Antigen-Antibody Interactions: Specificity, Types, and Applications

Posted on May 5, 2024 in Biology

Antigen-Antibody Interactions: Specificity

Antigen-antibody interactions exhibit high specificity due to the complementary shapes of the antigen-binding site (paratope) on the antibody and the specific epitope of the antigen.

Types of Bond Interactions

The interactions between a paratope and an epitope involve noncovalent forces, resulting in reversible binding. These forces include:

  • Hydrogen Bonds: Formed between oppositely charged or polar groups containing hydrogen atoms.
  • Electrostatic Interactions: Attractions between oppositely charged ionic groups.
  • Van der Waals forces: Weak, short-range attractions between non-polar regions.
  • Hydrophobic Interactions: The tendency of non-polar groups to cluster together, avoiding water molecules.

Affinity and Avidity

  • Affinity: The strength of the interaction between a single antigen-binding site and its corresponding epitope. High-affinity antibodies bind more strongly to their antigens.
  • Avidity: The overall strength of binding of a multivalent antibody to an antigen with multiple epitopes. Avidity depends on both the affinity of individual binding sites and the valency (number of binding sites) of both the antibody and the antigen.

Types of Antigen-Antibody Interactions

  1. Precipitation Reactions: Soluble antigens cross-link with antibodies, forming large, insoluble complexes (precipitates).
  2. Agglutination Reactions: Antibodies bind to cellular antigens (e.g., bacteria, red blood cells), causing cell clumping.
  3. Complement Fixation: Binding of specific antibody classes (IgM, IgG) to an antigen triggers the complement cascade, leading to various immune outcomes.
  4. Neutralization: Antibodies block the activity of toxins or pathogens by binding to critical functional sites.
  5. Opsonization: Antibodies “tag” a pathogen, enabling recognition and engulfment by phagocytes.

Applications of Antigen-Antibody Interactions

  • Diagnostic Tests: Various lab techniques utilize antigen-antibody interactions for detecting pathogens, antibodies, or other molecules in biological samples (e.g., ELISA, Western blot, immunofluorescence).
  • Therapeutic Antibodies: Monoclonal antibodies engineered to recognize specific antigens on cancer cells or pathogens are used for targeted therapies.

2.3.1 Precipitation Reactions

Principle

Precipitation reactions occur when soluble antigens interact with specific antibodies, creating large, insoluble complexes that become visible as precipitates.

Mechanism

  1. Antibody Structure: Precipitation is primarily driven by IgG and IgM antibodies. Their Y-shaped structure with two identical antigen-binding sites (Fab regions) allows them to bind multiple antigens simultaneously.
  2. Antigen Multivalency: Soluble antigens often possess multiple identical or similar epitopes, enabling cross-linking by multiple antibodies.
  3. Lattice Formation: Antibodies bridge together multiple antigens, forming an extensive three-dimensional network held together by noncovalent interactions.
  4. Critical Mass and Insolubility: As the lattice grows, its size, complexity, and molecular weight increase, leading to reduced solubility and visible precipitation.

Factors Influencing Precipitation

  1. Zone of Equivalence: Maximal precipitation occurs when antigens and antibodies are present in an optimal ratio, allowing for the largest possible lattice formation. Excess of either component can hinder precipitation.
  2. Antibody Characteristics:
    • Isotype: IgG and IgM, due to their multivalency, are most effective.
    • Affinity: High-affinity antibodies form stronger, more stable interactions within the lattice.
  3. Electrolytes and pH: The presence of electrolytes (salts) and the pH of the solution can affect the solubility of immune complexes and influence precipitation reactions.

Types of Precipitation Reactions

  1. Tube Precipitin Test: Quantitative; measures approximate antigen or antibody concentration via serial dilutions.
  2. Gel Immunodiffusion: Performed in a semi-solid medium (agarose) allowing for visualization of precipitin bands:
    • Single diffusion (Ouchterlony): Analyzes a single component (antigen or antibody)
    • Double diffusion: Analyzes both antigen and antibody, useful for comparing multiple antigens.

