Chapter 4 - B cell (Exam 3) Flashcards
Functions of antibodies
- Recognize and bind antigens
- Target bound antigen to other cells
Number of Ab molecules one can produce is limited by
Number of B cells
Before antigen binding
Ig is bound to B cell surface receptor
After antigen binding
B cell proliferates and differentiates into plasma cell
Secretes antibody
Different antibody structures
Light + Heavy
Variable + Constant
Fab fragment
Fragment antigen binding
Fc fragment
Fragment crystallizable
Proteolytic cleavage
Two Fab and one Fc
5 isotypes of heavy chains
IgG, IgM, IgD, IgA, IgE
Secreted as monomers?
IgG, IgD, and IgE
Secreted as monomer and dimer?
IgA
Secreted as pentamer?
IgM
What are the two isotypes of light chains?
kappa and lambda
each antibody has EITHER one
H and L chains arranged in
immunoglobulin domains
Differences in ____ lead to variability and different binding properties in light chains.
Loops
Globular
Soluble in aqueous environment
-hydrophilic aa out
-hydrophobic aa in
Immunoglobulin-like domain
present in other proteins as well
immunoglobulin superfamily
Hypervariable regions (HV)
major differences in aa sequences reside
Complementary determining regions (CDRs)
Framework regions (FR)
low variability in amino acid sequence
Types of antigen binding sites
Pocket
Shallow groove
Extended surface
Protrusion
Antigen binding sites
Non-covalent forces bind chemical groups in antigen to amino acids of Ab
Non-covalent forces
- electrostatic interactions
- hydrogen bonds
- van der waals forces
- hydrophobic interactions
what do antibodies recognize?
proteins, carbohydrates, glycoproteins, proteoglycans, glycolipids, etc
antibodies recognize _______________ on he outer surface
native structure
B cell differentiates into a ___________ which secretes ____________
plasma cell
antibody
epitope
antigenic determinant
part of antigen to which antibody binds
cluster of amino acids
antibodies are most effective at binding
surface epitopes
multivalent antigen
more than 1 epitope
antibody epitopes can be
linear or discontinuous
polyclonal antibody
different antibodies recognize different epitopes
monoclonal antibody
same antigens, same epitope
way to produce antibodies in research
immunize an animal, collect antisera from blood (pure) and produce polyclonal antibodies
monoclonal antibodies in research
B cells from mouse and tumor cells fused together
select antigen specific hybdridoma and grow in large numbers
uses of antibodies in research
identify many unknown surface proteins
monitor specific surface protein expression
flow cytometry
use of fluorescent antibodies to measure cell population in blood
detect diseases
antibodies in the clinic are used to _______
how?
treat diseases
inject mouse antibodies into deficient patients, prevent transplant rejection
modifications to mouse monoclonal antibodies to prevent human immune system attack
chimeric
humanized
human
chimeric modifications
drug example
mouse V regions, human C region
rituximab - lymphoma
humanized modifications
drug example
mouse CDRs, all other sequences human
omalizumab - allergic asthma
human modifications
drug example
completely human antibody sequence
antibody from mice to have human Ig genes
adalimumab (HUMIRA)
another name for human modifications is
human hybridomas
ELISA
enzyme linked immunosorbent assay
measures concentration of an antibody/antigen that can bind to an antibody
quantification of ELISA
absolute concentration or antibody titer
RIA
radioimmunoassay
measures the concentration of an antibody/antigen that can bind an antibody
coombs test
tests for presence of antibodies and complement that can bind to RBCs
results in red blood lysis/anemia
western blot
detects presence of a single protein in a complex mixture of proteins
immune system needs to generate a _______ population of antibodies that can recognize any pathogen at infection site and target it for destruction
diverse
what happens before antigen binding?
generate billions of B cells
containing a surface immunoglobulin with a unique variable region
what happens after antigen binding?
secrete antibody
modify variable region to bind more tightly
switch heavy chain
why do we switch heavy chain after antigen binding?
to eliminate the pathogen most effectively
things that happen before antigen binding
somatic recombination
junctional diversity
combination of different heavy chains and light chains
antibodies cannot be _________ until they are __________
expressed
rearranged
light chain variable region contains
variable (V) and joining (J) segments
heavy chain variable region contains
variable (V), joining (J) and diversity (D) segments
rearranged V, D and J segments encode for
CDR loops of antibody
do constant regions have to be rearranged?
NO BITCH!
which constant region determines the isotope of the antibody?
the closest one
somatic recombination
random rearranging of antibodies
how many rearrangements in light chains?
1 (V-J joining)
how many rearrangements in heavy chains?
2 (DJ, V-DJ joining)
enzyme responsible for the recombination of genes
VDJ recombinase
examples of recombination activating genes
RAG-1 and RAG-2
what is involved in the recombination of genes?
VDJ recombinase
recombination activation genes
DNA repair enzymes
junctional diversity
2 sets of new nucleotides are randomly added (P and N)
Severe Combined Immunodeficiency (SCID)
genetic deficiencies RAG enzymes
B and T cells are not produced
3 contributions to antibody diversity
antibody DNA gene segments are randomly combined
additional nucleotides are inserted at junctions during somatic recombination
separate B cells have the same rearranged heavy chains with different rearranged light chains
allelic exclusion
only 1 allele for each is rearranged and expressed in a single B cell
allelic exclusion ensures that each B cell produces
antibody of a single antigen specificity
allelic exclusion allows for
clonal selection against only 1 pathogen
things that happen after antigen binding
surface Ig vs Secreted Ab
somatic hypermutation
isotope switching
first form of Ig is
membrane bound
membrane bound Ig
transmits signal of antigen binding to Ig into cell interior with the help of Iga and Igb coreceptors
after antigen binding, Ig is
secreted as antibody (plasma B cell)
somatic hypermutation
random introduction of single nucleotide mutations through entire rearranged V region DNA
somatic hypermutation is performed by the enzyme
AID (activation induced cytidine deaminase)
AID causes a __________ and is made only by _____________
point mutation
proliferating B cells
result of somatic hypermutation
B cells with mutant surface Ig
somatic hypermutation leads to
selection of antibodies with higher affinity for infection pathogen
isotope switching
splice out C region of previous isotope
what antibody is made in the primary immune response?
IgM
why does the isotope need to switch from IgM?
so the antibodies can eliminate the pathogen properly
isotope switching depends on the
AID enzyme
switch region
highly repetitive DNA
facilitates recombination
switching can occur
from m to any other isotype or sequentially
Hyper IgM immunodeficiency
no somatic hypermutation/class switching
only low affinity IgM in large amounts
antibody effector functions
neutralization cells
opsonization
complement activation
recruiting NK cells to kill
sensitization of mast cells
what determines antibody effector functions?
isotype!
IgM
pentameric
low affinity, high avidity
avidity
overall strength of multiple binding sites
IgG
smaller, more flexible
transferred across placenta during pregnancy
IgA
secreted as monomer in blood, dimer in mucosal surfaces
in lymphoid tissue under mucosa
in lumen of gut and in milk, saliva, sweat, tears
what is the most abundantly produced Ig?
IgA
IgE
binds to mast cells, eosinophils, basophils
eradicates parasites/response to allergen
IgG antibody has the most _______ functions
why?
diverse
hinge, elbow and tail regions give IgG flexibility
IgG has ______ different subclasses and they differ in _________________
four (G1-G4)
structure of the hinge
IgG4 can exchange
1 heavy and light chain in circulation (functionally monovalent)
because IgG4 is functionally monovalent, it
can reduce immune response
is elevated in allergic individuals