1 antibody structure

Question 1

what are antibodies and what cell type produces them?

Answer 1

• antibodies are the effector arm of humoral immunity

- they are produced by B lymphocytes and bind to antigen

Question 2

B cells and T cells are both types of?

Answer 2

lymphocytes - part of the adaptive immune system

Question 3

difference between adaptive and innate immune systems?

Answer 3

adaptive is faster and has memory

Question 4

role of T cells?

Answer 4

• cytotoxic

- help other cells in the immune response

Question 5

how can innate immunity can be divided?

Answer 5

humoral or cellular

Question 6

humoral immunity?

Answer 6

mediated by macromolecules found in extracellular fluids such as secreted antibodies, complement proteins, and certain antimicrobial peptides.

Question 7

cell mediated immunity?

Answer 7

doesn’t 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.

Question 8

what is the surface bound AB and what happens upon binding?

Answer 8

BCR

-upon binding, the ab signals internally to stimulate the B-cell to proliferate and produce secretory antibody

Question 9

how do the different clones of B cells differ?

Answer 9

they each have an individual specificity for antigen

Question 10

how many chains is an antibody made of?

Answer 10

4 chains
2 light and 2 heavy

The binding site of the antibody involves all 4 chains.

Question 11

describe antibody structure

Answer 11

4 polypeptide chains: 2 identical heavy & light chains
held together by S-S bonds
glycosylated to allow for specific confirmation
Fab region - determines binding specificity
Fc region - determines function

Question 12

Fab region

Answer 12

antigen binding region, composed of the variable regions of the heavy and light chain

determines the specificity and the affinity and avidity of the interaction with antigen.

Question 13

proteases split the antibodies into?

Answer 13

2 parts - fab and fc regions

Question 14

Fc region

Answer 14

Confirms the functional properties of antibody

Recognised by FcR (cell receptors) and binds complement (effector molecule of the immune system)

Question 15

how are antibody classes defined?

Answer 15

defined by the heavy chain constant region
9 (sub) classes of antibody (in humans) determined by:
4x γ
1x μ
2x α
1x δ
1x ε

Question 16

what antibody is a pentamer?

Answer 16

IgM, hence the large molecular weight

Question 17

what antibody is a dimer?

Answer 17

sIgA

Question 18

the majority of our serum contains what antibody?

Answer 18

Majority in the serum is IgG

-IgG has a higher half life, its is stimulated when you have been primed against an antigen

Question 19

different antibody functions?

Answer 19

1. immune complex, complement classical pathway (C1q)
   - antigen binds to fab2 regions forms complex which complement can bind to
2. opsonisation + phagocytosis
   - antibody binds to antigen, and antibody can bind to the Fc receptor on phagocytic cells
3. target cell
   - Fc receptor on killer cells - infected cell with antigen on surface
4. primed cell, sensitisation
   - IgE produced in first encounter, second encounter the IgE binds to antigen and mast cells degranulate, histamines and anaphylaxis

Question 20

IgM

Answer 20

5 identical IgMs linked together by a J chain
They each have a low affininty, but when you put them together you get a high avidity

Heavy chain encoded by μ gene
Constitutes ~ 10% serum Ig
Major antibody in primary response - first one made
4 CH domains, stabilised by J-chain
Monomer forms the BCR on most B cells in association with Igα and Igβ chains

Question 21

IgG

Answer 21

Because IgG is small and monomeric, it can cross the placenta and protect fetal development

Monomeric, heavy chain γ gene,3 CH domains.
Major circulating Ig (70-75%)
Major antibody in secondary response
4 sub-classes G1; G2; G3; G4
Activates complement (Classical)
Opsonin (FcR)

Question 22

IgA

Answer 22

Range of polymers (shapes) from monomer to pentamer - mostly monomer and dimer in secretions.

1 J chain per polymer - the J chain stabilises it

Heavy chain encoded by α-gene

15-20% circulating Ig

Epithelium adds secretory component which protects against degradation due to proteolysis; aids release into mucous.

2 subclasses IgA1 in serum IgA2 in mucous

Found in tears, milk, saliva, sweat etc

Protection of external surfaces- first line of defence

Can get localised mucosal response, different to systemic response - It is secreted onto the mucosa to protect against enteric pathogens

Does not activate complement (by classical pathway)

Question 23

IgA secretion

Answer 23

IgA dimers bind to Poly-Ig receptors on basolateral surface of the epithelial cells.

