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Cell and Tissue Culture

General Concepts

Ece ÖZYAĞCI
Selay TORNACI
Ebru TOKSOY ÖNER
Marmara University, Department of Bioengineering Organization Levels of the Body

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts General Concepts
Biologists recognize 12 levels of organization in the living world.
From the simplest to the most complex, these levels include

ü atoms,
ü molecules,
ü organelles,
ü cells,
ü tissues,
ü organs,
ü organ systems,
ü organisms,
ü populations,
ü communities,
https://www.youtube.com/watch?v=HmMeOrgcuKo ü ecosystems,
ü biospheres.
Organization of Life
from atoms to Biosphere…

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General Concepts General Concepts

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts General Concepts

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts General Concepts

•Epithelial tissue is made up of cells that line

•Connective tissue is made up of cells that
form the body’s structure. Examples include
bone and cartilage.
inner and outer body surfaces, such as the skin and
the lining of the digestive tract. Epithelial tissue
protects the body and its internal organs, secretes
substances such as hormones, and absorbs
substances such as nutrients.

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts General Concepts

•Muscle tissue is made up of cells that have
the unique ability to contract, or become
shorter. Muscles attached to bones enable the
body to move.
•Nervous tissue is made up of neurons, or
nerve cells, that carry electrical messages.
Nervous tissue makes up the brain and the
nerves that connect the brain to all parts of the
body.

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General Concepts General Concepts

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts General Concepts - What is cell culture?

It is estimated that the ü Observing human cells is crucial for understanding

human body contains
about 37.2 trillion
cells.
Many cells assemble
into a specific type
of “tissue.” One or
more tissues work
together as an
“organ.”
human health and treating diseases.
ü By studying cells, we can gain insight into how they
function and use this knowledge to diagnose, monitor,
and treat various conditions.
ü To effectively study cells, it is necessary to have a way to
observe them continuously.
ü Ideally, we should be able to “pet” cells – growing them
in a fish tank so that we can observe them closely. This
technique is called “Cell Culture”.

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts - What is cell culture?
Cell culture refers to the removal of cells from an
animal or plant and their subsequent growth in a
favorable artificial environment.
General Concepts - What is cell culture?
Cell culture is a technique that involves growing cells in a
laboratory setting, such as in a culture dish or flask. These cell
culture containers are transparent to allow us closely observing
cells under a microscope.

The cells may be removed from the tissue directly and

disaggregated by enzymatic or mechanical means
before cultivation, or they may be derived from a cell
line or cell strain that has already been established.

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts - What is cell culture? General Concepts - What is cell culture?
Ø Scientists can tell whether their
cultured cells are happy or not. For
example, by providing the cells with All of these studies rely on the
different nutrients, researchers can ability to grow human cells in
learn more about the body’s energy the laboratory, which is what
metabolism. we refer to as cell culture.
Ø By introducing unknown chemicals
or toxins to the cells, researchers This technique is critical for
can study toxicology and
understand how certain substances understanding how cells work
affect cell health. and is an essential tool in the
Ø By manipulating the genes of the study of human health and the
cells, researchers can learn more treatment of diseases.
about the functions of specific
genes.

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts - What is cell culture? General Concepts - What is cell culture?
To keep your cells happy, it is important to replicate their natural habitat To keep your cells happy, it is important to replicate their natural habitat
as closely as possible. Scientists have spent decades researching the ideal as closely as possible. Scientists have spent decades researching the ideal
conditions for keeping cells happy in culture dishes. conditions for keeping cells happy in culture dishes.

