Cell Culture

Question 0

What does recombinant DNA technology make?

Answer 0

Enzymes, synthetic hormones,monoclonal antibodies, anticancer agents

Question 1

What do biotechnology produce?

Answer 1

Manufacture of viral vaccines

Question 2

What is transfection?

Answer 2

Introduction of foreign DNA
Express a protein of interest
RNAi constructs: suppress expression of a gene/protein

Question 3

What is cell culture?

Answer 3

Removal of cells from animal /plant to be grown in a favourable artificial environment

Question 4

What is primary culture?

Answer 4

Cells isolated from tissue & proliferated until they occupy all available substrate

Question 5

What is a cell line?

Answer 5

1st subculture (subclone)

Question 6

What is a cell strain?

Answer 6

Select a subpopulation (e.g. Cloning)

Question 7

What is a finite cell line?

Answer 7

Normally cells only divide a limited number of times, then stop proliferating (senescence)

Question 8

What is a continuous cell line?

Answer 8

Virally induced transformation to become “immortal”

Question 9

What is the media for cell culture?

Answer 9

Dulbeccos modified eagle medium

Question 10

What basic equipment is needed?

Answer 10

Cell culture good
Incubator
Water bath
Centrifuge
Refrigerator and freezer (-20oC)
Automated cell counter 
Inverted microscope
Liquid nitrogen for storage
Sterilizer (autoclave)

Question 11

What is used to make benches aseptic?

Answer 11

70% ethanol spraying

Question 12

What do the hood do?

Answer 12

Protect working environment from dust & airborne contaminants

Question 13

What is immortalized cell line?

Answer 13

Acquired ability to proliferate indefinitely

Question 14

What does the medium contain?

Answer 14

Essential nutrients: amino acids, carbohydrates, vitamins,minerals
Growth factors & hormones
Gases: oxygen & carbon dioxide
Right pH, osmotic pressure & temperature

Question 15

How can you tell if bacteria is been contaminated into your culture?

Answer 15

Culture cloudy (turbid) with thin film on surface. Drop in pH, tiny moving granules among cells

Question 16

How can you tell if your culture is contaminated with yeast?

Answer 16

Turbid, pH rises when heavy, ovoid/spherical particles

Question 17

How can you tell if contained with molds?

Answer 17

Hyphae: connected filaments

Question 18

How can tell if viruses in it?

Answer 18

v. difficult to detect in culture

Question 19

How are infections prevented in the cell culture?

Answer 19

Antibodies or antifungals are added to media

Question 20

How do you prevent cryopreservation?

Answer 20

Treat with protective agent (DMSO or glycerol)

Stored at -130oC: liquid nitrogen

Question 21

Name some cell types?

Answer 21

Fibroblasts
Epithelial
Lymphoblast

Question 22

What is plating density?

Answer 22

No. cells per volume of culture medium
Suspension
Adherent: extracellular matrix (collagen/laminin)

Question 23

What is confluence?

Answer 23

No. Of adherent cells in culture dish

Question 24

What are some technical considerations?

Answer 24

Nutrient depletion/changes in pH in the media
Accumulation of dead cells
Contact inhibition m: cell to cell contact stimulates cell arrest
Cellular differentiation stimulated by cell to cell contact
Adherent:extracellular matrix

Question 25

What is passinging?

Answer 25

Transferring a small number of cells into a new vessel

Question 26

How do you avoid technical considerations?

Answer 26

Passaging of cells

Question 27

What is trypsin-EDTS used for ?

Answer 27

Passinging of adherent cells