Viability Assays

Question 1

What is the solution we grow cells in called?

Answer 1

Growth medium

Question 2

What does growth medium contain?

Answer 2

amino acids, vitamins, salts & sugars

Question 3

How do we supplement protein into cell culture?

Answer 3

• using fetal calf serum

Question 3

Describe Fetal Calf Serum

Answer 3

• fraction purified from blood of bovine calves
• non definded mixture of proteins, growth factors & cytokines

Question 4

What flasks are typically used in cell culture?

Answer 4

T25s & T75s

Question 5

What is the base area of a T25 flask?

Answer 5

25cm ^2

Question 6

What is the base area of a T75 flask?

Answer 6

75cm^2

Question 7

What does anchorage dependant mean?

Answer 7

• many human cells are anchorage dependant, meaning they need to stick to the base of the flask to grow

Question 8

How do cells adhere to the flask?

Answer 8

• cells secrete ECM

Question 9

How do we remove adhered cells from flask walls?

Answer 9

• need to destroy the ECM without damaging the cells
• this is carried out using Trypsin

Question 10

What is the biggest risk to tissue cultures?

Answer 10

Microbial infection

Question 11

What is included to help alleviate infection in growth culture?

Answer 11

penicillin & streptomyocin

Question 12

Define Viability

Answer 12

• capable of living
• capable of growing/developing

Question 13

Describe Trypan Blue

Answer 13

• diazo dye -> its a dye exclusion method
• routine test to determine percentage viability of a populations of cells
• viable = alive
• percentage viability tells us what proportion of cells are dead & alive

Question 14

How does Trypan Blue enable us to calculate viability?

Answer 14

• cells which have taken up the dye are blue & we know are dead
• not taken up dye = colourless & are alive

Question 15

Describe the principle of Dye Exclusion

Answer 15

• principle that intact membrane of a living cell will exclude a given substance (dye)
• when cells dye the membrane becomes compromised & substances that were once excluded can now traverse the membrane

Question 16

Describe a Haemocytometer

Answer 16

• modified microscope slide with a counting grid etched into it
• liquid dispensed onto gird using a pipette & seal with coverslip

Question 17

What is the equation for percentage viability ?

Answer 17

(live cells/ total cell number ) x 100