This document provides an overview of cell culture techniques. It discusses the history of cell culture beginning in the late 1800s. Key terms like cell line, normal cell line, and continuous cell line are defined. The document outlines the biology of cultured cells including adhesion, proliferation, differentiation, and energy metabolism. It also discusses the origin of cultured cells and characterization techniques for authenticating cell lines such as morphology, chromosome content, and DNA analysis. The applications of animal cell culture and conclusions are presented.

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By
KAUSHAL KUMAR SAHU
Assistant Professor (Ad Hoc)
Department of Biotechnology
Govt. Digvijay Autonomous P. G. College
Raj-Nandgaon ( C. G. )

2. SYNOPSIS
• Introduction
• History
• Important terminology
• Biology of culture cell
• Characterization of culture cell
• Application of animal culture
• Conclusion
• References
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3. INTRODUCTION
• Cell culture refers to the process by which cells
are grown in a controlled artificial environment.
• Cells can be maintained in vitro outside of their
original body by this process which is quite
simple compared to organ and tissue culture.
• In a cell culture technique, cells are removed from
an animal or a plant, and grown subsequently in a
favorable environment.
• The cells may be removed directly or by
mechanical or enzymatic action.
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4. HISTORY
• 1885: Roux maintained embryonic chick cells in a saline
culture.
• 1897: Loeb demonstrated the survival of cells isolated
from blood and connective tissue in serum and plasma.
• 1911: Lewis and Lewis made the first liquid media
consisted of sea water, serum, embryo extract, salts and
peptones.
• 1916: Rous and Jones introduced proteolytic enzyme
trypsin for the subculture of adherent cells.
• 1923: Carrel and Baker developed ‘Carrel’ or T-flask as
the first specifically designed cell culture vessel.
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5. IMPORTANT TERMINOLOGY
• Cell line: Once a primary culture is sub-
cultured or passaged.
• Normal cell line: Divides a limited number of
times.
• Continuous cell line: cell line having the
capacity for infinite survival.
• Characterization: it is the refer many traits of
the cell line..some of which may be unique.
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6. BIOLOGY OF CULTURED CELL
• The Culture Environment
• Cell Adhesion
• Cell Proliferation
• Differentiation
• Cell Signaling
• Energy Metabolism
• Origin of Cultured Cells
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7. CELL ADHESION MOLECULE
• Cell Adhesion Molecules
• Intercellular Junctions
• Extracellular Matrix
• Cytoskeleton
• Cell Motility
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10. ORIGIN OF CULTURED CELL
• Initiation of the Culture
• Evolution of Cell Lines
• Senescence
• Transformation and the Development of
Continuous Cell Lines.
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11. EVOLUTION OF CELL LINES:
•After the first passage (or subculture), the primary
culture becomes known as a cell line and may be
propagated a number of times.
•By the 3rd passage, the culture becomes more stable.
•Normal cells can divide only a number of times and
then will die out a phenomenon known as senescence.
• Senescence is attributed to the lack of telomerase
(responsible for replicating the telomere sequence),
which results in shortened telomeres and stop of cell
proliferation.
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12. CHARACTERIZATION OF THE
CULTURE CELL
• Need for characterization
• Authentication
• Provenance
• Parameter of characterization
• Cell morphology
• Chromosome content
• DNA analysis
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13. NEED FOR CHARACTERIZATION
• Authentication, i.e., confirmation that the cell
line is not cross-contaminated or misidentified.
• Confirmation of the species of origin.
• Correlation with the tissue of origin.
• Identification of specific cell lines within a
group from the same origin, selected cell
strains, or hybrid cell lines.
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14. PARAMETER
• if molecular technology is readily available, then
DNA profiling or analysis of gene expression
likely to be of most use.
• Whereas a cytology laboratory may prefer to use
chromosome analysis coupled with FISH and
chromosome painting.
• A laboratory with immunological expertise may
prefer to use MHC analysis (e.g., HLA typing)
coupled with lineage specific markers. Combined
with a functional assay related to your own
interests.
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15. CELL MORPHOLOGY
• Study of the size, shape and structure of the cell.
• Most cells in culture can be divided into five
basic categories based on their morphology.
a) Fibroblastic
b) Epithelial-like
c) Lymphoblast-like
d) Endothelial
e) neuronal
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16. CHO-K1
VERO CELL LINE
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17. CHROMOSOME CONTENT
• Chromosome preparation
• Chromosome banding
• Chromosome analysis
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18. •Chromosome content or karyotype is one of the most characteristic and best-defined
criteria for identifying cell lines and relating them to the species and sex from which
they were derived.
•Chromosome analysis can also distinguish between normal and transformed cells
because the chromosome number is more stable in normal cells.
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19. NORMAL HUMAN KARYOTYPE
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21. DNAANALYSIS
• DNA Hybridization.
• DNA Fingerprinting.
• DNA Profiling
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22. DNA HYBRIDIZATION
• Hybridization of specific molecular probes to unique DNA
sequences (Southern blotting) can provide information about
species specific regions, amplified regions of the DNA, or
altered base sequences that are characteristic to that cell line.
• cells over expression of a specific oncogene, or oncogenes in
transformed cell lines or deletion, or loss, of heterozygosity in
suppressor genes.
• Although DNA aberrations can be detected in restriction
digests of extracts of whole DNA, this is limited by the
amount of DNA required.
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23. DNA FINGERPRINTING
• DNA contains satellite DNA, non-transcribed regions that are highly
repetitive, and of variable length, with minisatellite.
• These regions are not highly conserved because they are not transcribed,
and they give rise to regions of hyper variability.
• When the DNA is cut with specific endonucleases, sequences of interest
may be probed with cDNAs that hybridize to these hyper variable regions
or they may be amplified by PCR with specific primers.
• Electrophoresis reveals variations in fragment length in satellite DNA
(restriction fragment length polymorphisms, RFLPs) that are specific to the
individual from which the DNA was derived.
• When analyzed by polyacrylamide electrophoresis, each individual’s DNA
gives a unique hybridization pattern as revealed by autoradiography with
radioactive or fluorescent probes.
• These patterns are known as DNA fingerprints and are cell line specific
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24. CHARACTERIZATION OF CELL LINES AND
CELL STRAINS
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25. APPLICATION OF CELL CULTURE
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26. CONCLUSION
• biological studies of culture cell such as cell proliferation,
differentiation and their maintenance, cell cycle and their
regulation is very important in the field of genetic
engineering.
• Characterization of cell line is the first indispensible step after
each cell line is generated for determining its functionality,
authenticity, contamination, origin etc.
• Morphology, chromosome and DNA analysis have now
became the major standard procedure for cell line
identification.
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27. REFERENCES
• Culture of animal cells: R. IAN FRESHNEY (6th
Edition).
•
https://www.slideshare.net/varijnayan1/characte
rization-
•
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC
5661806/
• file:///C:/Users/dell/Downloads/Chap6%20Anim
al%20Cell%20Culture%20biology.pdf
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