6. Antibodies and biotechnology

Question 1

difference between monoclonal and polyclonal Ab?

Answer 1

monoclonal - Ab produced which can only recognise 1 type of Ab
polyclonal - many Ab produced - many epitopes recognised

Question 2

what cells allow proliferation of cells?

Answer 2

tumour/myeloma cells

Question 3

Advantage of polyclonal Ab

Answer 3

good precipitation and agglutination

Question 4

advantages of monoclonal Ab - 2

Answer 4

1. pure sample - specific binding

2. no batch variation

Question 5

disadvantages of polyclonal Ab - 3

Answer 5

1. not pure = non-specific binding
2. limited serum supply = batch variation
3. ethics - animals

Question 6

disadvantages of monoclonal Ab -2

Answer 6

1. POOR agglutination and precipitation

2. costly - labour intensive

Question 7

4 types of blotting techniques - what do each of them detect?

Answer 7

western = proteins 
eastern = carbs/lipids 
Southern = DNA
northern = RNA

Question 8

what 3 things does western blotting tell us?

Answer 8

1. presence/absence of protein
2. drug effect
3. subcellular distribution of proteins, glycosylation and size

Question 9

method used for extracting pure sample of epitope

Answer 9

immunoaffinity production

Question 10

2 types of immunoaffinity production

Answer 10

1. affinity chromatography

2. immunoprecipitation

Question 11

3 commonly used proteins for purification in immunoprecipitation

Answer 11

1. 6 x histidine
2. Gluthione-S-transferase
3. Maltose binding protein

Question 12

what does immunohistochemistry detect?

Answer 12

distribution of antigens in tissue

Question 13

what does immunocytochemistry detect?

Answer 13

specific ag in whole cells

Question 14

in IHC - what is used to preserve the cells?

Answer 14

paraffin wax

Question 15

2 uses of ICC in clinical practice?

Answer 15

a) cancer diagnosis

b) change in disease with drug

Question 16

what does ELISA stand for?

Answer 16

enzyme-linked immunosorbent assay

Question 17

4 types of ELISA

Answer 17

1. direct
2. indirect
3. sandwich
4. competitive

Question 18

purpose of competitive elisa?

Answer 18

test Ag concentration

Question 19

which ELISA start with an Ab coated well?

Answer 19

indirect

Question 20

which ELISA only uses 1 AB?

Answer 20

direct

Question 21

how to detect outcome in ELISA?

Answer 21

add substrate which will react with enzyme on conjugated Ab - changing colour of solution

Question 22

what method is used in diagnosing blood disorders?

Answer 22

flow cytometry

Question 23

FACS

Answer 23

fluorescence-activated cell sorting

Question 24

in FACS - what does forward scatter determine?

Answer 24

size and volume of cell

Question 25

in FACS - what does side scatter determine?

Answer 25

granularity, nucleus structure, vesicles inside of cell

Question 26

which cells have large forward and side scatter?

Answer 26

monocytes and granulocytes

Question 27

3 problems of lab methods

Answer 27

1. non-specific binding 2, tissues are auto-fluorescent 3. Ab need to be optimised

Question 28

solution to non-specific binding

Answer 28

use positive and negative controls

Question 29

solution to auto-fluorescent tissues

Answer 29

enzymatic markers

Question 30

solution to Ab optimisation

Answer 30

alter conc to allow max binding

Question 31

2 enzyme markers

Answer 31

1. horse-radish peroxidase | 2. alkaline phosphatase

Question 32

2 fluorochromes

Answer 32

1. fluorescein isiothiocyanate | 2, Alexa 488

Question 33

how can Ab be altered so that they don't cause an autoimmune response in humans?

Answer 33

changing constant region of Ab to human

Question 34

name 8 diseases Ab can be used as target therapy

Answer 34

1. cancer 2. transplant rejection 3. crohn's disease 4. multiple sclerosis 5. inflammatory lesions 6. bacterial infections 7. sepsis 8. viral infections

Question 35

example of Ab treatment used to combat HER2 on cancer cells

Answer 35

Herceptin

Question 36

2 mechanisms of Herceptin

Answer 36

1. binds to EGF on tumour cell - stops proliferation | 2. binds to Fc domain - NK cell apoptosis

Question 37

4 ways Ab can be used against cancer

Answer 37

1. naked 2. attach to toxin 3. attach radioactive substance 4. attach enzyme which activates drug

Question 38

hormone which is elevated in pregnancy?

Answer 38

hCG