Immunology: Antibodies and Antigens

Antibodies

• Antibodies are proteins that bind specifically to antigens.
• Antibodies belong to the immunoglobulins (Igs) class of proteins.
• Antibodies are secreted by differentiated B cells known as plasma cells.

Antigens and Immunogens

• Antigens are molecules that can bind to antibodies or generate peptide fragments recognized by T cell receptors.
• Immunogens are substances that elicit an adaptive immune response.
• All antigens have the potential to elicit specific antibodies, but some need to be attached to an immunogen.
• All immunogens are antigens, but not all antigens are immunogenic.

Immunogenicity Requirements

• Foreignness
• High molecular weight
• Chemical complexity
• Degradability

Haptens

• Small organic molecules with simple structures that fail to induce immune responses.
• An immune response can be provoked if a hapten is attached to a protein carrier.

Epitope, Paratope, and Antigen

• Epitope: The specific portion of an antigen that binds to an antibody or T-cell receptor.
• Paratope: The part of an antibody that recognizes and binds to the epitope (antigen binding site).

Antibody Molecule Fragments

• Antibodies can be cleaved into functionally distinct fragments by proteolytic enzymes.
• Two identical Fab (antigen binding) fragments
• One Fc (crystallisable) fragment

Antibody Structure

• Antibodies are composed of two types of protein chains:
  • Heavy chains (2)
  • Light chains (2)
• The antigen binding region varies considerably between antibody molecules and is known as the variable or V region. The V region determines antigen specificity.
• The constant region, or C region, is less variable and determines the effector functions of the antibody.

Immunoglobulin Classes

• Five different immunoglobulin classes: IgM, IgD, IgG, IgA, and IgE.
• Nine different isotypes.

Human Antibody Properties

• IgM:
  • The predominant antibody early in an immune response.
  • It has 10 potential antigen-binding sites.
  • Most effective antibody at agglutinating bacteria and activating complement.
• IgD:
  • Involved in B cell activation.
  • Serum function is unknown.
• IgG:
  • The most prevalent antibody in serum.
  • Able to cross the placenta to allow maternal protection of newborns.
• IgE:
  • Evolved to protect against parasitic infections.
  • Involved in allergic reactions.
• IgA:
  • Present in serum and seromucous secretions.
  • IgA1 is predominant in serum (over 80% is a monomer).
  • IgA2 is the predominant antibody in seromucous secretions (saliva, colostrum, tracheobronchial, genitourinary secretions) and is a dimer.

Antibody Variations

• Isotype: Variation in the constant region of the heavy chain that results in different antibody classes (IgG, IgM, IgA, IgD, IgE).
• Allotype: Variation between individuals due to minor sequence differences in the heavy or light chains of antibodies.
• Idiotype: Variation in the antigen binding region of different antibodies.

Antigen-Antibody Interactions

• Affinity: The strength of the interaction between a single antigen binding site (paratope) and its epitope.
• Valence: The number of different molecules that an antigen or antibody can combine with at one time.
• Avidity: The strength of the interaction between multivalent antibodies and multivalent antigens.

Antigen-Antibody Bonds

• Antibodies form multiple non-covalent bonds with antigens:
  • Hydrogen bonds
  • Electrostatic bonds
  • Van der Waals forces
  • Hydrophobic forces
  • Cation-pi interactions

Specificity, Cross-Reactivity, and Non-Reactivity

• Reactions can show a high level of specificity.
• Reactions can also be cross-reactive, binding to structurally related but different antigens.

Immune Response and Antibody Secretion

• Seroconversion: The phase of an infection when antibodies against the infecting agent are first detectable in the blood.
• Primary Response: The first exposure to an antigen; slow, sluggish, short-lived, and low levels of antibodies.
• Secondary Response: Second exposure to an antigen; faster, stronger, and longer-lasting response with higher antibody levels.

Clonal Selection Hypothesis

• Antigens activate specific clones of lymphocytes.
• This hypothesis explains how the immune system generates a highly diverse range of antibodies to target specific pathogens.

Antibody Repertoire Development

• The immune system generates billions of different antibodies with a limited number of genes.
• The diversity of the antibody repertoire is generated by:
  • Somatic recombination
  • Somatic hypermutation
  • Class switching

V(D)J Recombination

• Somatic recombination involves the rearrangement of gene segments within immunoglobulin during B cell development, in the absence of antigen.
• The variable (V) region of an immunoglobulin heavy and light chain is encoded by more than one gene segment.

Somatic Hypermutation

• Somatic hypermutation is the random mutation that occurs in the V region and diversifies the antibody repertoire.
• This process alters the affinity of the antibody for antigens.
• It occurs at the same time as class switching and both processes involve the enzyme activation induced cytidine deaminase (AID).

Affinity Maturation

• Affinity maturation results from somatic hypermutation and increases antibody affinity for an antigen.
• B cell receptors are selected for progressively higher affinity during an immune response

Class Switching

• Class switching involves the C region of the heavy chain only and increases the functional diversity of antibodies.
• Antigen specificity is retained.
• Cytokines and T cells play a major role in class switching.
• Isotype switching is greatly affected by the tissue environment, partly due to different cytokines released by T cells.

Antibody Applications

• Diagnostics:
  • Detection of infections
  • Measurement of biological markers
  • Recognition of allergies
• Therapeutics:
  • Treatment of cancer, infectious diseases, and autoimmune diseases.
• Research:
  • Immunohistochemistry
  • Western blots
  • Flow cytometry

Polyclonal Antibodies

• Antibodies generated in a natural immune response or after immunization are polyclonal.

Monoclonal Antibodies

• Monoclonal antibodies are produced by hybridoma technology or recombinant DNA technology.
• Hybridoma technology: Fusion of an immortal myeloma cell with an antibody-producing B cell.
• Recombinant technology: Synthetic antibodies or antibody fragments generated by recombinant DNA technology.

Antibody Levels After Immunization

• After immunization, IgM levels increase first.
• With repeated immunization, IgG levels increase.

Learning Objectives

• Describe the basic structure of antibodies.
• Understand the functions and properties of antibodies.
• Define key terms related to immunology: antibody, antigen, epitope, paratope, immunogen, hapten, affinity, avidity, valency, idiotype, isotype, allotype.
• Discuss antigen-antibody interactions.
• Discuss the generation of the antibody repertoire.
• Discuss the use of antibodies in therapeutics and diagnostics.
• Discuss the preparation of monoclonal antibodies.