Transcript Introduction to Cell Culture

Introduction to Cell Culture
Biotechnology I
Cell Culture
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Definition: the in vitro growth of cells isolated
from multi-cellular organisms
Process: Cells will continue dividing until they
fill up the container; cell to cell contact stops
cell division
Uses: vaccines, research of all kinds
including stem cell, recombinant DNA,
production of antibodies
Types of Cell used
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Mammalian cells are commonly used but
they do grow slower and are more fragile
than bacterial cells. Also have more complex
nutrient requirements
Bacterial cells were first used to make
genetically engineered products and they
reproduce very quickly
Growing Bacterial Cells
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Choose bacteria – E. coli most common
Make media
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Petri plates use agar in media (Luria Broth,
nutrient agar)
Liquid cultures use broth (LB, nutrient broth)
Sterilize media in autoclave
Pour media plates
Innoculate media
Grow cells in incubator (37oC)
Equipment Required for Mammalian Cells
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CO2 incubator: for
proper atmospheric
conditions
Laminar flow hood: for
sterility
Hemocytometer: for
cell counting
Inverted Microscope:
for viewing cells on
hemocytometer
Cell Culture Process
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Contaminating the cell culture is area of
concern
To avoid contamination mfg. techs do the
following:
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Sterilize all containers used
Change the media frequently
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Media is the source of nutrition, pH indicator often times
included in media to help monitor pH which is critical to
cell survival
Cell Culture Process
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Count cells
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Using a hemocytometer the cells are counted to
determine if they are ready to harvest
Can stain cells with trypan blue to determine if
they are still alive. All cells which exclude the dye
are viable. All stained cells are dead.
Dead cell
Live cell
Cell Culture Process
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View cells
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Inverted microscope is used
to determine cell growth and
activity
Adherent cells are ones which
are stuck to the flask or vessel
Cell counting video for cell
culture