for its use in screening experiments. At small-scale, deep-well plates and shaker flasks are good options, as are small-scale, high-throughput, single-use bioreactors. The obtained results can be confirmed later in a stirred-tank vessel.


When it comes to the final manufacturing stage, the bioreactor has to deliver according to what was observed at small scale. In addition, the bioreactor has to support manufacturing of regulated products under the Quality Management System in place and provide the automation architecture necessary to ensure process control. For production volumes of more than 2000 L, a traditional stainless steel bioreactor is still the most common option. In certain cases, stainless steel vessels might also be economically viable in smaller scale, for example, when setting up a single-product facility operated with a high-batch frequency. However, a single-use bioreactor system can be a more suitable choice in other cases where increased flexibility, reduced set-up costs, and minimized environmental impact are required.

The technology of the single-use production bioreactor should allow uniform scalability from the process development bioreactor, without compromising on basic parameters such as mixing and gas transfer. In addition, the bioreactor turn-down ratio needs to be considered. Many cultures have split ratios of approximately 1:5. If the single-use bioreactor can be operated at 20% or less of the final volume, the vessel will be able to support two seed-stage cultures with just one single intermediate in-bag medium addition step. A lower turn-down ratio will require one bioreactor vessel per stage of the seed-train, and might also compromise with certain process strategies including batch-refeed and high-volume feeding.

The final, complete bioreactor set-up depends on the manufacturer, the specific product to be produced, and the design of the manufacturing facility where restrictions in terms of ceiling height and floor space might need to be considered. A common strategy is to use a rocking bioreactor at the seed stage, followed by a stirred-tank bioreactor for the production stage.

Certain products are manufactured in small volumes. Examples of small-volume processes include cell expansion for therapy applications and virus expansion for the production of live, attenuated vaccine. Both of the above mentioned examples would benefit from a single-use bioreactor strategy in terms of minimizing the risk of contamination and carryover from previous batches. Each specific business case will determine the amount of product needed and hence the smallest bioreactor size required for the requested production volume. The main requirements are again compliance with regulatory demands, reliability, and accuracy. A rocking bioreactor that meets these needs is a good option in many cases. However, certain applications might benefit from the stirred-tank bioreactor format.


Before a decision is made on a single-use bioreactor investment, important aspects should be considered, not only in the immediate term but also in the mid and long term. A purpose-developed, robust, single-use bioreactor with a proven performance and a design based on a solid technology is a good starting point. It is also recommended to take a close look at the bioreactor manufacturer to evaluate security of supply, quality aspects, and customer support capabilities. Choosing a bioreactor that is well suited for its purpose is vital. Time spent upfront for investigating the options available will lead to a good return of investment in the end.

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