Complex endocytosed and still bound to membrane.

Vesicles fuse with the luminal surface and receptor cleaved by proteolysis.

Dimer released attached to secretory component

Question 24

IgD

Answer 24

Heavy chain coded by δ gene

Monomer, very like IgG but slight differences in heavy chain constant region
<1% serum Ig

Has specific antigen binding activity but NO effector functions

Sensitive to proteolytic degradation and heat

Expressed on the surface of B cells with monomeric IgM

Involved in antigen triggered B-cell differentiation

Question 25

IgE

Answer 25

Heavy chain coded by ε-gene

Heavy chain has 4 constant domains (looks like monomer IgM)

Trace levels in serum (elevated in allergic or heavy parasitic infection)

Majority bound to mast cells and basophils through high affinity FcεR1

Key to allergic response

Important role in parasitic infections, mast cells very effective at killing multicellular organisms

Question 26

what do most B cells express?

Answer 26

monomeric IgM and IgD

• Both Ig have the same specificity on each individual cell
• 10% of circulating B cells express IgG, IgA or IgE (after memory and differentiation)
• Some tissue bias, eg mucosal B cells express IgA

Question 27

structure of surface bound Ig?

Answer 27

very short intracytoplasmic tail, thus is associated with ‘accessory’ molecules to form the BCR

Igα (CD79a) and Igβ (CD79b) - (similar to TCR and CD3)

Question 28

BCR binding effects

Answer 28

Ligation of the BCR results in ITAM phosphorylation (Immunoreceptor Tyrosine-based Activation Motifs).

Leads to downstream cascade of events resulting in plasma cell differentiation and antibody production.

CD79a and b are associated with the B cell receptor to cause the transmission and activation of B cells

Question 29

Outcome of BCR

ligation

Answer 29

When a B cell becomes activated, it can become a plasma cell or a memory cell

Clonal expansion: B cell becomes activated and becomes a clone of cells with exactly the same BCR specificity

A small proportion of cells remain as long-lived memory cells

The majority of cells become effector cells (plasma cells) which produce antibody, lots of it!

Plasma cells have a limited life-span and apoptose after a few days

Question 30

Class switching

Answer 30

First Ab response is IgM

Secondary response is switched to other classes of Ab, especially IgG

Mutate their variable region gene sequences- somatic hypermutation

Affinity maturation

Needs T cell help

Question 31

role of mutation in class switching?

Answer 31

Ab’s mutate their binding site in a random fashion to increase the affinity of the binding site for the antigen

In the secondary response – we get a different type of antibody and that has a greater capacity for binding its antigen

Question 32

Monoclonal antibodies

Answer 32

The single most important immunological discovery of the century!

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.

Question 33

examples of new checkpoint inhibitors:

Answer 33

Ipilimumab:- anti-CTLA-4
Nivolumab:- anti-PD1

Question 34

problems with monoclonal antibodies?

Answer 34

Constant region heterogeneity
Most early McAb were murine
Anti-antibodies
Inflammation

solutions: Chimeric antibodies: Murine antigen recognition sites grafted to human constant regions…. Humanised.

Fully humanised antibodies: ‘phage display (in vitro McAb) and humanised mice!

Question 35

main functions of monoclonal antibodies

Answer 35

anti inflammatory or anti cancer

Infiximab used for athrities (anti TNF-alpha).

TNF-a = inflammatio, partcuilalry in joints. Helped
athritis a lot. Mops up inflam mediators, causing disease, (so doesn’t realy stop disease)

Rituximab binds to molecule found on B cell, to treat B cell tumour.

Herceptin – binds to receptor for growath factor.

Zumab – humanized molecule antibody has a bit of change, to look like human molecule.

Ximab at the end = chimeric antibody. (part of human and mouse antibody – they do this to stop it
from becoming an antigen itself)

new checkpoint inhibitor: Nivolumab is anti-PD1, stops switching immune system off.

Question 36

affinity vs avidity

Answer 36

affinity is the strength of a single interaction between an antibody and its epitope

Avidity is the sum of the different affinities (some antibodies will be multimeric and will have several binding sites)