General Procedure for Cell Culture

The cell culture methods are either applied to free-living organisms (bacteria or eukaryotic microorganisms), or to cells removed from multicellular tissues. Cells can be cultured for a prolonged time if they split regularly. After that the growth medium is replaced and the cells are diluted (after first detaching them by trypsin of NaOH from the support).

For culturing cells efficiently, the environment in which they found themselves should be maintained before they are transplanted to the artificial environment (an extracellular fluid derived from blood). This extracellular fluid is recreated using Ham’s tissue culture medium (commonly for mammalian cells).

Some cells (like those in bloodstream) survive without attaching to a surface; while others (like those derived from solid tissues) require a special surface. There are also cells like yeast and some bacteria types which survive under either condition and exhibit different phenotypes depending on whether or not they are attached to a surface.

In most of the cells derived from tissues, nutrients are provided by a liquid broth that washes the cells attached to a surface. Some cells require an “air-liquid interface” for their growth; the cells, in this case, are grown on a raft of organic material floating on the surface of a nutrient broth and acting like a wet sponge to feed the cells from underneath while the tops are exposed to the air.

For bacteria and yeast, small quantities of cells are grown on a solid support enriched with nutrients embedded in its stiff nutritious Jello, while large-scale cultures are grown with the cells suspended in a nutrient broth.

A population of cells derived from a single parental cell is called a clone, which may be derived from continuous cell lines or from primary culture. Cloning within a cell population reduces the degree of genetic and phenotypic variation.

Initiation of cell culture can be discussed under the following heads:

  1. Preparation and sterilisation of the substrate and culture vessel,
  2. Preparation and sterilisation of the medium,
  3. Isolation of cells and tissues,
  4. Disaggregation of tissues,
  5. Primary cell culture,
  6. Sub-culture,
  7. Cloning of cell lines, and
  8. Contamination in cell culture.

Animal Cell Culture

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Santhakumar Raja

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