Antibodies Flashcards
The Immune System: The Cells
What are the 2 catagories of immunity cells? and how are each of these decribed?
- Innate immunity=rapid response
- Adaptive immunity= slow response
The Immune System: The Cells
All cells of the immune system share a common progenitor – What is it?
- The hematopoietic stem cell
- Multipotent
The Immune System: The Cells
What produces the vast array of ”white blood cells” that make up our immune system?
Rounds of differentiation and specialisation
The Immune System: The Lymphoid Lineage
What are the 3 types of cells found in this category?
And where do they all start life?
- T- cells
- B-Cells
- NK-Cells
All start life in the bone marrow, receiving signals from stromal cells
The Immune System: The Lymphoid Lineage
Where do T cells mature and what is their function?
- Mature in the Thymus
- Regulatory and Cytotoxic
The Immune System: The Lymphoid Lineage
Where do B cells mature and what is their function?
- Mature in the Bone Marrow
- Produce antibodies
The Immune System: The Lymphoid Lineage
Where do NK cells mature and what is their function?
- Mature in the Bone Marrow and also secondary lymphoid tissues
-Cytotoxic
B-cell development:
What is B-Cell development dependent on?
Dependent on the transcription factor Pax5
B-cell development:
The development of the B-cells and the B-cell receptor, involves a complex series of what? And what does this produce?
- Involves a complex series of gene recombination
- To produce the vast array of antibodies we need
Antibody Development:
When are IgG and IgM detectable by?
By around 2 weeks
Antibody Development:
When may detection of IgG and nAb decline by?
After 8 weeks
Antibody Development:
What are the 2 theories of antibody development?
- Instructive theory
- Selective theory
Antibody Development:
Who came up with the Instructive theory and give a description of the theory
Instructive Theory: Linus Pauling -1940
- The antigen acts as a template that directs the folding of the unfolded antibody chain
- So a specific amino acid chain has the propensity to form a variety of antibodies, depending on the antigen present at the time
Antibody Development:
Who came up with the Selective theory and give a description of the theory
Selective theory: Macfarlane Burnet, Niels Jerne, David Talmage, and Joshua Lederberg – 1950s
- The binding site of the antibody is already pre-determined before antigen is even encountered
- The presence of antigen only affects how much of that specific antibody you get
Antibody development – The Instructive Theory
What does the instructive theory suggest? And is it correct?
- Suggests that if the antibody protein was unfolded and then refolded without antigen present it would lose its affinity
- The theory was incorrect, and the binding of antigen is dependent on the amino acid sequence
Antibody development – The Selective Theory
What does the selective theory suggest and is it correct?
- Suggests that each antibody producing cell, makes a single kind of antibody. And this commitment is determined before the antigen is even present
- this theory is correct
Antibody development – The Selective Theory
Each cell has a distinctive base sequence in its DNA, what does this determine?
- Determines the amino acid sequence
Antibody development – The Selective Theory
As the cell matures…What does each antibody producing cell make? What then happens to the cell if it binds to a molecule during foetal development?
- Makes a small amount of cell surface-bound antibody.
- The cell is killed – therefore preventing B-cells that act against itself
Antibody development – The Selective Theory
Later when the mature cell is activated, what process does it begin to undergo?
Begins to undergo clonal expansion when exposed to antigen
Antibody development – The Selective Theory
What does a Heterogeneous cell population equal?
Heterogeneous cell population = heterogeneous antibodies
Classes of Antibody
What are the 5 antibody classes?
- IgA
- IgD
- IgE
- IgG
- IgM
Classes of Antibody:
What are the properties of the antibody IgA? and draw its structure
- Found in mucous, saliva, tears and breast milk
- Protects against pathogens
Classes of Antibody:
What are the properties of the antibody IgD? and draw its structure
- Part of the B cell receptor
- Activates basophils and mast cells
Classes of Antibody:
What are the properties of the antibody IgE? and draw its structure
- Protects against parasitic worms
- Responsible for allergic reactions
Classes of Antibody:
What are the properties of the antibody IgG? and draw its structure
- Secreted by plasma cells in the blood
- Able to cross the placenta into the fetus
Classes of Antibody:
What are the properties of the antibody IgM? and draw its structure
- May be attatched to the surface of a B cell or secreted into the blood
- Responsible for early stages of immunity
Antibody Structure - Heavy and Light Chains
What is Fab and Fc in antibody structure?
Fab = Antigen binding Fragment
Fc = Crystallisable region fragment
Antibody Structure - Heavy and Light Chains
What is IgG made up of?
- 2 polypeptide chains:
Light Chain x1
Heavy Chain x1
H2L2: Heavy/Light
Antibody Structure - Heavy and Light Chains
What is the antibody structure held together by?
Held together by disulphide bridge
Antibody Structure - Heavy and Light Chains
Both chains consist of what 2 regions? and how many amino acids are in each?
