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Mammalian Cell Culture

Mammalian Cell Culture. What is cell culture, exactly?. Cells, previously growing in a human or animal modified to grow in plastic or glass In the body = in vivo On plastic or glass = in vitro Kept in an incubator to stay at body temperature

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Mammalian Cell Culture

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  1. Mammalian Cell Culture

  2. What is cell culture, exactly? • Cells, previously growing in a human or animal modified to grow in plastic or glass • In the body = in vivo • On plastic or glass = in vitro • Kept in an incubator to stay at body temperature • We use special media with nutrients so the cells can grow and divide

  3. If we treat the cells right • They might act similar to those in vivo • Making the same proteins • Then scientists can change the environment

  4. Animals are complex • Many different cells • Many different proteins • Interacting continuously • Difficult to watch individual events in vivo • Animals are usually harmed to observe biological events

  5. Using cultures has advantages • Fewer animals are harmed • Can control all external factors • Can easily test what the cells are doing • Cells are easy to manipulate and propagate • All of the cells are the same • Results of experiments will be consistent • Cheaper to maintain

  6. What can we do with cells? • Test pharmaceutical drugs • Watch disease mechanisms • Design potential treatments • Observe the regenerative process • How do cells and tissues repair themselves after damage from illness or injury? • Observe the developmental process

  7. What is in the media? • Dulbecco’ Modified Eagle’s Media (DMEM) • Contains glucose, some proteins, and essential salts • Contains a pH indicator (phenol red) Media looks pink/red at pH 7.2 • Acidic -yellow or orange (cell growth, bacterial growth) • Basic -purple (no cell growth, not enough CO2)

  8. More media components • Antibiotics (penicillin and streptomycin) • Prevent bacterial contamination • Salts and buffers • To simulate in vivo environment • Serum • Portion of blood after the cells and fibers have clotted • From cow, horse, sheep • added to media as a nutrient source for growing cells • Lipids, proteins

  9. Phosphate Buffered Saline • Used to wash/remove excess serum that inhibits the function of TRED. • Must be warmed in the water bath before use so cells are not shocked by cold liquid.

  10. Trypsin EDTA (TRED) • An enzyme used to detach the cells from a culture dish. • EDTA binds calcium ions in the media that would normally inhibit trypsin.

  11. Bleach • Used to destroy any remaining cells in dishes and tubes before they are tossed in the trash can. • Add enough to change media to clear, • wait 5 minutes, • rinse solution down sink • throw away the dish/flask/plate in the trash can.

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