Single-use systems for biotechnology products
Single-use systems have become an important tool in the development and scale up of biotechnology processes. Single-use systems save space, increase flexibility in scale and space planning and, to a large extent, eliminate cleaning costs in development and change over.
However, single-use systems come with their own challenges, including complete integration, scalability, and most of all with extractables and leachables. A comprehensive technical report has been published to guide potential new users through the process of implementing single-use technologies throughout their bioprocesses.1
Upstream processing has most fully embraced single-use technologies. From the thaw of the vial to the harvest of the bioreactor, it is possible to construct a fully disposable process stream for a cell culture process. However, limitations in heat and oxygen transfer have excluded bacterial and yeast processes. In this article, we review the available technologies and advances that have been made.
Vial thaw and cell expansion
The first steps of a cell culture biotechnology process are typically the thaw of a seed bank vial and the expansion of the cells in a series of shake flasks. In principle, these steps have always been performed in disposable containers, because the containers are small and cheap. However, there has been a change from glass shake flasks to plastic, since these are easier to transport. Since the materials are not as fragile, they can be gamma irradiated for sterilisation. This increases convenience, and increases sterility assurance, as gamma irradiation does not depend on the condensation of saturated steam in small material imperfections.
Although the materials may have changed in these steps, the footprint has not. Humidified CO2 incubator space and a sterile hood are still required. But it is possible to conduct the early stages of cell expansion without an autoclave, using all disposable, gamma-sterilised containers and implements. Eliminating the need for an autoclave also eliminates the requirement for high-quality water to produce clean steam. The utilities savings will be covered throughout this article.
Intermediate cell expansion
The intermediate steps of cell expansion are now dominated by disposable bags on rockers. This technology, pioneered by Wave, now owned by General Electric, relies on a tilting platform to mix the contents of the bag and transfer heat and oxygen. Expansion bags are largely differentiated by their materials of contact and a mixing pattern that is etched into their bases that affect the mixing pattern. These systems seem optimal up to a 20-litre total volume, 10-litre working volume, although they are available up to 1,000L total, 500L working. They have been proposed as production bioreactors, but they are not generally constructed to accommodate feeds, off-gas analysis and other features of high cell density high productivity bioreactors.
Intermediate cell expansion bags are generally easy to use and manipulate. They come pre-sterilised by gamma irradiation and do not require an autoclave. Sterile media is introduced directly through use of a sterile tubing welder, eliminating the need for an aseptic connection or a filtration step. Cells are also introduced through a tubing welder connection. Temperature and pH are monitored through integrated disposable probes, but dissolved oxygen is not typically measured.
There has been some concern with the polymer films used in cell expansion bags, as well as single-use bioreactors. In one often cited study,2 a chemical found at low levels in extraction studies was found to inhibit the growth of CHO cells when introduced at much higher concentrations. These findings have not translated consistently into manufacturing difficulties. However, research into biocompatible films continues.3