Applications

  • Diagnostics: Historically used for:
    • Detecting specific pathogens or their antigens
    • Identification of antibodies in a patient’s serum
    • Quantifying antigen or antibody levels
  • Research:
    • Studying the structure and composition of antigens
    • Characterizing antibody specificity and affinity
    • Forensic applications (in the past)

2.3.2 Agglutination Reactions

Principle

Agglutination reactions occur when antibodies bind to antigens present on the surface of cells or particles, leading to visible cross-linking and the formation of clumps (agglutinates).

Mechanism

  1. Antibody Binding: Antibodies, primarily IgM and sometimes IgG, bind to multiple, repeating epitopes expressed on the surface of cells or other large particles.
  2. Cross-Linking: The bivalent (or multivalent, in the case of IgM) nature of antibodies allows them to bridge multiple particles, forming a network. The size of the agglutinate depends on the number of cells or particles linked together.
  3. Visible Clumping: As the lattice-like network of antibodies and particles grows, the clumps become large enough to be visible to the naked eye or with simple microscopy.

Types of Agglutination Reactions

  • Direct Agglutination: Involves the direct interaction between antibodies and cellular antigens. Examples include:
    • Blood typing: Antibodies against blood group antigens (A, B, Rh) cause clumping of respective red blood cells.
    • Bacterial agglutination tests for the identification of specific bacteria.
  • Indirect (Passive) Agglutination: Antibodies are first attached to inert particles (e.g., latex beads). If the patient’s sample contains antigens that correspond to those antibodies, visible agglutination of the antibody-coated particles occurs.

Factors Affecting Agglutination

  • Antigen-Antibody Ratio: As with precipitation, optimal concentrations are needed. Excess antibody can sometimes inhibit agglutination, known as the prozone effect.
  • Antibody Characteristics:
    • Isotype: IgM, due to its pentameric structure and thus higher valency, is generally a more potent agglutinin than IgG.
    • Affinity: Higher affinity antibodies lead to stronger cross-linking and greater agglutination.
  • Time and Temperature: Optimal incubation time and temperature can influence the rate and extent of agglutination.

Applications

  • Diagnostics
    • Blood typing and crossmatching before blood transfusions.
    • Detection of specific antibodies or antigens in patient samples for diagnosis of infections (e.g., Widal test for typhoid fever).
    • Rapid diagnostic tests (e.g., antibody-coated latex beads for detecting specific pathogens).

Complement Fixation

Principle

Complement fixation refers to the consumption of complement proteins by antigen-antibody complexes. It primarily involves the activation of the classical complement pathway, initiated by specific antibody isotypes (IgM and certain subclasses of IgG) when they bind to antigens.

Mechanism

  1. Antibody Binding: IgM or specific IgG subclasses bind to their corresponding antigens, forming immune complexes.
  2. C1 Complex Activation: The Fc region of the bound antibodies interacts with and activates the C1 complex (composed of C1q, C1r, and C1s), the first component of the classical complement pathway.
  3. Complement Cascade: The activated C1 complex initiates a proteolytic cascade, cleaving and activating subsequent complement proteins (C4, C2, C3, etc.).
  4. Complement Consumption: As the cascade progresses, complement components are either deposited on the target surface or form active fragments with potent effector functions. This process “consumes” active complement proteins, reducing their availability in the surrounding fluid.

Complement Fixation Test

  • Historical and Diagnostic Tool: The complement fixation test historically was used to detect specific antibodies or antigens in a sample. It relies on a two-stage indicator system:
  1. Stage 1 (Test System): Patient sample (potentially containing antibodies) is combined with a known antigen and a fixed amount of complement. If specific antibodies are present, they form complexes that consume complement.
  2. Stage 2 (Indicator System): Sheep red blood cells (SRBCs) coated with antibodies against SRBCs are added.
    • If complement was consumed in Stage 1, there is insufficient complement left to lyse SRBCs. This indicates a positive test for the antibodies present in the patient sample.
    • If no antibodies were present in the patient sample, complement is left intact and causes SRBC lysis, resulting in a negative test.

Applications

  • Serology: Historically used to diagnose various infectious diseases (syphilis, some viral and fungal infections) by detecting the presence of specific antibodies in patient serum.

Limitations and Modern Techniques

  • Complexity: Complement fixation tests are technically demanding and require careful standardization
  • Alternatives: Largely replaced by more sensitive and specific immunodiagnostic techniques such as ELISA, immunofluorescence, and Western blot.