Mammalian Cell Culture

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Mammalian Cell Culture
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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

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If we treat the cells right

• They might act similar to those in vivo
• Making the same proteins
• Then scientists can change the environment

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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

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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

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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

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What is in the media?

• Dulbecco Modified Eagles 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)

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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

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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.

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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.

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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.