Table. Optimal cell culture conditions for most mammalian cells.

pH 7.0–7.4

Osmolarity 280–320 mOsmol/kg

CO2 5–10%

Temperature 35–37 oC

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts - What is cell culture?
Cells are grown in a nutrient-rich medium that
provides them with the necessary nutrients, growth
factors, hormones, and gases (O2) to thrive. This
medium also provides a stable physical and chemical
environment with a suitable pH, osmotic pressure,
and temperature.
Scientists try to replicate the conditions of our body
fluids in the cell culture medium, with a neutral pH
(pH 6.8 ~ 7.4), body temperature (37oC), and
isotonic osmotic pressure. In addition, necessary
nutrients such as amino acids, vitamins, inorganic
salts, and a carbon source like glucose are added to
the medium according to the cells’ physiological
needs. These formulations are known as basal
media.
General Concepts - What is cell culture?
Serum is a component of blood that contains many
proteins, hormones, enzymes, and growth factors
that support the cells in our body.
It is common to add a percentage (5-20%) of fetal
bovine serum (FBS) to the cell culture medium as a
supplement. Some cell types may require additional
growth factors, such as neural growth factors (NGF)
for the growth of neural cells.
Cell culture medium is often supplemented with an
antibiotic combination of penicillin and
streptomycin as a measure to prevent bacterial
growth.

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General Concepts - What is cell culture? General Concepts - What is cell culture?
The growth medium controls the pH of the cell culture and helps to Some cells require a solid or semi-solid substrate to form an adherent
buffer the cells against changes in pH. This is typically done by culture as a monolayer. Researchers typically use plastic cell culture
including an organic buffer, like HEPES, or a CO2-bicarbonate containers, such as dishes, flasks, and multi-well plates, to grow these
buffer. The pH of the medium is dependent on the balance of cells. These containers are chosen for different experimental purposes.
dissolved CO2 and bicarbonate, and changes in atmospheric CO2 can
alter the pH of the medium. To prevent this, it is necessary to use
exogenous CO2 when using a CO2-bicarbonate buffer. Most
researchers use a concentration of 5% CO2.

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts - What is cell culture? General Concepts - What is cell culture?

Blood cells can be grown floating in the

culture medium as a suspension culture. Cells are grown in flasks, dishes,
Small-scale suspension cultures can be and plates that are placed inside an
done in cell culture flasks or plates. Large- incubator with strictly controlled
scale suspension cultures are typically done humidity, temperature, and
in special spinner flasks (i.e., stirrer bottles), CO2 concentration. This helps to
which are designed for suspension cell provide the optimal environment for
culture. These flasks allow for better gas cell growth and proliferation.
exchange and can accommodate larger
volumes of cells.

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Cell and Tissue Culture
General Concepts - What is cell culture?
Successful cell culture depends heavily on the practice of an aseptic
technique to keep the cells free from contamination by
microorganisms such as bacteria, fungi, and viruses. The elements of
the aseptic technique include a sterile work area, good personal
hygiene, sterile reagents and media, and sterile handling.
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Cell and Tissue Culture
General Concepts - What is cell culture?
Most laboratories have dedicated cell culture rooms. This is good not only
for preventing the contamination of cell cultures, but also to eliminate
exposure of laboratory workers and the outside environment to potentially
harmful biological agents.
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Cell and Tissue Culture
General Concepts - What is cell culture?
Sterile handling of cell culture is usually done inside a cell culture hood
(also known as laminar flow hood). The hood consists of a work surface
enclosed in a cabinet with a front opening, through which the user can
access the work area. A laminar flow of air is generated within the hood,
which helps to remove contaminants from the air and maintain the sterility of
the work area.
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Cell and Tissue Culture
General Concepts - What is cell culture?
The Centers for Disease Control (CDC) in the United States sets regulations
and recommendations for biosafety in laboratory settings. Cell culture
laboratories can be assigned to one of four biosafety levels based on the
risk associated with handling a particular agent.
Biosafety Level 1 (BSL-1) is the basic level of protection and is appropriate
for agents that do not cause disease in normal, healthy humans.
Biosafety Level 2 (BSL-2) is appropriate for moderate-risk agents that can
cause human disease through ingestion, inhalation, or injection. Most cell
culture labs should be at least BSL-2.
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Cell and Tissue Culture
General Concepts - What is cell culture?
Biosafety Level 3 (BSL-3) is appropriate for agents that have the
potential for aerosol transmission and may cause serious and potentially
lethal infections.
Biosafety Level 4 (BSL-4) is the highest level of containment and is
appropriate for agents that pose a high individual risk of life-threatening
disease through infectious aerosols and for which no treatment is
available. These agents are restricted to high-containment laboratories.
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Cell and Tissue Culture
Source of Cells – Primary Cell v.s. Cell Lines
There are various ways to classify cell culture. One way
to classify cell culture is by the source of the cells. The
cells may be removed from the tissue directly and
disaggregated by enzymatic or mechanical means before
cultivation. Or they may be derived from a cell line or cell
strain that has already been established.
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Cell and Tissue Culture
General Concepts - What is cell culture?
In a BSL-4 cell culture room of Boston University,
scientists are studying a Level-4 pathogen, the
Ebola virus.
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Cell and Tissue Culture
Source of Cells – Primary Cell v.s. Cell Lines
Primary cells are those that are directly extracted from a tissue or organ
and are usually used in a single experiment. They are often preferred in
research studies that require the use of cells that closely resemble those
found in the body. Primary cells are more representative of the original
tissue, leading to more accurate results. However, their lifespan is limited,
as they can only divide a few times (one to two times) in cell culture.
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Source of Cells – Primary Cell v.s. Cell Lines General Concepts - What is cell culture?
Cell lines, on the other hand, are cells cultured in the laboratory for an
extended period of time and have the ability to grow and divide
indefinitely. Cell lines are derived from primary cells and are often used
in research and other applications because they are easy to grow and
maintain in the laboratory.

