Immunology Chapter 7 Flashcards
(40 cards)
What is polyclonal antisera
mixture of antibodies purified from a serum
occurs when an animal is injected with an antigen and antibodies can be isolated from a serum
what is a serum
liquid portion of the blood (plasma) that doesn’t have platelets but have antibodies
the serum contains antibodies that recognizes….
different epitopes
antigens have multiple different
epitopes
what do these epitopes activate
B cells
what does the B cell do after it is activated by an epitope
It proliferates and differentiates into plasma cells
what do these plasma cells do
secrete antibodies for that epitope
What are the limitations of polyclonal antisera (3)
not standardized (different for every batch)
limited supply (bcs u get it from an animal)
not useful when u need one specific epitope
what is a better option if you want to isolate one specific epitope
monoclonal antiserum
What are the benefits of polyclonal antisera
it has a mix of antibody specificity
so if you want protein Y, protein X’s antibodies may bind to Y because they are similar
What are monoclonal antibodies
antibodies produced by the descendents of 1 clone of B cells
what is the result of monoclonal antibodies (experiment) (after selection)
hybridoma and all of its descendents will make the same antibody as the original B cell so the antibodies will bind to the same epitope because 1 B cell makes an antibody of 1 specificity
Why are hybridomas needed (compare it to B cells)
B cells live for a few days in the culture
But hybridomas live and divide forever and continues making antibodies
B cells + Myeloma cells =
Hybridoma cells
Differentiate the characteristics of all three: B cells, Myeloma cells and Hybridoma cells in terms of
Life span, antibody secretion and drug tolerance
B cells
- mortal
- secrete antibodies
- drug resistance
Myeloma cells
- immortal = plasma cell tumor
- don’t secrete antibodies
- drug sensitive
Hybridoma cells
- immortal
- secrete antibodies
- drug resistance
What is the raw result of monoclonal experiment (like before sorting)
mixture of hybridomas making antibodies of different epitopes
What if I want to get a desired antibody from monoclonal antibodies, what should be done? (selection)
place 1 cell per well in a 96 well place so each one can grow into a clone
use ELISA to test which antibodies bind to the target epitope
What is ELISA, and what are the two versions
an assay that measures proteins, hormones and immunoglobulins in a solution
version 1: detect antibodies
version 2: detect cytokine/hormone –> antigen
Why can’t ELISA be used to detect cells?
because the machine can’t tell the difference between light that’s absorbed vs light that’s scattered
In antibody-detection ELISA, what is the first step?
coat ELISA plate with the target antigen
What is the use of washing away the plate
so the unbound antibodies can go away we only need the bounded ones (bound to the well)
What binds to the antigen in the well during antibody detection ELISA?
primary antibodies
What is the secondary antibody?
What is it’s purpose?
What region does it bind to?
mouse IgG produced by hybridomas
to see which wells have antibodies bound to antigens and is linked to an enzyme for detection
constant region (Fc) of mouse IgG (primary antibody)
How does ELISA produce a visible result?
when you add a colorful substrate and there is color present meaning that there is an antibody
