Download Types of Cell Lines

CELL LINES
In tissue culture, the cells growing in the first or later subculture from a primary culture.
It is a clone of cultured cells derived from an identified parental cell type.
Types of Cell Lines:
Finite Cell Lines:
The cells in culture divide only a limited number of times, before their growth rate declines and they
eventually die.
Continuous Cell Lines:
Cell lines transformed under laboratory conditions or in vitro culture conditions give rise to continuous
cell lines.
Maintenance of Cell Lines:
For the routine and good maintenance of cell lines in culture (primary culture or subculture) the
examination of cell morphology and the periodic change of medium are very important.
Cell Morphology:
The cells in the culture must be examined regularly to check the health status of the cells, the absence
of contamination, and any other serious complications (toxins in medium, inadequate nutrients etc.).
Replacement of Medium:
Periodic change of the medium is required for the maintenance of cell lines in culture, whether the cells
are proliferating or non-proliferating. For the proliferating cells, the medium need to be changed more
frequently when compared to non-proliferating cells. The time interval between medium changes
depends on the rate of cell growth and metabolism.
Cell Cloning:
In the traditional culture techniques, the cells are heterogenous in nature. Isolation of pure cell strains is
often required for various purposes.
Cell cloning broadly involves the processes connected with the production of a population of cells
derived from a single cell. Cloning of continuous cell lines is much easier compared to that of the
primary cultures, and finite cell lines.
Cloning may be carried out by two approaches- monolayer and suspension cultures:
1. Monolayer culture:
Petri dishes, multi-well plates or flasks can be used for cloning by monolayer culture. It is relatively easy
to remove the individual colonies of cells from the surfaces where they are attached.
Dilution Cloning:
Dilution cloning is the most commonly used technique for cloning of monolayer cells, and involves the
following stages
1. Trypsinization of cells (at log phase) to produce single cell suspensions.
2. Dilution of the cells to about 10-100 cells/ ml.
3. Seed the cells in multi-well dishes, petri dishes or plastic bottles.
4. Incubate under appropriate conditions for 1-3 weeks.
5. Isolate individual colonies.
2. Suspension culture:
Cloning can be carried out in suspension by seeding cells into viscous solutions (Methocel) or gels (agar).
As the daughter cells are formed in suspension, they remain intact and form colonies in suspension.
Suspension Cloning:
Certain cells, particularly the transformed fibroblasts and hematopoietic stem cells, can be more
conveniently cloned in suspension rather than monolayers. Suspension cloning can be carried out by
using agar or Methocel, which can hold the cells of a given colony together, and prevent mixing of
colonies. The technique of cloning in agar suspension is carried out in the following stages
1. Cells from suspension culture or monolayers can be used. The monolayer cells have to trypsinized
while the suspension cells can be directly used.
2. Count the cells and dilute serially so that 10-200 cells/ml are finally present.
3. Freshly prepared agar medium with appropriate dilution is used.
4. The agar medium is inoculated with the diluted cells.
5. Incubation for 1-3 weeks given clones in culture.
CELL SYNCHRONIZATION
Cell Synchronization is a process by which cells at different stages of the cell cycle in a culture are
brought to the same phase.
CELL PRESERVATION AND CHARACTERISATION
PRESERVATION
The preservation of cells is an extremely important aspect of cell culture. The only effective means of
preservation of animal cells is by freezing, which can be accomplished with either liquid nitrogen or by
employing cryogenic freezers.
The freezing process involves slowly reducing the temperature of prepared cells to -30 to -60°C followed
by a transfer to temperatures less than -130°C. Once at ultralow temperatures, the cells are biologically
inert and can be preserved for years.
Cryopreservation is defined as the viable freezing of biological material and their subsequent storage at
ultra-low temperatures.
Liquid nitrogen is most widely used material for cryopreservation, dry ice can also be used.
STEPS INVOLVED IN CRYOPRESERVATION:
 SELECTION OF PLANT MATERIAL
 PREGROWTH
 ADDITION OF CRYOPROTECTANTS
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Vitrification
Cryoprotective dehydration
Encapsulation and Dehydration
 FREEZING
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Rapid freezing
Slow freezing
Stepwise freezing
 STORAGE
 THAWING
 DETERMINATION OF SURVIVAL/VIABILITY
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TTC Method of staining
Evans blue staining
Fluorescein diacetate (FDA) staining
CHARACTERIZATION
STEM CELL LINES
A stem-cell line is a group of stem cells that is cultured in vitro and can be propagated indefinitely. Stemcell lines are derived from either animal or human tissues and come from one of three sources:
embryonic stem cells, adult stem cells, or induced stem cells.
By definition, stem cells possess two properties:
(1) they can self-renew, which means that they can divide indefinitely while remaining in an
undifferentiated state; and
(2) they are pluripotent or multipotent, which means that they can differentiate to form specialized cell
types.
TYPES OF STEM CELL LINE
1. Embryonic stem cell line
An embryonic stem cell line is created from cells derived from the inner cell mass of a blastocyst,
an early stage, pre-implantation embryo.
2. Adult stem cell line
Adult stem cells are found in juvenile or adult tissues. Adult stem cells are multipotent: they can
generate a limited number of differentiated cell types.
3. Induced pluripotent stem-cell (iPSC) line
Induced pluripotent stem cell (iPSC) lines are pluripotent stem cells that have been generated
from adult/somatic cells.