antibody structure Flashcards
how is immunity classified?
innate and adaptive
- innate is the first response, quicker but no memory
- adaptive can be split into humoral (antibodies) and cell-mediated (cytotoxic T cells, phagocytes)
what makes antibodies?
B lymphocytes
BCR is a…?
surface bound antibody
how many diff antibody specificities does the human body have?
10^8
antibody structure
consists of 4 polypeptide chains, 2 identical heavy chains and 2 identical light chains.
chains held together by disulphide (S-S) bridges - stabilised
the antibodies are glycosylated, which makes a particular type of confirmation
Fab (determines antigen binding specificity) and Fc region (biological activity, complement will bind within here)
hinge region for flexibility
fab vs fab2?
F(ab’)2 and Fab, are antigen-binding fragments that can be generated from the variable region of IgG and IgM, by protease digestion
F(ab’)2 fragments consists of 2 antigen-binding regions joined by hinge through disulfides. This fragment is void of most of the Fc region.
domains?
Fab2 and Fc region have individual domains - domains can be duplicated to produce variability
Avidity
the overall strength of binding between an antibody and an antigen (sum of all the binding sites)
Humoral immunity
immunity mediated by macromolecules found in extracellular fluids such as secreted antibodies, complement proteins, and certain antimicrobial peptides.
Cell-mediated immunity
immune response that does not involve antibodies, but rather involves
the activation of phagocytes, antigen-specific cytotoxic T-lymphocytes, and the release of various
cytokines in response to an antigen.
Fab2 region
constitutes the antigen binding region
- Antigen recognition site composed of the variable region of both the heavy and light chain
- This determines both the specificity and the affinity and avidity of the interaction with antigen.
Fc region?
It’s the crystallizable fraction. Gives Ab its function.
- Fc region confirms the functional properties of antibody.
- Recognised by FcR (Fc receptors)
- Binds complement
how do antibody classes differ?
differ depending on their heavy chain constant region 9 subclasses of antibody in humans 4 gamma 1: mu, epsilon, delta 2: alpha
antibodies have 4 different functions - what are they?
- immune complex, complement classical pathway (C1q)
- antigen binds to fab2 regions forms complex which complement can bind to - opsonisation + phagocytosis
- antibody binds to antigen, and antibody can bind to the Fc receptor on phagocytic cells - target cell
- Fc receptor on killer cells - infected cell with antigen on surface - primed cell, sensitisation
- IgE produced in first encounter, second encounter the IgE binds to antigen and mast cells degranulate, histamines and anaphylaxis
IgM
- pentameric, 5 linked by J chain which stabilises the whole structure
- 4 heavy chain constant regions
- first class of Ab to evolve in the primary response, low affinity but high avidity overall –> ROUGH and READY, binds to lots of stuff but just weakly
- the IgM monomers mainly forms the BCR
- activates classical complement pathway
IgG
monomeric, heavy chain with 3 constant regions
highest conc in serum (circulating) (IgG 1, 2, 3, 4 subclasses)
can cross placenta and protect foetus
activates classical complement pathway
opsonisation
IgG has a long half life, once primed to antigen, IgG is secondary antibody produced
IgG split half and half between vasculature and outside
venous system. 50% of IgG is in tissues.
IgA
lots of diff polymers, in humans mostly monomers and dimer and 1 J chain per polymer
next highest circulating conc after IgG
2 subclasses, IgA1 in serum, IgA2 in mucous
found in milk, sweat, saliva, tears
lines mucosal defences
does not activate complement
how does IgA become secretory IgA?
transported across mucosal epithelium from the basolateral surface to the luminal surface
IgA dimer binds to a poly-Ig receptor and forms a complex with it. this gets endocytosed into the cell and enclosed in a transport vesicle. the receptor gets cleaved by proteolysis and the IgA dimer with new secretory component is released - now secreted IgA!
IgD?
like IgG but a few diff in heavy chain constant region
specific antigen binding sites but no effector function, expressed on B cell surface with monomeric IgM
very low serum conc
IgE?
heavy chain - 4 constant regions
Majority bound to mast cells and basophils through high affinity FcεR1
involved in defence against parasitic infections, eg. helminths - mast cells good at killing multicellular organisms
Trace levels in serum (elevated in allergic or heavy parasitic
infection)
surface bound Ig?
this Ig has a short intracytoplasmic tail, therefore needs accessory molecules to form the BCR - CD79 which forms a complex with the BCR after antigen binding, and is made of 2 chains: CD79a and CD79b
most B cells express monomeric IgM or IgD, both have same specificity on each cell
(some tissue bias, e.g. mucosal B cells express IgA)
what does BCR ligation (recognition) lead to?
leads to ITAM phosphorylation (immunoreceptor tyrosine based activation motifs), which leads to downstream cascade resulting in differentiation into plasma cell and antibody production.
results in clonal expansion where the B cell activated gets cloned and more copies of it are made. small % live as memory cells, majority become effector cells (plasma cells) which produce antibody. plasma cells have a short lifespan and apoptose after a few days.
class switching?
f1st Ab response is IgM, class switching occurs and other classes made, esp IgG.
somatic hypermutation (mutations of variable region gene sequences) which increases affinity of binding site for antigen and affinity maturation
how are antibodies used in clinic?
monoclonal antibodies!
developed by Kohler and Milstein in 1975
Inject the antigen you want to make antibodies against into the animal, and harvest the B cells from spleen after a few weeks.
Fuse B cells with myeloma cells (constantly
growing lymphoid cells). Myeloma cells are immortal cells from a B cell tumour, they lack the HGPRT gene.
Ethylene glycol is used to help fuse - melts membranes. Forms hybidroma which are the only cell type that can grow in the HAT medium.
The hybridoma will spontaneously grow like myeloma. And also produce Ab.
