Lecture 3

Question 1

What are the 4 basic things that must be controlled to grow cells in culture?

Answer 1

O2, temp, humidity, pH.

Question 2

what can maintain the environment for cells?

Answer 2

Incubator.

Question 3

what is the optimum pH for mammalian cells

Answer 3

7-7.4

Question 4

Under what pH is mammalian cell growth inhibited?

Answer 4

6.8

Question 5

What can regulate the pH of a medium?

Answer 5

Buffering systems, e.g. sodium bicarbonate-CO2. (bicarbonate added to medium, CO2 pumped in from gas tank when needed).

Question 6

What is osmolality?

Answer 6

Concentration of a solute in a solution measured by mass.

Question 7

Does osmolality affect cell growth and function?

Answer 7

Yes. Can be determined by components of growth medium.

Question 8

What is Mr Frosty filled with before cryovials full of substrate are added?

Answer 8

Isopropanol.

Question 9

Does the pH of the medium increase of decrease over time with mammalian cells? why?

Answer 9

Decrease, because of metabolic products, e.g. lactic acid.

Question 10

What indicator can be used to determine pH of growth medium? What is the colour change on becoming too acidic?

Answer 10

phenol red.

Turns yellow.

Question 11

Are established cells often aneuploid?

Answer 11

Yes.

Question 12

Do established cell lines have normal doubling times?

Answer 12

No, often shortened.

Question 13

What are 2 main ingredients in medium?

Answer 13

Glucose and glutamine.

Question 14

Does CO2 increase or lower pH?

Answer 14

Lowers.

Question 15

What is transformation?

Answer 15

Genetic modification allowing long term propagation in culture, causing a phenotypic modification.

Question 16

What 4 does the extracellular matrix do?

Answer 16

Support, segregate, aid communication, act as a depot.

Question 17

Does the matrix change over time? When? What enables this?

Answer 17

Yes, during growth and development (by use of proteinases and matrix metalloproteinases), and wound healing (fibroblasts).

Question 18

Components of ECM?

Answer 18

Proteoglycans (e.g. chondroitin).
Polysaccharides (e.g. hyularonic acid).
Fibres (e.g. collagen, elastin).
Fibronectin, laminin.

Question 19

Types of cell adhesion

Answer 19

Focal adhesion (ECM interacting w/ actin).
hemidesmosome-ECM interactions.
integrin.

Question 20

Do cells bind to each other, or to filaments?

Answer 20

Both.

Question 21

Do fibroblasts interact with the matrix?

Answer 21

Yes.

Question 22

what happens to signals from the extracellular matrix when they bind to a receptor on a cell?

Answer 22

Signal transduction pathways -> cytoskeleton -> EITHER

Organisation and movement of molecules.
OR
Cell shape movement and contraction.

Question 23

2 reasons to do adhesion assays?

Answer 23

Understand how tissue structures form.

Understand how tumours bind during tumour metastasis.

Question 24

Basic mechanism of adhesion assay?

Answer 24

Coat microplates with matrix of choice.
Cells stained with colorimetric/fluorescent dye, usually calcein AM.
Cells added to micro plates, left, then washed to remove cells that have not adhered.
Fluorescence/colour then measured.

Question 25

What matrices are usually used in adhesion assays?

Answer 25

Fibronectin, collagen, gelatin, poly-t-lysine.

Question 26

What is a fungal toxin that binds F-actin?

Answer 26

Phalloidin.