CMB: Cell Culture, Cell Lines, and Stem Cells

What is cell culture?

Cells, previously growing in a human, animal, or plant, are modified to grow on plastic or glass under controlled laboratory conditions.

What is the difference between in vivo and in vitro?

In vivo = inside a living organism. In vitro = outside the organism, such as cells growing on plastic or glass in the laboratory.

Why is cell culture needed?

Cultured cells are model systems for studying basic cell biology, interactions between disease-causing agents and cells, drug effects, aging, nutrition, toxicity, cancer, viruses, and genetic engineering.

What are the major benefits of cell culture in research?

It allows study of cell behavior individually, control of the growth environment, reproducible experiments, exposure to defined concentrations of drugs or chemicals, and reduction of legal, moral, and ethical problems associated with animal experimentation.

What is a clone?

A population of cells derived from a single cell; the cells are genetically identical.

What is a sub-culture?

The transplantation or transfer of cells from one culture vessel to another.

What is a passage number?

The number of successive sub-cultures from the original primary culture.

What is a primary cell culture?

A culture derived directly from animal tissue. It has a finite lifespan, generally retains differentiated characteristics, is mainly anchorage-dependent, and exhibits contact inhibition.

What is an established, stable, or continuous cell line?

A primary culture that has become immortal due to transformation and can be continuously propagated in culture. Chinese Hamster Ovary (CHO) cells are commonly used.

What is the Hayflick limit?

After about 40–60 cell divisions, telomeres become critically short, triggering stress signals. p53 and Rb pathways can then cause cell-cycle arrest, senescence, or apoptosis.

What happens when both p53 and Rb pathways are inhibited?

The normal controls on cell proliferation are lost, allowing continuous proliferation and contributing to formation of an immortalized cell line.

What do cells need to grow in culture?

An adhesion surface, liquid culture medium, appropriate environment including CO₂, temperature around 37°C, humidity, appropriate oxygen tension, and sterility.

How does 5% CO₂ help maintain cell-culture pH?

Cell-culture media contain sodium bicarbonate (NaHCO₃). CO₂ supplied by the incubator works with bicarbonate as a reversible buffering system to maintain pH around 7.4.

What is the purpose of basal culture media?

To maintain pH and osmolarity and provide nutrients and an energy source. It contains glucose, proteins, essential salts, minerals, and often the pH indicator phenol red.

What does phenol red indicate in cell culture media?

It is a pH indicator. Around pH 7.2 the medium appears pink/red. Acidic conditions produce yellow/orange, while basic conditions produce purple.

What is the purpose of serum in cell culture?

Serum provides nutrients and growth-promoting components, including amino acids, vitamins, lipids, proteins, and growth factors.

What are PBS and trypsin-EDTA used for?

PBS is used to wash away excess serum. Trypsin-EDTA is used to detach cells from a culture dish. EDTA binds calcium ions that would normally inhibit trypsin.

What are the three major types of stem cells based on their ability to differentiate?

Totipotent, pluripotent, and multipotent. Totipotent cells can develop into a new individual; pluripotent cells can form over 200 body cell types; multipotent cells can form a specific range of tissue types.

Why are hematopoietic stem cells considered multipotent?

They can generate different blood-cell types, including red blood cells, white blood cells, and platelets, but cannot normally generate unrelated cell types such as neurons, hepatocytes, or muscle cells.

What is a hybridoma and why is it important?

A hybridoma is a hybrid cell formed by combining an antibody-producing cell with a cell capable of continued proliferation. Hybridomas are important for large-scale production of monoclonal antibodies used in diagnosis and therapeutics.