Humoral Immunity Flashcards
Key Points about Antibodies
- Antibodies Neutralize or Eliminate Extracellular microbes and their Soluble Products (Toxins).
- Circulating Antibodies Neutralize microbes by **Blocking their entrance into Host cells, before they proliferate and spread.
- Antibodies are produced against Proteins, Lipids, Nucleic Acids, Carbohydrates, and Polysaccharides, and small chemicals.
- Antibodies are particularly effective against microbes with Capsules rich in Polysaccharides and Lipids.
- Most Effective Vaccines Stimulate Antibody Production.
How to Develop Circulating Antibodies
Congenital: Passive
- -Newborns acquire Antibodies Transplacentally prior to Birth
- -And from Milk after birth.
- -Providing immunity in infancy.
Infections: Active
–Primary Immune Response leaves Memory B cells that secrete low levels of Antibody for Months or even many Years.
Immunization by Vaccination:
–Whole Microbe or Part or its toxin is given under controlled circumstances to Induce a Primary Immune Response –> To Prevent Subsequent Infection.
B cell Maturation
Occurs in Bone Marrow.
Progenitors give rise to Pro-B cells.
1) Pro-B cells undergo Ig Heavy chain Recombination, mediated by VDJ Recombinase.
2) Pre-B cells are those that contain Ig mu Heavy chain protein in cytoplasm.
- –They then express the Pre-BCR Complex on Cell Surface, composed of mu Heavy chain (with 2 surrogate light chains and with the Ig alpha and beta signaling molecules).
- –Pre-BCR complex delivers intracellular signals: (1) Promote survival and proliferation; (2) Ig Light chain Recombination.
- –(3) Allelic Exclusion inhibits VDJ Recombinase from acting at other chromosome’s heavy chain.
3) Immature B cell: Expresses a complete IgM Antigen Receptor on cell surface, composed of a mu Heavy chain and a Light chain (kappa or lambda).
4) Mature B cell: Expresses both IgM and IgD on cell surface.
5) Positive Selection of Weak Self-Antigen Recognition.
- -Negative selection of strong or no recognition of self antigen.
Immunoglobulin Structure
4 Polypeptide Chains: 2 identical Heavy (H) chains; 2 identical Light (L) chains.
- -Each Light chain is attached to 1 Heavy chain.
- -The 2 Heavy chains are attached to each other.
Light Chains contain 1 Variable region and 1 Constant region.
Heavy Chains contain 1 Variable region and 3-4 Constant regions.
Each Variable region of the either chain contains 3 Hypervariable regions (CDRs).
- -CDR3 has the greatest variability.
- -CDR3 is located at the junction of the V and C regions.
Light Chains are not attached to cell membrane.
Heavy Chains are either membrane-bound or part of soluble Ig.
Light chains do not switch and remain fixed throughout the life of that B cell. They do not affect Antibody function.
2 Types of Light Chains (by Constant regions): Kappa and Lambda
5 Classes/Isotypes of Heavy Chains (by Constant regions): IgA (alpha type C region) IgM (mu type C region) IgG (gamma type C region) IgE (epsilon type C region) IgD (delta type C region)
Fab Regions
Fab Region (Fragment, Antigen-Binding):
- —Region of the Antibody that contains a whole Light chain attached to the V and first C domains of a Heavy chain.
- —Is the portion of the Antibody required for Antigen Recognition.
There are 2 identical Fab regions on an Ig.
Fc Region
Fc Region (Fragment, Crystalline)
There is one Fc region on the Ig.
It is the region of the Antibody containing the remaining Heavy chain C domains.
Function:
- -**Most of the biologic activity and Effector functions of the Antibody.
- -**Binds to cells on their Fc Receptors only if Fab is Bound to Antigen!!
Hinge Region
Located between the Fab and Fc regions.
Flexible.
Allows the two Antigen-binding Fab regions to move independently of each other, enabling them to simultaneously bind Antigen Epitopes that are separated from one another.
Epitope / Antigenic Determinant
The region of the Antigen recognized by Variable Domains.
- Each Antibody typically recognizes 1 Epitope.
- Because there are 2 Fab sites on a given Ig, this can bind to 2 Identical Antigen Epitopes.
- Microbes have multiple Antigen, each one with multiple Epitopes.
- -Some may be *Highly Repetitive.
Affinity
= Fab binding strength to an Antigen Epitope.
