animal tissue culture lecture 2
TRANSCRIPT
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Introduction to
Animal Tissue culture
LECTURE OF SUBJECT :
Dr. sharafaldin Al-musawi
College of Biotecholgy
LECTURE: 2
SUBJECT: Animal Tissue culture
LEVEL: 4
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Why do we need Cell culture?Research
To overcome problems in studying cellular behavior such as:• confounding effects of the surrounding tissues. • variations that might arise in animals under experimental
stress.Reduce animal use.
Commercial or large-scale production.Production of cell material: vaccine, antibody, hormone.
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Initiation of culture
Tissue
Primary culture
Cell line Continuous cell line
Subculture
Stored Stored
Animal Plant
Finite numbers Indefinite numbers
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Types of Cell culture
1. Primary Cultures
Derived directly from excised tissue and cultured either as:
Outgrowth of excised tissue in culture
Dissociation into single cells (by enzymatic digestion or mechanical dispersion).
Primary Culture
Preparation
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Characteristics of Primary Cultures
5
Primary Culture
Preparation
Characteristics:
Morphologically similar to the parent tissue.
Limited number of cell divisions.
Best experimental models for in vivo
situations.
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Advantages & Disadvantages• Advantages:
• usually retain many of the differentiated characteristics of the cell in vivo
• Disadvantages:
• Initially heterogeneous but later become dominated by fibroblasts.
• The preparation of primary cultures is labor intensive.
• Can be maintained in vitro only for a limited period of time.
• Difficult to obtain.
• Relatively short life span in culture.
• Very susceptible to contamination.
• May not fully act like tissue due to complexity of media.
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Types of Cell culture2. Continuous Cultures
derived from subculture (or passage, or transfer) of primary culture
Subculture : The process of dispersion and re-culture the cells after they have increased to occupy all of
the available substrate in the culture.
usually comprised of a single cell type.
can be serially propagated in culture for several passages.
There are two types of continuous cultures
Cell lines
Continuous cell lines
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Types of continuous culture1) Cell lines
• Cell lines derived from primary cultures have a limited life span.
• After the first subculture, the primary culture becomes cell line.
• finite life, senescence after approximately thirty cycles of division.
• usually diploid and maintain some degree of differentiation.
• It is essential to establish a system of Master and Working banks in order to
maintain such lines for long periods.
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Types of continuous culture
2 )Continuous cell lines
can be propagated indefinitely
generally have this ability because they have been transformed by:
tumor cells.
viral oncogenes
chemical treatments
Spontaneously
disadvantage: having very little of the original in vivo characteristics
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Transformation VS Transfection
• Transformation
• Spontaneous or induced permanent phenotypic changes resulting from change in DNA
and gene expression that result and effect in:
• growth rate
• mode of growth )loss of contact inhibition)
• specialized product formation
• longevity
• loss of need for adhesion
• Transfection
• Introduction of DNA into a cell (like viral DNA)
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Cell Culture Morphology• Morphologically cell cultures take one of two forms:
1. growing in suspension )as single cells or small free-floating clumps) such as: cell lines derived from blood )leukemia, lymphoma).
2. growing as a monolayer that is attached to the tissue culture flask. Such as: Cells from solid tissue )lungs, kidney, breast), endothelial, epithelial, neuronal, fibroblasts.
Hela-Epithelial
MRC5-Fibroblast SHSY5Y-Neuronal
BAE1-Endothelial MCF-7 breast
HT1080- kidney 3LL - lungs
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• Excellent model systems for studying:The normal physiology and biochemistry of cells.
The effects of drugs and toxic compounds on the cells.
Mutagenesis and carcinogenesis.
• Used in drug screening and development
• Large scale manufacturing of biological compounds
(vaccines, insulin, interferon, other therapeutic protein)
Cell culture application