Culture of animal cells and tissue is rather more difficult than that of microorganisms and plants because the later synthesize certain chemical constituents from inorganic substances. However, the culture media provide the optimum growth factors (e.g. pH, osmotic pressure, etc) and chemical constituents (unlike microbes). There are two types of media used for culture of animal cell and tissue, the natural media and the synthesized media.
Natural media are the natural sources of nutrient sufficient for growth and proliferation of animal cells and tissue. These are of three types : (i) coagulans or plasma clots (it is used since long time but now available in market in the form of liquid plasma kept in silicon ampoules or lyophilized plasma. Plasma may also be prepared in laboratory taking out blood from male fowl and adding heparin to prevent blood coagulation), (ii) biological fluid (it is obtained in the form of serum from human adult blood, placental, cord blood, horse blood, calf blood or in the form of biological fluids such as coconut water, amniotic fluid, pleural fluid, insect haemolymph serum, culture filtrate, aqueous humour (from eyes), etc. The most commonly used fluids are human placental, cord serum and foetal calf serum. Before use its toxicity should be checked, and (iii) tissue extract (extract from some tissues such as embryo, liver, spleen, leukocytes, tumour, bone marrow, etc. are also used for culture of animal cells, where embryo extract is of most common use. Tissue extract should be used before a week or stored at 27°C.
Synthetic media are prepared artificially by adding several nutrients (orgtmie and inorganic), vitamins, salts, O2 and CO2 gas phases, serum proteins, carbohydrates, cofactors, etc. However, different types of synthetic media may be prepared for a variety of cells and tissues to be cultured. It can be prepared for different functions. Basically, synthetic media are of two types, serum-containing media (i.e. the media containing serum) and serum-free media (i.e. media devoid of serum). Example of some of the media are: minimal essential medium (MEM) (Eagle, 1955), 199 (Morgan et al. 1950), CMRL 1066 (Parker et al, 1957), RPMI 1640 (Moore et al, 1967) and F12 (Ham, 1965). Some of these media are given in Table 6.1.
Table 6.1. Chemical composition of different media (with serum) used for animal cell and tissue culture (quantities are in mg/l).
When the synthetic media are devoid of serum in culture medium, it is called serum-free media. Example of some serum-free media for certain cells and cell line are given in Table 6.2. By doing so the medium could be made selective for a particular type of cells because each type of cells requires different chemical constituents and physical factors. Serum-free media should not be used commonly until cheap and better serum-free media are available. Serum itself has several disadvantages as given below: (i) it deteriorates within a year and differs with batches, (ii) a number of batches are required if more than one cell types are used which make difficult for maintaining and co-culturing of cells difficult, (iii) supply of serum is less than its demand, therefore, medium becomes several times costly, and (iv) undesirable growth stimulation and inhibition may occur. Fiechter (1996) has enlisted the advantages and disadvantages of using the serum in culture media (Table 6.3).
Table 6.2. Serum-free medium for certain cell and cell lines.
Table 6.3. Advantages and disadvantages of serum in culture media. Serum contains a complete set of essential growth factors, hormones, attachment and
spreading factors, binding and transport proteins.It binds and neutralizes toxins.It contains protease inhibitors.It increases buffering capacity.It provides trace elements and other nutrients.
Source : based on Fiechter (1996).
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