Principles of Cell Culture
Principles of Cell Culture
Principles of Cell Culture
Hessah Alshammari
1885: Roux maintained embryonic chick cells alive in saline solution for
short lengths of time
1912: Alexis Carrel cultured connective tissue and showed heart muscle
tissue contractility over 2-3 months
1970s: Gordon Sato et al. published the specific growth factor and media
requirements for many cell types
1979: Bottenstein and Sato defined a serum-free medium for neural cells
Secondary culture
Sub-culture
Cell Line
Single cell isolation
Immortalization
Successive sub-culture
Loss of control
of cell growth
Senescence
Genetically unstable
Fibroblastic
Epithelial
Endothelial
Neuronal
HEPES Strong chemical buffer range pH 7.2 7.6 (does not require CO 2)
Glucose
Energy Source
Containment level 2
cell culture laboratory
Sub-culture (passaging)
Count cells
Mr Frosty
Used to freeze cells
Cryopreservation
Passaging Cells
Resuspend in serum
containing media
70-80% confluence
100% confluence
Cryopreservation of Cells
Passage cells
Resuspend cells in serum
containing media
Centrifuge &
Aspirate supernatant
Resuspend cells in
10% DMSO in FCS
Transfer to cryovial
Freeze at -80oC
Transfer to liquid
nitrogen storage tank
Contamination
2-Biological Contamination
Bacteria and yeast
Viruses
Mycoplasmas
Cross-contamination by other cell culture
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