- a variable region and a constant region
Variable= 1 – 108 amino acids
Constant= 109 – C-terminus
Antibody Structure - Homologous Domains
Vl and Vh are both what?
Very similar
Antibody Structure - Homologous Domains
The heavy chain consists of what?
3 equal thirds
Antibody Structure - Homologous Domains
Where is the intrachain disulphide bond located?
- Located in the same place on every domain of both the H and L chains
Antibody Structure – Immunoglobulin Folding
What does this consist of?
Broad sheets of anti-parallel β-sheets
Antibody Structure - Homologous Domains
What is CL very similar to?
The 3 CH domains
Antibody Structure – Immunoglobulin Folding
Hydrophobic side chains are tightly packed between the anti-parallel beta sheets. What are they held together by?
Held together with disulphide bridges
Antibody Structure – Immunoglobulin Folding
What are the constant and variable domains?
Constant domains:
3 β-sheets in one strand
4 β-sheets on the other
Variable domains:
2 additional β-sheets
3 loops that compose the part of the antigen binding site
Antibody Diversity:
What did William Dryer and Claude Bennett discover in 1965?
- Multiple variable genes
- 1 Constant gene
- These genes are separated, and one V becomes joined to the C during differentiation and maturation of the B-cell
Antibody diversity:
What did Susuma Tonegawa confirm in 1985?
- Confirmed that V and C genes are far apart in the embryonic DNA
- And are closely associated in B-cells
- Immunoglobulin genes are translocated during the differentiation of B-cells
Antibody Diversity:
How many different kinds of antibodies can be made by an animal?
Number in the millions
Antibody Diversity:
What are the several hundred genes for the variable regions of a mouse?
- 300 for the light chain
- 300 for the heavy chain
- 9x10^4 different specificities
Antibody Diversity:
There is a large discrepancy between the germline variation and adult variation. What does this mean must be generated?
Must be an alternate source generated during the lifetimes of an animal and the differentiation of its lymphocytes
Antibody Diversity – V(D)J Recombination:
What does the V gene not encode for and when does it stop?
- The V gene does not encode for the entire polypeptide chain.
- Stops at amino acid 95, 13 residues too short
Antibody Diversity – V(D)J Recombination:
Light chain:
Where is the stretch of DNA that encodes for this region found?
- Found nearer the Constant region
- an array of 4 joining regions
Antibody Diversity – V(D)J Recombination:
- What can happen to 1 of the several hundred V genes?
-Can become spliced to one of the 4 Joining genes
-Further diversity added as this splicing can occur in different joining frames
Antibody Diversity – V(D)J Recombination
Heavy chain: The heavy chain has an additional region, What is it?
- The D region
D = Diversity region
Antibody Diversity – V(D)J Recombination
How many genes are in the D region?
15 genes within this region
Antibody Diversity – V(D)J Recombination
What is special about the enzyme deoxyribonucleotidyl transferase and what does it do?
- It is a special polymerase that does not use a template
- It inserts extra nucleotides between the V and D regions
Antibody Diversity – V(D)J Recombination
What does the light and heavy chain consist of?
LIGHT CHAIN:
-250 V genes
- 4 J genes
- At least 3 possible joining frames
- 250 x 4 x 3 = 3000
HEAVY CHAIN:
- 250 V genes
- 5 J genes
- 15 D genes
- At least 3 possible joining frames
- 250 x 5 x 15 x 3 = 56,250
- 3000 x 56,250 = 1.7x10^8 possible variations
Antibody Diversity – Class Switch Recombination
What do B-cells only produce at the beginning of life?
Begin life only producing IgM
Antibody Diversity – Class Switch Recombination
What is Class Switching recombination needed for?
- To produce the other types of antibody
Antibody Diversity – Class Switch Recombination
Different classes of heavy chains depend on what?
The antibody type
G – γ
M - µ
A – α
D - δ
The Germinal Centre reaction:
What does somatic Hypermutation result in?
- Results in mutations being introduced into the V region of the heavy chain and light chain
-Altering the affinity of the immunoglobulin for its antigen
The Germinal Centre reaction:
In comparison, What happens in Class Switch Recombination?
- The initial heavy‐chain C regions are replaced by another isotype
- Modifying the effector activity of the immunoglobulin but not its specificity
The Germinal Centre reaction:
Where does somatic hypermutation and class switch recombination occur? And what other cell types are required?
- These occur in secondary lymphoid tissues as part of the Germinal Centre reaction
- Requires other cell types:
CD4+ T-cells
Dendritic Cells
Practical Uses of Antibodies:
What are 3 practical uses of antibodies and provide examples for each.
- Staining
- Microscopy
- Flow cytometry
- Western blotting - Antigen Capture
- ELISA - Drugs
- Highly specific to the target, so reduced off target effects
- Chemotherapy
