Antibodies are a class of glycoproteins called
immunoglobulins
Antibodies are produced by
plasma cells (stimulated B cells)
IgM
First antibody produced in blood and lymph in a primary antibody responce
8% Ig pool
IgG
makes up the vast majority of antibody in blood and lymph
Principal antibody in secondary antibody response against the antigen
80% of Ig pool
IgA
found in body cavities where it binds bacteria and viruses before they can infect tissue.
12% of Ig pool
IgE
important in allergic reactions. Trace amount in serum
IgD
normally bound to B cells as a cell surface receptor. Trace amounts in serum
Plasma cells secrete antibody of the same antigen specificity as the membrane bound immunoglobulin expressed by
b-1 precursor
IgM
is a pentameric antibody
IgA
is a dimeric antibody
IgG, IgD, IgE
monomeric antibodies
Antibodies are composed of
polypeptides with variable and constant region
Epitope recognition requires antibodies to have special structure of
2 identical heavy chains
2 identical light chains
Each light and heavy chains has
constant region
variable region
antigen binding site
Constant regions
determines the location and functional class of antibody
Variable region
which contains different amino acids for the many antibodies produced
An antigen- binding site is formed from
the hypervariable regions of a heavy chain V domain and a light chain V domain
The variability allows formation of the specific antigen binding site
Immunoglobulin chains are folded into
compact and stable protein domains
Treatment of IgG with enzyme papain results in
proteolytic cleavage of the hinge of each heavy chain
reduction of the disulfide bonds that connects the two hinges
The flexible hinge of the IgG molecule allows it to bind with both arms to many
different arrangements of antigens on the surface of pathogens
The antigen-binding site of an immunoglobulin is formed from?
Paired V regions of a single heavy chain and a single light chain
Antigen-binding sites vary
in shape and physical properties
Epitopes can bind to
pockets, grooves, extended surfaces, or knobs in antigen-binding sites.
A liner epitope of a protein antigen is
formed from contiguous amino acids
A discontinuous epitope is formed from
amino acids from different parts of the polypeptide that are brought together when the chain folds
Monoclonal antibodies produced from
a clone of an antibody-producing
Rituximab used for
non-hodgkin b cell lymphoma
Adalimumab for rheumatoid arthritis
anti TNF alpha
Production of a mouse monoclonal antibody
Lymphocytes from a mouse immunized with the antigen are fused with myeloma cells by using polyethyleneglycol.
The cells are then grown in the presence of drugs that kill myeloma cells but permit the growth of hybridoma cells. unfused lymphocytes also die.
Individual cultures of hybridoma are tested to determine whether they make the desired antibody. The cells are then cloned to produce a homogeneous culture of cells making a monoclonal antibody.
Myelomas are tumors of
plasma cells; those used to make hybridomas were selected not to express heavy and light chains
Hybridomas only express
the antibody made by the b-cell partner
Generation of immunoglobulin diversity in B cells before encounter with antigen
Generation of immunoglobulin diversity in B cells before encounter with antigen
The DNA sequence encoding a V region is assembled from two or three gene segments
...
The two types of gene segment that encode the light chain V region are called
variable- V
joining -J
segments
Two light-chain loci version
kappa or lambda only one is used
Each kappa or lambda has two types of gene segment (V,J)
The heavy chain locus includes an additional set of
diversity (D) gene segment for a total of three segments
Random somatic recombination of gene segment produces
diversity in the antigen-binding sites of immunoglobulins.
enzyme responsible for recombining V,D,J
V(D) J recombinase
Gene segments encoding the variable region are joined by recombination signal sequences recognized by the
RAG complex
Developing and naive B cells use
alternate mRNA splicing to make both IgM and IgD
The isotype of an antibody is determined
by its heavy chain
The only heavy chains made by mature B cells before they encounter antigen are
mu μ and delta δ corresponding to IgM and IgD, respectively,on B cell surfaces
Simultaneous expression by both forms from the same heavy chain locus is accomplished by
differential splicing of the same primary RNA transcript (no gene rearrangement)
Each B cell produces immunoglobulin of
a single antigen specificity
Allelic exclusion
in developing b-cells ensures that only one heavy chain and one light chain are expressed, which results in B cells producing antibodies of a single antigen specificity.
B cells are
monospecific
monospecific
an encounter with a given pathogen engages a subset of B cells that will make antibodies of a single antigen specificity
----- this is clonal selection. Focus of B cell/ antibody response to a specific antigen
Immunoglobulin is first made in a
membrane- bound form that is present on the B cell surface
When B cells first make IgM and IgD, they are
associated with cell membrane. they need anchors to stick to the plasma membrane
Igβ and Igα invariant chains are trans membrane
proteins that anchor the antibody heavy chain, constant region, to the plasma membrane
Diversification of antibodies after B cells
encounter antigen
Secreted antibodies are produced by alternative pattern of heavy chain
RNA processing
Gene rearrangement with immature B cells leads to the expression of functional heavy and light chain and to the
production of membrane-bound IgM and IgD on the mature B cells
After an encounter with antigen,
the secreted antibodies are produced by the B-cell ( now known as the plasma cell)
HOW? ^^^
Alternative RNA splicing (no gene rearrangement going on here) The membrane bound has a hydrophobic anchor sequence at the end of the heavy chain, where the secreted one has a hydrophilic one.
The surface and secreted forms of an immunoglobulin are derived from the same
heavy-chain gene by alternative RNA processing
Rearranged V-region sequences are further diversified by
somatic hypermutation
Once a B cell has been activated by antigen
further diversification of the whole V-domain coding sequences occurs thru somatic hyper mutation
There is random point mutation at
a very high rate throughout the V-region of the heavy chain and light chain genes
Constant regions are not affected
Somatic hypermutation results in B cells bearing mutant antibodies at the
variable region
somatic hypermutation is dependent on the enzyme
activation-induced cytidine deaminase (AID)
also converts cytosine to uracil normally
Some of these mutant antibodies will bind antigen
better (higher affinity)
B cells containing these mutant receptors will compete for the antigen and are
preferentially selected to mature in to plasma cells
The almost random variation produced by somatic hypermutation allows
selection of variant immunoglobulins with improved antigen-binding sites.
Somatic hypermutation targets the rearranged gene segments encoding the
variable region
IgM and igD are coexpressed on naive cells by a process called
aleternative mRNA splicing
Isotype switching produces immunoglobulins with different
C regions but identical antigen specificities
Like somatic hypermutation isotype switching is dependent on the enzyme
activation-induced cytidine deaminase (AID)
Further DNA recombination allows V regions to be joined with
different C regions
Isotype switching only occurs in
B cells that have been activated
isotype switching involves
recombination between specific switch regions
Antibodies with different C regions have
different effector functions
identify which of the following is associated with activation induced cytidine deaminase activity?
diversification of the VH domain but not the VL domain
The derivation of antibodies from a single clone of B lymphocytes that have identical antigen specificity is reffered to as
monoclonal antibody production