Generation of Antibodies

Question 1

Describe monoclonal antibody production

Answer 1

-homogenous antibody production
-All antibodies possess the same amino acid sequence
All antibodies are of the same isotype
Originated from one mouse B cell

Question 2

What are the characteristics of monoclonal antibodies?

Answer 2

High specificity, one epitope recognised
Variety in nature, all molecules identical
Unlimited supply

Question 3

Overview of hybridoma production

Answer 3

Myeloma + B lymphocyte—>PEG—>hybridoma

Question 4

which myeloma cells should you use?

Answer 4

ones that have an enzyme deficiency necessary

Question 5

What selection medium is used?

Answer 5

HAT- Hypoanthine aminoperterin thymidine
Aminoperterin blocks DNA synthesis
pathways

Question 6

How do you assess hybridomas?

Answer 6

Identity fusions= hybridoma, they grow
Grow cells assess the supernatant
Expand desired clones
Make stocks of hybridomas to allow unlimited supply

Question 7

What are the considerations when assessing a hybridoma?

Answer 7

Antibodies are usually low
Assays to assess antibodies need to be:
-Reliable
-Fast
-Sensitive

Question 8

Screening assays for mAbs?

Answer 8

ELISA

Western blot

Question 9

What are the advantages to the PAb?

Answer 9

Low technology, easy to make, quick, low skill set required

Question 10

What are the advantages to the MAb?

Answer 10

Single specificity
Unlimited supply
Use as therapeutic agent