The ideal animal cell culture media is expected to possess certain physiological and chemical properties such as pH, oxygen, carbon dioxide, buffering, osmolality, viscosity, temperature in such as way that it should support growth and proliferation of cultured animal cells.
Normally most of the animal cells can grow at pH 7-7.4, although there may be some slight variations in the required optimum pH depending on the type and origin of cells. The pH indicator phenol red is most commonly used for visible detection of media pH.
Carbon dioxide, Bicarbonate and Buffering:
1. Carbon dioxide in the animal cell culture media is in a dissolved state and its concentration depends on the temperature and also the atmospheric carbon dioxide tension. Carbon dioxide in the medium exists in many forms such as carbonic acid, HCO3 and hydrogen ions.
2. Concentration of carbon dioxide, bicarbonate and pH are interdependent. That is for example by increasing the concentration of atmospheric carbon dioxide decreases the pH of the medium, this in turn will make cell culture medium acidic. Addition of sodium bicarbonate as bicarbonate buffer neutralizes the bicarbonate ions, which are present in the media.
3. The presence of pyruvate in the animal cell culture media increases the endogenous production carbon dioxide by the cells.
1. The proliferation and scale up of animal cell culture depends up on the oxygen supply without causing cell damage.
2. Oxygen is sparingly soluble in cell culture media. But cultured cells depend on the dissolved oxygen present in the media. But high concentration of the same may act as toxic to the cultured cells due to the generation of free radicals. Therefore oxygen must be supplied in such a way that it full fills all the cell requirements and avoids toxic effects on cultured animal cells.
3. Free radical scavengers such as glutathione are used to nullify the toxic effect caused by the high concentration of oxygen.
4. Addition of Selenium to the culture medium also reduces the oxygen toxicity, as selenium act as cofactor in the production of glutathione.
5. Cultured cells when compared to in vivo cells undergo anaerobic form of glycolysis and also the depth of cell culture media influence the rate of oxygen diffusion.
1. Animal cells are more sensitive to temperature higher than the body temperature, that is 37c. But depending on the origin of cultured cells optimum temperature varies, for example cell culture from birds is around 38.5c and cells cultured from cold blooded animals is around 15-25c.
2. Animal cell culture media has low thermal conductivity, high temperature gradient can occur with heating and mixing of the media.
3. In a small or compact bioreactor low gradient heating system can be achieved using a thermo stated draught tube.
4. Temperature of the culture media influences the animal cell growth, and also temperature also affects the oxygen solubility, that is higher temperature increases the solubility of oxygen.
1. Osmolality for most of the cultured animal cells is around in the range of 260-320mosm/kg, comparable to the osmolality of human plasma, which is around 290mosm/kg.
2. Once the osmolality for a cell culture media is selected, then it must be maintained at that level throughout the culture.
3. Addition of acids, bases, drugs or any other compounds to the cell culture medium alters the osmolality and the same is measured using Osmometer.
Growth factors promote in vitro proliferation and differentiation of cultured cells. Growth hormones such as insulin and hydrocortisone are most commonly used in serum free ar artificial animal cell culture media. For example combination of steroid hormones such as hydrocortisone, estrogen and also androgen are used in maintaining the mammary epithelial cells in the laboratory.
Animal cell culture media also contains nutrients such as cholin, ethanol amine, linolic acid, and also minerals like iron, copper, selenium and much more are added, in such a way that combination of all these nutrients promote the proliferation of cultured animal cells.
Most commonly added protein into animal cell culture media is Bovine Serum Albumin (BSA). This protein promotes the cell growth and it also increases the survival rate of animal cells.
Likewise polyamines and protease inhibitors are also added into the animal cell culture media.
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