Animal Cell Culture: Growth of animal cells in culture.
Introduction:
Animal cell culture is a microbiological technique in which animal cells and tissues are obtained and maintained in a suitable environment.
It is further divided into,
Organotypic: Animal Organ Tissue Culture.
Histotypic: Animal Cell Culture.
Animal cells can be grown only upto certain generations.
Their growth requires typical conditions and a physicochemical environment.
Animal cell culture prepared from collecting tissue or cells from an organism is called a “Primary culture.”
The sequence of subcultures prepared from primary culture is called “Cell lines”.
History:
It was Ross Harisson from America who first cultured animal cells successfully in the laboratory.
Campbell, Wilmut and colleagues at Roslin Institute in Scotland successfully cloned the first animal cell using nuclear transfer technique and gave birth to a sheep named “Dolly” in 1996.
Growth of Animal Cells in Culture.
Growing animal cells in cultures is a difficult task, it requires specially made culture media.
The culture media used for animal cell culture are classified as,
Natural,
Artificial,
Synthesized.
The selection of culture media depends on the objective of experiment and type of cell to be cultured.
Natural Culture Media:
As the name indicates these media are obtained from natural resources like, tissue extracts, biological fluids, plasma clots or coagulants etc.
Blood Plasma:
It provides good nutrition and support to many types of animal cell culture.
During subculturing it provides good protection against traumatic damage and better attachment of cells to glass surfaces.
Chicken plasma is most favored for its clarity even on dilution.
The plasma is obtained by centrifugation before coagulation.
This tissue or cells are placed in such plasma and then coagulation is initiated by addition of thrombin.
Blood Serum:
Blood serum is plasma without fibrinogen.
Blood serum with or without nutritional substances is used as a culture media to grow animal cells.
The sera commonly used in animal culture are bovine, horse, and human.
Calf and fetal bovine serum are most widely used sera in animal cell culture.
Human serum is used sometimes but is needed to be screened for presence of hepatitis and HIV.
The composition of a serum is highly complex as it is a mixture of various plasma proteins, peptides, lipids, carbs, enzymes and minerals.
Tissue Extracts:
Commonly used tissue extracts in animal cell culture are, embryo, spleen, liver, bone marrow etc.
CHick embryo extract is the most commonly used tissue extract in animal cell culture, it is obtained from 10-12 days old embryos.
2) Artificial Media:
Synthetic or artificial media are prepared by adding organic or inorganic substances, nutrients, vitamins, proteins, carbs, gaseous phases.
Different artificial media are prepared for different types of cells as per their requirement.
Some common examples of artificial media are,
Minimal Essential Medium (MEM),
CMRL 1066,
RPMI 1640.
Synthetic media re classified as,
Serum Containing Media.
Serum Free Media.
Serum Containing Media:
It is prepared by adding 5 to 20 % serum in serum free medium.
Serum is one of the best sources of many basic nutrients, contains hormones, growth factors, proteins, and minerals.
Serum also acts as a buffer.
Serum combines and neutralises many toxins.
Although its a best source of nutrients it has following disadvantages,
Most expensive of all ingredients of the culture medium.
It increases difficulties and cost of downstream processing.
Its content varies from batch to batch.
Extensive testing is required to check similarity on serum change.
Supply gets affected in drought or cattle disease spread.
Source of contamination: May contain viruses, bacteria, prions etc.
Some growth factors may be inadequate and can not be supplemented.
Serum Free Media:
They are developed to overcome limitations of serum containing media.
These media can make the medium more selective for particular cells as per their requirements e.g. melanocytes can be cultivated in absence of keratinocytes.
As serum is not present there is no problem of toxicity, serum change tests.
Bioassays and other processes become easy due to absence of the serum.
However, they also has some disadvantages like,
Slower growth.
Useful only for a few generations.
Purity of ingredients.
pH control and temperature control required is more.
Most media are specific to a particular cell type.
Preparation time is more.
Physicochemical Parameters needed for growth animal cell culture:
Control on different physicochemical parameters is necessary during animal cell culture.
The temperature of the culture has to be the same as that of the body temperature of the animal from which cells are collected.
Osmolarity between 260 m Osm/kg and 320 m Osm/kg is good for many cell lines.
Buffers are incorporated for maintaining the pH as per requirement of the cell culture.
Most cell cultures grow well at a pH of 7.4, phenol red is commonly used indicator to detect the pH change in medium.
External Carbon dioxide addition is needed for some cell cultures to replenish CO2 loss.
Oxygen is a component of the gas phase as most cells require oxygen for respiration.
Cell damage in the culture is prevented by increasing viscosity of the medium by addition of CMC or PVP.
Balanced Salt Solution (BSS) is used as a diluent for vitamins, amino acids to make complete media.
Commonly Asked Questions:
What is the difference between Blood Plasma and Blood Serum? (For Viva)
Write in short about culture media used in animal cell culture.
Labels: Microbiology
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