Normal cells have a limited capacity for proliferation and will eventually
stop dividing, a process known as senescence. However, some cell lines
can become immortal through a process called transformation.
Transformation can occur naturally, as is the case with cancer cells, or it
can be induced by external factors such as chemicals or viruses.

When a finite cell line undergoes transformation and gains the ability to
divide indefinitely, it becomes a continuous cell line. Continuous cell
lines are able to divide and proliferate indefinitely, unlike normal cells
which have a limited capacity for proliferation.

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts - What is cell culture?
Pluripotent stem cells have characteristics of both cell lines and primary
cells. They are called “pluripotent” because they have the potential to give
rise to any cell type. In an undifferentiated status, pluripotent stem cells can
divide and proliferate indefinitely. This makes them similar to cell lines.
General Concepts - What is cell culture?
However, unlike cell lines, which become immortal through transformation,
pluripotent stem cells have the ability to divide indefinitely due to their nature
as stem cells. This means that they also retain the properties of normal
primary cells.

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General Concepts - What is cell culture? General Concepts - What is cell culture?
There are two main types of pluripotent stem cells: embryonic stem cells
(ESCs), which are derived from the inner cell mass of a blastocyst during
early embryonic development, and induced pluripotent stem cells (iPSCs),
which are adult cells that have been genetically reprogrammed to behave
like pluripotent stem cells.

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Cell and Tissue Culture Cell and Tissue Culture

General Concepts - What is cell culture? General Concepts - What is cell culture?

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General Concepts - What is cell culture? General Concepts - What is cell culture?
Bacterial and yeast culture

Insect cell culture Bacteria and yeasts are typically grown in small quantities on a solid
Insect cells can be cultured in a medium containing nutrients, such as agar. For large-scale cultures,
nutrient-rich media, which allows them these cells are grown suspended in a nutrient broth. This method allows
to grow and divide. Insect cells are for the proliferation and study of bacteria and yeasts in a laboratory
relatively easy and inexpensive to setting.
culture compared to mammalian cells,
making them popular for producing Viral culture
proteins, vaccines, and other products
for research or industrial purposes. Viruses cannot live alone. In order to culture viruses, host cells of
mammalian, plant, fungal, or bacterial origin are required. These host
cells provide an environment where the virus can grow and replicate.
Large-scale cultivation of viruses is used to produce vaccines.

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Ece ÖZYAĞCI
Selay TORNACI
Ebru TOKSOY ÖNER
Marmara University, Department of Bioengineering

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