Affinity Maturation
During Immune responses, the quality of the Antibody improves, increase its Affinity.
Cross-Reactivity
A particular clone of B cells produces Antibody specific for 1 Antigen Epitope.
Antibody to one particular Antigen Epitope may also exhibit binding, although weaker, to another epitope that is structurally similar = cross reactivity.
Avidity
= The total strength of Antigen-Antibody binding, not just one interaction.
IgG exists as a monomer and thus can form 2 Ag-Ab bonds (b/c 2 V domains).
IgA forms a *Dimer; can bind 4 Antigens.
IgM forms a *Pentamer; can bind 10 Antigens; has Highest Avidity.
Polyclonal Antibody Production vs. Monoclonal Antibody Production
Polyclonal: Antibodies against microbes are made to Multiple Antigen Epitopes by Multiple Clones of B cells.
*Polyclonal B cells have an even kappa/lambda ratio.
Monoclonal: When only a single clone of B cells produces Antibody.
- -> *This is Pathological and typical produced by B cell Tumors.
- -Will show an unequal kappa/lambda ratio.
- -Also done in lab via Hybridoma Technique: (1) Pregnancy Test that uses Antibodies to HCG;
(2) Drugs, e.g. Herceptin, used to treat Breast Cancer.
IgD
Remains membrane-bound.
Never secreted.
Function: Antigen receptor on B cell.
IgM
IgM is the First Ig Secreted during Primary Immune Response.
–The Best Activator of Complement.
(–May opsonize, but less effectively than IgG.
–Not a good activator of NK cells.)
Effector Functions:
–Complement Activation (Classical) (Best!)
IgG
IgG is the *Most Abundant Ig in Blood. 4 subclasses: IgG1-4.
- -*Best to Neutralize Microbes;
- -*Best Antigen Opsonizer;
- -An okay *Activator of Complement;
- -Best NK cell Activator.
Effector Functions:
1) Neutralization of Microbes and Toxins (Best!)
2) Opsonization (Best!) for Fc Receptor-Dependent Phagocytosis: IgG1 and IgG3 bind to Fc receptors on Neutrophils and Macrophages specific for gamma Heavy chain.
3) Antibody-Dependent Activation of NK cells (Best!)
4) Complement Activation (Classical)
5) Feedback Inhibition of B cell Activation
6) Neonatal Immunity (Placental transfer)
IgA
IgA is the Most Abundant Ig in the Whole Body. Is Located on Mucosal Surfaces.
–*Is Mainly Secreted into the Respiratory, Urinary, and Genital Tracts to provide “Mucosal Immunity.”
Effector Functions:
–Mucosal Immunity (IgA transported through epithelia)
IgA is produced by MALT in submucosa, then actively transported across the epithelia to Mucosal Surface to bind and *Neutralize Microbes (prevent them from penetrating epithelia).
The *Poly-Ig Receptor on *Basal Epithelial Cells binds IgA Fc region and triggers Endocytosis.
Locally produced TGF-beta promotes Isotype switch to IgA.
B1 B cells migrate to MALT and respond to Non-protein Antigen.
IgE
IgE is typically *Bound to Mast Cells
- -Mediates Cytotoxicity of Basophils, Mast Cells, and Eosinophils against Parasites, especially worms.
- -Also involved in Allergic Reactions.
Effector Functions:
- -Immunity against Helminths
- -Mast Cell Degranulation (immediate hypersensitivity)
IgE activates *Mast cells and *Eosinophils against Helminthic Parasites and in *Allergic Reactions.
Eosinophils have *Fc-eRI and IL-5 Receptors; activation releases their granular contents and is toxic to Helminths.
*IgE binds to Fc-eRI on Mast cells, which secrete Histamine and Chemokines, *Attracting more Eosinophils.
Follicular B cells
Follicular B cells:
- -Located in Follicles of Spleen and other Lymphoid organs
- -Recognize Protein Antigen with help from Helper T cells
- -Undergo Isotype-Switching
- -Produce High-Affinity Antibodies (Affinity Maturation)
- -Are Long-lived Plasma Cells
- -Can become Memory B cells
Marginal Zone B cells
- -Located in the Spleen and other Lymphoid Organs.
