CELL CULTURE
(August 2004)
Animal or plant cells, removed from tissues, will continue to grow if supplied with the appropriate nutrients and conditions. When carried out in a laboratory, the process is called Cell Culture. It occurs in vitro (‘in glass’) as opposed to in vivo (‘in life’). The culture process allows single cells to act as independent units, much like a microorganism such as a bacterium or fungus. The cells are capable of dividing, they increase in size and, in a batch culture, can continue to grow until limited by some culture variable such as nutrient depletion.
Cultures normally contain cells of one type although mixed cultures, especially of bacteria, are common in food sciences and wastewater treatment related studies. The cells in culture may be genetically identical (homogenous population) or may show some genetic variation (heterogeneous population). A homogenous population of cells derived from a single parental cell is called a clone. Therefore all cells within a clonal population are genetically identical.
Why Grow Cells in Culture?
There are a number of applications for animal cell cultures:
- To investigate the normal physiology or biochemistry of cells. For instance studies of cell metabolism.
- To test the effect of various chemical compounds or drugs on specific cell types (normal or cancerous cells, for example).
- To study the sequential or parallel combination of various cell types to generate artificial tissues e.g. artificial skin. Possibility of generating artificial tissues is an emerging and intensively studied area of biotechnology known as “tissue engineering”.
- To synthesize valuable biologicals from large scale cell cultures. The biologicals encompass a broad range of cell products and include specific proteins or viruses that require animal cells for propagation. For example, therapeutic proteins can be synthesized in large quantities by growing genetically engineered cells in large-scale cultures. The number of such commercially valuable biologicals has increased rapidly over the last decade and has led to the present widespread interest in animal cell culture technology.
The major advantage of using cell culture for any of the above applications is the consistency and reproducibility of results that can be obtained from using a batch of clonal cells. The disadvantage is that, after a period of continuous growth, cell characteristics can change and may become quite different from those found in the starting population. Cells can also adapt to different culture environments (e.g. different nutrients, temperatures, salt concentrations etc.) by varying the activities of their enzymes.
Milestones of Cell Culture
The history of cell culture dates back to early twentieth century. The original impetus for the development of cell culture was to study, under the microscope, normal physiological events such as nerve development. The growth rate of animal cells is relatively slow compared with bacteria. Whereas bacteria can double every 30 minutes or so, animal cells require anywhere from 18 to 24 hr to double. This makes the animal culture vulnerable to contamination, as a small number of bacteria would soon outgrow a larger population of animal cells. Consequently, animal cell culture did not become a routine laboratory technique until the 1950s.
The need for cell culture, especially at large scales, became apparent with the need for viral vaccines. Major epidemics of polio in the 1940s and 1950s promoted a lot of effort to develop an effective vaccine. When it was shown in 1949 that poliovirus could be grown in cultures of human cells, considerable interest was shown to develop large quantities of the polio vaccine using cell culture. The polio vaccine, produced from de-activated virus, became one of the first commercial products of cultured animal cells.
Recombinant DNA technology (also known as genetic engineering) was developed in the 1970s to express mammalian genes in bacteria. It soon became apparent that large complex proteins (and especially those having therapeutic value) couldn’t be produced in bacteria, as they do not have the appropriate metabolism to add sugar chains to these proteins. Therefore, genetically engineered animal cells were developed for large-scale commercial production of such important proteins.
Another milestone in the animal cell culture technology came in 1975 with the production of hybrid cells (known as hybridoma) from the fusion of two or more cells capable of continuous production of a single type of antibody. These antibodies have diagnostic and therapeutic value and are now produced commercially in kilogram quantities from large-scale cultures of hybridomas.
Plant Cell Culture
Plant cells have been cultured to produce many ingredients needed by the food industries. Tremendous progress has been made in recent years in understanding the basics of plant metabolism and in the development of bioprocesses as well as design and operation of large-scale bioreactors for plant cell culture. A wide range of food ingredients including flavors, colorants, essential oils, sweeteners and antioxidants have been produced in culture. Japan has so far been the most successful country in the world to carry out plant cell culture on commercial scale. Ginseng products derived from cell suspension cultures of Panax ginseng used as an additive in wine, tonic drinks and herbal liquors have been produced by a company in Japan since 1990 with a net sale of 3 million dollars in 1995.
Types of Mammalian Cultures
Freshly isolated cultures from mammalian tissues are known as primary cultures until sub-cultured. At this stage, cells are usually heterogeneous but still closely represent the parent cell types as well as in the expression of tissue specific properties. After several sub-cultures onto fresh media, the cell line will either die out or ‘transform’ to become a continuous cell line. Such cell lines show many alterations from the primary cultures including change in morphology, chromosomal variation and increase in capacity to give rise to tumors in hosts with weak immune systems.
Animal cells can be grown either in an unattached suspension culture or attached to a solid surface. Suspension cultures have been successfully developed to quite large bioreactor volumes, with successful production of viruses and therapeutic proteins.
The range of commercially available biologicals produced by cell culture technologies has increased rapidly over the last decade. This is particularly so for therapeutic proteins synthesized from selected or genetically engineered mammalian cells. They are needed in large quantities and hence require careful study of the underling biochemical, cell biological and engineering principles for control of production processes.