In hybridoma production, B cells are fused with myeloma cells that lack the enzyme HGPRT, and the mixture is grown in HAT medium. Why do only hybrid cells survive?
AHAT medium kills all B cells immediately
BUnfused myeloma cells cannot make nucleotides when aminopterin blocks the main pathway because they lack HGPRT, and unfused B cells die naturally in culture; hybrids get HGPRT from the B cell and immortality from the myeloma
CHybrids are larger and are separated by filtering
DMyeloma cells produce the desired antibody on their own
Aminopterin blocks de novo nucleotide synthesis. Cells can survive only by the salvage pathway, which needs HGPRT. Myeloma cells lack it and die. Normal B cells have it but stop dividing and die within days. Only fused cells have both the enzyme and the ability to divide indefinitely.
Question 2 True / False
A monoclonal antibody preparation contains antibodies that recognize many different epitopes on the same protein.
TTrue
FFalse
Answer: False
All antibodies in a monoclonal preparation come from one clone and have the same antigen-binding site, so they bind one epitope. Mixtures that recognize many epitopes are polyclonal.
Question 3 Short Answer
Why are therapeutic antibodies such as trastuzumab humanized rather than given as pure mouse antibodies?
Think about your answer, then reveal below.
Model answer: Patients' immune systems recognize mouse antibody regions as foreign and make anti-mouse antibodies, which clear the drug faster and can cause allergic reactions. Humanizing replaces most mouse sequence with human sequence while keeping the parts that bind the target, so the drug is less immunogenic.
Under the naming convention used for most approved antibodies, -ximab marked chimeric antibodies (mouse variable regions, human constant regions), -zumab humanized antibodies and -umab fully human ones. Rituximab, trastuzumab and adalimumab are examples of each.
Question 4 Multiple Choice
In a sandwich ELISA, what is the role of the second (detection) antibody?
AIt coats the plate and captures the antigen
BIt binds a different epitope on the captured antigen and carries an enzyme that produces a measurable color
CIt destroys unbound antigen
DIt blocks the enzyme reaction
The capture antibody on the plate binds the antigen. The detection antibody, linked to an enzyme, binds a second epitope. After washing, a substrate is added, and the enzyme converts it into a colored product. More antigen means more bound detection antibody and more color.