- -Recognize Polysaccharides, Lipids, etc. (T-Independent)
- -Have limited diversity Antigen Receptors
- -Produce Mainly IgM
- -Are Short-lived Plasma cells
- -Cannot become Memory B cells
B-1 cells
- -Located in Mucosal Tissues and Peritoneal Cavity
- -Recognize Polysaccharides, Lipids, etc. (T-independent)
- -Have limited diversity Antigen Receptors
- -Produce Mainly IgM
- -Are Short-Lived Plasma cells
- -Cannot become Memory B cells
Primary Response vs. Secondary Response
Primary Response:
- -First exposure of B cell to Antigen, regardless of type of Antigen.
- -Low amount of Antibody produced; predominantly IgM.
Secondary Response:
- -Repeated exposure
- -Higher amount of Antibody; predominantly IgG
- -Helper T cells cause Isotype Switching to occur and increased Affinity
Antigen Recognition of B cells
1) Antigen is transported and concentrated in B cell rich Follicles and Marginal Zones of Peripheral Lymphoid Tissues.
2) B cells recognize whole, unprocessed Antigen via Membrane-Bound IgD Antigen Receptors.
3) IgAlpha and IgBeta accessory molecules of the BCR Complex transduce intracellular activation signals via their ITAMs motifs.
4) BCR complex clustering Cross-Links and Phosphorylates ITAMs, initiating signaling.
- *Proteins initiate Weak B cell Signaling due to Limited Cross Linking: (Proteins rarely have repeated epitopes)
- -> Therefore, T-Dependent
**Polysaccharides, Lipids, and Nucleic Acids contain High numbers of Identical Epitope, leading to Abundant Cross-Linking and strong activation signals on their own:
T-Independent
Costimulation of B cells
Naive B cells require Costimulators.
1) Microbial Activation of Complement generates C3d.
2) C3d binds to Microbe.
3) Complement Receptor Type 2 (CR2) on B cells recognizes the C3d bound to the microbe, while IgD receptor recognizes the Antigen.
- -> Second Signal.
4) Microbes also produce Direct Second Signals:
- -B cells have Receptors for Microbial products, such as *TLR
- -Activation of these receptors stimulate B cell proliferation and Ig secretion.
CD4+ Helper T cell Costimulate B cells
1) B cells recognize Unprocessed Protein Antigen as a whole, ingest it, digest it, and Display it on Class 2 MHC (Simultaneously acting as an APC)
2) CD4+ Helper T cells recognize it and activate B cell via CD40L - CD40 interaction and cytokine secretion.
3) Activated B cells express B7, Costimulator for T cell (CD28).
CD40L-CD40 interactions results in further Costimulation, lowering the threshold for B cell Activation, Ig Secretion, Heavy Chain Class Switching, and Affinity Maturation. (Analogous to T cell-mediated Macrophage Activation).
Although B cell and CD4+ T cells recognize different Epitopes of the Protein Antigen, Specificity is Maintain for the same Antigen Protein.
B cells are Very Efficient APCs.
B cell Ig receptor has very High Affinity and can Recognize Antigen in Very Low Concentrations.
Due to the Weak or complete lack of Antigen cross-linking, B cells recognizing Protein Antigen still cannot initiate signaling, even after Peptide Antigen is bound to BCR and Second Signals are delivered by Microbes and by Complement.
*Additional costimulation is required from CD4+ Helper T cells by binding of CD40L on T cell to CD40 on B cells.
This reduces signaling threshold, overcoming weak cross-linking and BCR clustering.
Functional Consequences of B cell Activation
1) *Initiates B cell Proliferation and differentiation: Very strong for Polysaccharides and Lipids
2) *Sensitizes B cells for CD4+ Helper T cell interaction via expression of Cytokine Receptors for CD4+ Helper T cell-produced Cytokines.
3) *Increases B cell expression of B7, costimulators for CD4+ T cells.
4) *Decreases B cell expression of B cell Zone Chemokine Receptors
5) Low-level IgM secretion
Heavy Chain Isotype Switch
CD4+ Helper T cells induce B cells to Isotype switch by CD40L mediated signals and Cytokine signals.
CD40 induces Activation-Induced Deaminase (AID) expression,
–> Switching VDJ recombination from C-mu to other Downstream C genes.
CD4+ Helper T cell Cytokines:
IFN-gamma –> IgG
IL-4 –> IgE
Cytokines produced by Mucosal tissues (e.g. TGF-beta) –> IgA
Ig Affinity Maturation
Antibody Affinity for Antigen increases with Prolonged exposure to Antigen.
