Read this article to learn about the development, advantages, disadvantages and commonly used serum-free media for animal cells and also its benefits over the serum media.
Addition of serum to the culture media has been an age-old practice. However, in recent years, certain serum-free media have been developed. It is worthwhile to know the disadvantages associated with the use of serum, and the advantages and disadvantages of serum-free media.
Disadvantages of serum in media:
Variable composition:
There is no uniformity in the composition of the serum. It is highly variable (source, batch, season, collection method, processing). Such differences in the composition significantly influence the cells in culture.
Quality control:
To maintain a uniform quality of the serum, special tests have to be performed with each batch of serum, before its use.
Contamination:
It is rather difficult to get serum totally free from all pathogens, particularly viruses.
Presence of growth inhibitors:
In general, the concentration of growth promoters in the serum is much higher than the inhibitors. But sometimes, the growth inhibitors such as TGF-P may dominate and inhibit cell proliferation.
Availability and cost:
There is a dependence on the cattle for the supply of serum. Hence the availability may be restricted on several occasions for political and economic reasons. Further, cost also is another factor for discouraging the use of serum.
Downstream processing:
The presence of serum in the culture medium interferes with the isolation and purification of cell culture products. For this reason, several additional steps may be required for the isolation of the desired product.
Advantages and Disadvantages of Serum-Free Media:
Advantages:
The limitations associated with the use of serum in the media (described above) are eliminated in the serum-free media. In addition, there are two more distinct advantages.
Selection of media with defined composition:
The main advantage of serum-free medium is to control growth of the cells as desired, with a well-defined medium. This is in contrast to the use of serum wherein the growth frequently proceeds in an uncontrolled fashion.
Regulation of differentiation:
It is possible to use a factor or a set of factors to achieve differentiation of cells with the desired and specialised functions.
Disadvantages:
Slow cell proliferation:
Most of the serum-free media are not as efficient as serum added media in the growth promotion of cells.
Need for multiple media:
A large number of serum-free media need to be developed for different cell lines. This may create some practical difficulties in a laboratory simultaneously handling several cell lines. Another limitation of serum-free medium is that a given medium may not be able to support the different stages of development even for a given cell line. Hence, sometimes separate media may be required even for the same cell line.
Purity of reagents:
The native serum does possess some amount of protective and detoxifying machinery that can offer a cleansing effect on the apparatus and reagents. And therefore, in the absence of serum, pure grade reagents and completely sterile apparatus should be used.
Availability and cost:
In general, the serum-free media are costlier than the serum added media. This is mainly due to the fact that many of the pure chemicals added to the serum-free media are themselves expensive. Further, the availability of serum-free media is also another limitation.
Development of Serum-Free Media:
While designing serum-free media, it is desirable to identify the various serum constituents and their quantities. The most important constituents of natural serum with reference to their use in cell cultures may be categorized as follows.
i. Growth regulatory factors e.g. PDGF, TGF-β.
ii. Cell adhesion factors e.g. vitamins.
iii. Essential nutrients e.g. vitamins, metabolites, minerals, fatty acids.
iv. Hormones e.g. insulin, hydrocortisone.
For replacing the serum and development of serum-free media, several constituents should supplement the media. Some highlights are given below.
Growth factors:
A large number of growth factors (nearly 100) that promote in vitro cell proliferation and differentiation have been identified. Besides the factors described already (above), some others are listed below.
i. Erythropoietin (EPO).
ii. Eye-derived growth factors (EDGF 1 and EDGF 2).
iii. Interleukins (IL-1, IL-2).
iv. Hepatocyte growth factor (HGF).
v. Brain-derived neurotrophic factor (BDNF).
vi. Phytohemagglutinin (PHA).
vii. Lipopolysaccharide (LPS).
The growth factors may act synergistically or additively with each other or with other factors (e.g. hormones, prostaglandins).
Almost all the growth factors are now commercially available for the preparation of serum-free media.
Hormones:
Growth hormone, insulin and hydrocortisone are the most commonly added hormones into the serum- free media. A combination of steroid hormones- hydrocortisone, estrogen, androgen and progesterone are used in formulating serum-free media for the maintenance of mammary epithelium.
Nutrients:
Addition of certain nutrients-choline, ethanol- amine, linoleic acid, iron, copper, selenium etc., are added in most of the serum-free media.
Proteins:
Bovine serum albumin (BSA) is the most commonly added protein. It promotes cell survival and growth.
Polyamines:
Putrescine is the most widely added polyamine to the serum-free media. Polyamines promote cellular growth and differentiation.
Protease inhibitors:
Addition of protease inhibitors (e.g. soy bean trypsin inhibitor) is done to the serum-free media for the trypsin-mediated subcultures.
Commonly Used Serum-Free Media:
Several types of serum-free media have been developed for different cell lines. A selected list of cell lines and the media is given in the Table 34.5.
