Initially, serum-free medium was introduced into biopharmaceutical manufacturing because of the cost and process constraints
of serum. Serum-free medium (SFM) was first used in mammalian cell culture by Ham in 1965. Much of this early work involved
anchorage-dependent cell lines.7–9 To achieve growth-promoting effects similar to those of serum, SFM often was supplemented with animal-derived components
that replaced the role of serum, such as transferrin, albumin, insulin, and other biological extracts. Serum supplementation
continues to be used in many early stage research applications, clonal development, and vaccine production.
Establishing more defined media regimes reduced the problems associated with the batch-to-batch variability seen with serum
and allowed more consistent product and process control with simpler purification and downstream processing strategies. However,
having more defined media often resulted in extended cell adaption times, reduced growth rates, and decreased product titers,
all of which increased the costs of manufacture.
The most common serum-free supplements incorporated into a basal medium have been growth factor sources, transferrin, hydrolysates,
and albumin. Even today, some of these supplements are animal-derived (transferrin, albumin), contain animal components, or
are ill defined and are still used in manufacturing processes because of the lack of safe alternatives. The regulatory issues
of possible contamination of the final product with adventitious agents and ill-defined processes therefore remain. Chemically
defined SFM are commercially available, but for some cell lines it has not been possible to design a robust, animal-free,
and chemically defined medium that will perform as well as serum. Often, different cell lines and clones exhibit a large degree
of variability in their nutritional requirements to achieve optimal growth and performance, resulting in a lengthy and costly
media development program.
The remainder of this article will discuss four media supplements commonly used in the biopharmaceutical industry and recent
advances in their development.
Regulatory agencies have encouraged the development of serum-free media devoid of any animal-derived components to avoid the
risk of introducing adventitious agents such as viruses and prions. Protein hydrolysates have been shown to have beneficial
effects on cell growth and productivity and are a relatively effective alternative to the use of serum. Peptones derived from
bovine milk or animal tissues, such as Primatone RL (MP Biomedicals, Irvine, CA), are capable of supporting a number of different
cell lines in the absence of serum.10 However, non-animal hydroly-sates from microorganisms such as yeast11 and plants including soy and rapeseed12,13 are being investigated as supplements for supplying the nutritional requirements of mammalian cells in culture.
Although plant-based hydrolysates have been shown to promote growth and productivity,14 the industry has not fully embraced hydrolysates as a serum substitute. Primarily this has been because of a lack of chemical
definition and lot-to-lot variability leading to process and product inconsistencies. Currently, manufacturers of hydrolysate
supplements are addressing these concerns through novel enzymatic techniques and more refined processing to produce a more
Albumin, in the form of bovine or serum albumin (BSA, HSA) is commonly used in cell culture media formulations for nutrient
transport. SFM is frequently supplemented with serum albumin as a carrier for fatty acids, lipids, amino acids, and trace
elements. Additional advantages of albumin as a cell culture supplement include its ability to bind toxic components present
in culture and protect against mechanical damage such as shear stress in agitated cell culture systems. The successful replacement
of BSA or HSA in SFM with recombinant forms of albumin or synthetic compounds such as pluronic has been achieved, however,
requirements for albumin vary depending on the cell line. For example, the myeloma cell line, NSO, lacks the functional pathway
for cholesterol synthesis and therefore requires cholesterol. Albumin has been used as a carrier of cholesterol, although
cyclodextrins have been used as alternative carriers of cholesterol and other lipids in culture media.
In recent years, a variety of recombinant animal-free forms of albumin (rHA) have become commercially available. Up and coming
areas requiring commercial cell culture processes, such as advanced tissue and stem cell therapies and regenerative medicine,
are areas where rHA is likely to prove a compliant and consistent alternative to current albumin sources.16 Recombinant albumin, such as CellPrime rAlbumin AF-S (Millipore, Billerica, MA), and Recombumin (Novozymes, Bagsvaerd, Denmark),
produced in Saccharomyces cerevisiae has been shown to be structurally identical to native albumin. Safety, tolerability, pharmacokinetics, and pharmacodynamics
also have been studied, showing equivalence to native human albumin.17