Occurs in Germinal Centers where B cells divide quickly.
Process results from *Somatic Hypermutations in the Hypervariable Regions of Ig genes of Dividing B cells.
***AID plays a critical role by inducing Nucleotide Mutations.
1) B cell clones of varying affinities emerge.
2) *B cells undergo Apoptosis unless Primed by Antigen or Rescued by Helper CD4+ T cells.
3) *Circulating Antibody binds to Antigen and Complement to Form *Complexes.
4) *Follicular Dendritic Cells (FDCs) have Fc Receptors that bind to these Complexes and then Presents them to B cells.
5) *B cells uptake Antigen presented by FDC and present it to Helper CD4+ T cells who migrate to Germinal centers to provide Survival Signals to B cells.
6) *As the response proceeds, Antibody Increases and *Antigen Decreases to Low Concentrations.
7) *B cells with High Affinity Ig Receptors are Selected based on their Ability to Bind Antigen at Low Concentration and Survive.
Evasion of Humoral Immunity
Microbes can resist Humoral immunity by *Variation on their Surface Macromolecules, such as Glycoprotein Coat.
Also, Inhibition of Complement Activation.
Or Resistance to Phagocytosis.
Resistant strains become selected by Antibody responses.
Vaccination
*Purpose: Artificially induce protective immunity to Prevent Infection.
Can also *Prevent an Epidemic if enough people are immunized because microbe is less able to spread among population.
Instead of using “real”microbe, *Many vaccines use an agent that *Cannot Cause Disease, but that *Stimulates Specific Antibody Production using **Killed or *“Inactivated” Microbes or even **Toxins inactivated by *Heat or *Chemical treatment:
- -*Toxoid: Inactivated Toxin
- -*Killed Agent: Inactivated Bacteria or virus
- -*Subunits: Protein subunit of bacteria or virus.
Other strategies use **Live “Attenuated” Microbial *Strains.
**To produce a Protective CD8+ CTL Immune Response against Viral infection, a Live “Attenuated” Vaccine must be given.
–Only live viral infection of APC will result in Class 1 MHC-Antigen peptide presentation and CTLs.
**“Inactivated” Vaccines tend to be Less Effective for Some Viruses, require *Larger Dose, and Periodic *“Boosters” to maintain immunity.
–“Inactivated” Vaccines are *Generally used Against Bacteria or Toxins.
Active Immunization
With “Active” immunization, Vaccine is given prospectively to Initiate immune response with 2 Purposes:
1) To Prevent possible Disease in the person given the vaccine
2) To Create Immunity in Multiple Individuals to Decrease the Probability of Spreading/Epidemic.
Passive Immunization
With “Passive” immunization, *Ig (preformed Antibody) is *Given to an individual at the Time of Infection to:
1) **Stop the Continuation of the Infection and Minimize Damage
2) May also be given to persons who have Poor Immune Responses because they are **Immunocompromised, to *Prevent Infection.
Regulation of Humoral Immunity
–> Antibody Feedback:
*As Antigen declines, B cells die.
A special mechanism stops Ig secretion:
1) **Circulating IgG binds to Antigen, forming Complexes that bind to Fc Receptors on B cells
(Fc-gamma-RII Receptors; don’t need to know name)
2) These Fc Receptors provide *Inhibitory Signals when bound to IgG-Antigen Complexes
==> **Antibody Feedback Inhibition
Neonatal Immunity
Newborns have incompletely developed immune system.
Maternal Antibodies are Actively transported across Placenta to Fetus and Intestinal Mucosa in Neonate, providing Passive Immunity.
**Transfer relies on Neonatal FcRn Receptor.
**Maternal IgG binds to FcRn in Placenta and is *Actively Transported into Neonatal Blood.
Opsonization and Phagocytosis
Major mechanism of defense against Encapsulated Bacteria.
**Spleen contains large numbers of Phagocytes that clear Opsonized Bacteria.
**Splenectomy predisposes to infection by these bacteria.
Antibody-Dependent Cellular Toxicity (ADCC)
**NK cells bind and lyse Antibody-coated Host Target cells.
They have Fc-gammaRIII Receptor (CD16), which binds to IgG bound to Host target cells and *Promotes ADCC.
(Marker for NK cells)
*How often Infected cells express surface molecules recognized by Antibody is Unknown.
