Cell culture

Question 1

What is the medium in cell cultures? What does it contain?

Answer 1

It is a liquid or gel used to help cells grow in culture.

The culture medium should supply all essential nutrients for cell metabolism, growth and proliferation.

– It should have nutrients, water, ions, inorganic salts, growth factors – the basal medium (i.e. the defined component)

——–> a proportion (5-20%) of medium should be serum – which is the plasma after clotting (the undefined component)

– Most basal media are modifications of Eagle’s Minimum Essential Medium (MEM)

Question 2

What is serum? Why is it needed?

Answer 2

Animal cells derived from whole organisms and grown in culture often cannot grow without the addition of hormones or growth factors which usually occur in vivo.

This is provided by adding serum into the culture.

– Generally foetal calf/bovine serum is used, extracted from new-borns and it contains a lot of growth factors.

– Usually serum is tested before it is added to medium to see if it’s appropriate as batches of serum will vary from each other – it’s an undefined component.

– Serum also contains growth factors including IGF, FGF and EGF – can be tailored depending on cell type, e.g. VEGF to promote blood vessel formation.

– Hormones in the serum can also affect cell proliferation.

Question 3

What are serum-free systems? Why have they been developed?

Answer 3

—> People have developed serum-free systems due to the ethical concerns regarding extraction and harvesting of fetal bovine serum. There is also high demand and not much supply.

The technical disadvantages to using serum include its undefined nature, batch-to-batch variability in composition, and the risk of contamination.

Serum free systems allow a defined media, where components concentrations are known.

Finding out what the cell culture needs to obtain from serum showed three components where vital:

– The serum component provides carrier proteins (transferrin, albumin), hormones (insulin), trace elements (selenium) to maintain viability of cells. Insulin/transferrin/selenium (ITS) is essential in defined media.

Reduced serum culture systems are also used.

Question 4

Describe the cell-ECM interactions.

Answer 4

The matrix/ECM functions to –>

• Provide an anchor for cells for mechanical support
• Act as a substrate for cells to migrate through.
• Its composition can also have big effect on cells, as cells can detect composition of the environment.
• The ECM can act as a physical barrier to influence cell shape/orientation; it is also deformable where cells will deform in shape along with it.

Cells can also contract and physically deform their environment.

A lot has been learnt about cell ECM interactions from culture such as that cells need to be attached to ECM in order to reach their full function.

• Cells attach to matrix, stretch and spread out and acquire motility from acquired polarity ends – basal and apical, which is controlled by signals or random if no signals are present.
• Remodelling of cytoskeleton and energy is required for cell spreading – it isn’t a passive process, as energy is needed to modulate cell adhesion during spreading
• The equivalent of “spreading” and migrating for neurones is the formation of growth cones which are the tips of extending neurites.​