Adaptive Immunity: B-Cell Mediated Immunity III

Adaptive Immunity: B-Cell Mediated Immunity III

• Antibodies are soluble proteins produced by plasma cells.
• They are found in blood, lymph, and mucosal surfaces.
• Antibodies bind to extracellular pathogens/antigens.
• Antibodies directly disable pathogens/antigens and mark them for destruction by complement and/or phagocytes.
• Each antibody is specific, binding to one antigen or a small number of very similar antigens.

Typical Antibody Structure

• Antibodies have four polypeptide chains: two heavy and two light chains.
• Disulfide bonds link the chains.
• Light chains are either κ or λ.
• The N-terminus is at the top of the structure.
• The C-terminus is on the bottom.
• Antigen-binding sites are formed by the variable regions (VL + VH) of the light and heavy chains, respectively.
• The variable region's amino acid sequence determines antibody diversity and specificity.
• The hinge region is flexible, allowing for diverse antigen-binding abilities.
• Proteolytic cleavage can separate the antibody into Fab (two antigen-binding fragments) and Fc (crystallizable) fragments.
• Fc fragments are crucial for cell-receptor (FcR) binding and complement activation.

Immunoglobulin Domains

• Immunoglobulin domains are compact, stable, folded structures.
• They consist of approximately 100-110 amino acids.
• Two β-sheets connect via loops.
• Disulfide bonds and hydrophobic interactions stabilize the folding.

Variable Domains

• Variable domains have hypervariable regions (HVs) that contribute to antigen-binding specificity.
• Three HVs (complementarity-determining regions, CDRs) per light chain variable (VL) and heavy chain variable (VH) domain.
• These form the antibody's antigen-binding site.
• Framework regions (FRs) are structurally stable regions in the V domains.

Epitope (Antigenic Determinant)

• Epitopes are parts of an antigen that an antibody binds to.
• Antigens can be multivalent, having multiple epitopes, or multiple copies of the same epitope.
• Epitopes can be linear (contiguous amino acids) or discontinuous (noncontiguous amino acids).

Antigen-Binding Site Shapes

• Antibody binding sites can be pockets (for small, compact epitopes), grooves (for short, linear epitopes), or extended surfaces (for curved, globular epitopes).

Heavy Chain Constant Domains

• Heavy chains have constant (C) domains (either γ, μ, δ, α, or ε).
• These domains differ in location of disulfide bonds, carbohydrates, presence/absence of hinge regions and other structural characteristics.
• Antibodies can be IgG, IgM, IgD, IgA, or IgE, depending on the constant heavy chain region.

Antibody Affinity

• Affinity is the binding strength between an antibody and an antigen.
• Antibody affinity increases during the adaptive immune response because of somatic hypermutation.
• Antibody affinity maturation leads to stronger binding during the immune response.
• Affinity is determined by non-covalent attractions such as van der Waals forces, hydrophobic interactions, charge interactions, and hydrogen bonds, combined for a total binding affinity.

From BCR to Secreted Antibodies

• Secreted antibodies are produced from the same heavy-chain genes, with differences in RNA processing.
• This alternative processing allows for either membrane-bound or secreted antibodies.

IgM

• IgM is the first antibody class produced and secreted.
• IgM is found in blood and lymph.
• IgM has low affinity but high avidity (overall strength of binding to multiple epitopes on an antigen).

Secreted IgM

• Secreted IgM is a pentamer, containing 10 antigen-binding sites.
• Large size prevents easy entry to tissues but it has efficient complement activation properties.

IgG

• IgG is the most abundant blood-borne antibody.
• IgG has 2 high-affinity antigen-binding sites.
• IgG easily enters infected tissues.

Antibody Classes & Subclasses

• Each antibody class has different functions and properties based on the heavy chain constant regions.
• The tables provided in the notes (from pages 2-5) provide further details.

Dimeric IgA

• Produced by plasma cells in mucosal-associated lymphoid tissue (MALT).
• Found in gut, milk, tears, and saliva.
• Plays a crucial role in maintaining commensal populations, and prevents pathogen entry at mucous membrane locations.

Pentameric IgM & Dimeric IgA (Passive Immunity)

• Protect against microbes or toxins at mucosal surfaces and in the bloodstream.
• Maternal IgG crosses the placenta to provide passive immunity to the fetus.
• Breast milk provides dimeric IgA to newborns via the intestines.

Neutralizing Antibodies

• Prevent pathogen attachment to body cells.
• High-affinity binding sites are essential for neutralizing antibodies.

C'-Activating Antibodies

• Classical complement activation pathway is initiated by antigen-antibody complexes.
• IgM is a potent activator due to its pentameric structure.
• Some IgG subclasses (IgG3 and IgG1) can also efficiently activate complement.

Immune Complexes (ICs)

• Formed when antibodies bind to soluble multivalent antigens.
• Immune complexes can activate the complement system, leading to their removal by phagocytes.
• Removal is important to avoid them lodging in the basement membrane of small blood vessels, especially in the kidneys.

Circulating Immune Complexes in Kidney

• Immune complexes can damage kidney tissue (and other areas) through complement activation and deposition in glomeruli.

Anti-Parasite Defense Mechanisms

• Eosinophils and basophils play a role directly targeted at parasite destruction and elimination via IgE or IgG coating of the parasite.

Allergies & Asthma

• Allergens trigger a massive mast cell degranulation response that can lead to allergies and asthma.

Antibody Receptors (Fc Receptors)

• Fc receptors bind to the Fc region of antibodies.
• This binding can either activate effector cells or inhibit them in specialized immune situations.
• Some receptors are involved in antibody transcytosis or transport of antibodies across cells and tissues.

Polymeric Immunoglobulin Receptor (pIgR)

• Transports IgA and IgM across mucosal epithelial cells.
• This process is mediated by binding the J-chain on these molecules.
• The secretory piece of pIgR prevents antibody loss and allows them to function in the mucosal secretions.

FcyRI, FcyRII, and FcyRIII

• Activating Fc receptors for IgG are associated with cell activation, pathogen uptake & destruction by phagocytes; and antibody-dependent cell-mediated cytotoxicity (ADCC).
• Inhibiting Fc receptors of the IgG family moderate inflammatory response from effector cells.

Flow Cytometry

• Differentiates cells based on surface molecules and cytoplasmic structures that are labeled with fluorescent antibodies.
• Antibodies are used to tag particular cell surface molecules, which are identified and quantified by fluorescence labeling.

Evolution of Monoclonal Antibodies

• Monoclonal antibodies are produced through hybridoma technology.
• There are advancements in the development of chimeric, humanized and fully human forms.

Immunoassays

• Used for detection and quantitation of specific antigens.
• Immunoassays are based on antibody-antigen reactions.
• Examples include ELISA, lateral flow assays, and Western blotting.

Polyclonal Antisera vs. Monoclonal Antibodies

• Polyclonal antisera are mixtures of antibodies, each recognizing different epitopes on an antigen.
• Monoclonal antibodies are identical, specific for a single epitope.

Monoclonal Antibody Production

• A process used to produce a single type of antibody for a specific epitope.