Mammalian cells are most commonly used for the production of biopharmaceuticals. Over the past 25 years the focus has mainly been on protein-based therapeutic hormones, enzymes, antibodies, and vaccines.
Early mammalian cell culture systems have been increasingly replaced by high-productivity CHO and human-designed cell lines. The majority grow as suspended cells in serum-free, protein-free, or even chemically defined culture media and currently deliver up to 10 times higher product titers. The result has been a shrinkage in both batch sizes and bioreactor volumes during upstream processing.
As a result of a growing understanding of mammalian cellular processes at the molecular level, the number of potential product candidates for clinical research has been rising steadily.
Thus, the ability to utilize a range of different bioprocesses at varying scales has become necessary. In addition, the rising cost of healthcare has led to the need for shortened development and production times for biotherapeutics and to increased production capacities. Use of single-use systems, whose beginnings date back nearly 60 years to transfusion medicine, have made these advances possible.
In 1953 Fenwal’s invention of the plastic blood bag paved the way for single-use technology. For routine work in cell culture labs, plastic flasks, dishes, and 96-well plates (which have been commercially available since the 1960s) have increasingly replaced their counterparts made of glass. This changeover has contributed to a reduction in cleaning and sterilization times as well as to lower contamination rates.
Another important milestone in the history of single-use, the development of the first hollow-fiber reactor, was achieved by Knazek and his team in the early 1970s.
By carrying out perfusion culture operations with hollow-fiber membranes bundled in a single-use cartridge, mammalian cells can grow to high cell densities in in vivo like conditions. This ability has revolutionized the in vitro production of small amounts of hybridoma-derived antibodies.
In the mid ’70s, Nunc and Bioferon (now part of Rentschler Biotechnologie) started to manufacture cell factories from polystyrene. In the 1990s these multitray systems were shown to be capable of producing bioproducts (e.g., vaccines) with anchorage-dependent cells (while replacing roller bottles in GMP production).
The 1990s and 2000s saw a great increase in single-use technology for upstream production. In addition to the first single-use bags for storage and transport of buffers and media from HyClone (today a unit of Thermo Fisher Scientific), two-compartment membrane bioreactors and the first wave-mixed bag bioreactor system for mammalian cell cultures, the Wave Bioreactor 20, entered the market. The product was developed by Wave Biotech U.S, which was later acquired by GE Healthcare, and Wave Biotech Switzerland, which is now part of Sartorius Stedim Biotech.
Despite their lack of in-line sensors, single-use bioreactors prevailed in cell-expansion projects, screening experiments, and preclinical sample production at lab scale. Studies documented the advantages of wave-induced motion, which was initially viewed with skepticism, in surface-aerated bags manufactured from multilayer films. Positives included lower shear stress acting on cells, the opportunity for direct inoculation with cells pooled from T flasks (omitting intermediate cultivation steps), removal of need to add antifoam, and cost savings of 30–50%.
The Wave Bioreactor 20 and its successful application beyond cell expansion was a driver for the rapid further development of single-use technology in the 2000s.
In addition to aseptic connection devices, new sampling and transfer systems, single-use membrane filters and bioprocess containers, as well as stirred single-use mixers (e.g., LevTech’s Magnetic Mixer), have found their way into the development and commercial manufacture of small- and mid-volume protein therapeutics.
Since 2005 stirred single-use bioreactors have been available from HyClone and Xcellerex (XDR™ Disposable Stirred Tank Bioreactor) in addition to further wave-mixed systems (Tsunami Bioreactor and the AppliFlex from Applikon) for mammalian cells. In addition, all single-use bioreactors operating with bags have been available with optical single-use or standard sensors for in-process control of pH and DO.