32.2: Cell Culture
Most vertebrate cells grow in vitro attached to a substrate as a monolayer, called adherent cultures. The flasks and plates used to grow cells are chemically treated to facilitate cell attachment. However, a few cell types, such as hematopoietic cells, can grow in a suspension. In contrast to adherent cultures, suspension cultures can grow in non-treated cultureware using magnetic stirrers or spinner flasks to agitate the culture media
Culture conditions
The growth medium is a crucial component for the optimal growth of cells. Previously, natural growth media like blood plasma, chicken embryo extracts, and amniotic fluid were used. In recent times, however, synthetic media such as Dulbecco's Modified Eagle Medium (DMEM) and Roswell Park Memorial Institute (RPMI) are preferred. These basal media contain amino acids, vitamins, inorganic salts, and a carbon source such as glucose. They are supplemented with fetal bovine serum (FBS) as a source of growth and adhesion factors, hormones, lipids, and minerals. Antibiotics are added to the growth media to prevent microbial contamination, and culturing protocols are performed under aseptic conditions.
The pH of the medium is usually maintained at 7.4 for mammalian cells. The growth medium also contains a buffering system such as sodium bicarbonate with exogenous CO2 for pH regulation. A pH indicator like phenol red is incorporated into the culture medium to indicate pH changes. An optimum temperature of 37℃ and a CO2 concentration of 5% are maintained in the incubator.
Plant cell culture
Like animal cells, plant cells can also be cultured in vitro. Plant tissue, known as explant, is cultured in a nutrient medium containing plant growth hormones, micronutrients, and a carbon source. Relative proportions of the plant growth hormones auxins and cytokinins in the medium decide the tissue type developed in the culture. A balanced ratio of these hormones results in a mass of undifferentiated cells called the callus. By altering the hormone ratio, the callus can be differentiated into root or shoot, thus generating a complete plant. Explants are usually cultured on a solid nutrient medium, while single-cell suspensions can be grown in liquid media. Growth conditions like temperature, light intensity, and photoperiod govern the plant growth in culture. Plant cell cultures are used to produce improved hybrid plants and conserve endangered plants. Explant cultures are also used in the large-scale production of plant-derived products.
