ABSTRACT
Single-use bioreactors are commonly used for seeding stainless steel bioreactors or for producing material. The profitability
of this equipment has been well demonstrated for more than a decade, but few data on their scalability are published. In May
2010, Merck Serono Biodevelopment began a study to evaluate the performances of disposable bioreactors. Because different
technologies were available, this study compared performances of several types of mixing. The evaluation was performed both
for seeding application and for clinical material production. This study featured 20–50 L disposable bioreactors with gas
flow-rate scaled-down from seeding and production bioreactors. A fed-batch process producing a highly glycosylated molecule
was performed twice in four types of disposable bioreactors. The quality of the molecule together with molecule concentration
and cell growth were compared among the three single-use technologies. These process performances were also compared to 250
L and 1,250 L bioreactors and to a 3.6 L glass development bioreactor. These comparisons allowed Merck Serono Biodevelopment
to conclude on their uses and on the scalability (up and down) of these disposable systems. This study was completed by a
characterization of liquid–liquid and gas–liquid transfers inside each disposable bioreactor to estimate their potential in
terms of cell culture.
(Merck Serono Biodevelopment)
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To decrease sterilization, cleaning workload, and reduce costs, the disposables market has been diversified to meet the needs
of the biotechnology industry. At Merck Serono Biodevelopment (MSB), bags, tubing, and disposable cell culture containers
commonly are used in production processes. Hybrid processes with stainless steel bioreactors, however, remain because a lot
of factors have to be considered before implementing such technologies. Besides supplier support and quality, one critical
point is the scalability of these bags. Of course it is easier when you design a new plant, but when you have existing stainless
steel manufacturing bioreactors and you want to increase your capacity and produce material in disposable bioreactors, comparability
and scalability between bioreactors could be a problem. From May to September 2010, MSB performed a study to evaluate and
compare performances of different disposable bioreactors. Various equipment models are available on the market, so the options
were sorted based on maximum working volume, mixing technology, current good manufacturing practice compliance, availability,
data published, supplier experience, and internal requirements. To test different mixing principles, the following technologies
were evaluated: ATMI-Pierre Guerin single-use bioreactors; orbital shaking and stirred mixing, both from Sartorius. Processes
performed with this equipment began with a growth phase to evaluate bioreactor seeding performances, followed by a fed-batch
process producing a high glycosylated antibody-like molecule, to evaluate the impact of culture in disposable bioreactors
on molecule quality. Performances were compared to process development bioreactors and manufacturing bioreactors. The evaluation
finished with a characterization of transfers by KLa, mixing time measurements, and temperature mapping.
MATERIAL AND METHODS
Cell culture was performed twice in each disposable bioreactor under the same conditions.
Cell Culture
A vial of Chinese hamster ovary (CHO) cells was thawed in commercial medium in a static container. Cells were expanded every
two or three days in agitated containers until reaching 8.6.109 cells.
Cell Growth in Disposable Bioreactors
A sterility test was performed at 37 °C in each disposable bag two days before inoculation. The cells were then inoculated
in parallel in all disposable bioreactors at the minimum working volume provided by the supplier. Growth was then observed
for two days. Working volume was increased to the maximum working volume at the same cell density between bioreactors. Cell
growth at maximum working volume lasted three days.
Temperature was regulated at 37 °C. Stirring speed was fixed by the supplier.
Oxygen was regulated at 50% only with oxygen at a flow rate scaled-down from MSB 250 L seeding bioreactor for paddle and stirred
mixing bioreactors and at a flow rate fixed by the supplier and MSB according to its experiences with the orbital shaking
bioreactor.
