antigen antibody interaction

• Antigen-Antibody interaction


  The noncovalent interactions that form the basis of antigen-antibody (Ag-Ab) binding include hydrogen bonds, ionic bonds, hydrophobic interactions, and van der Waals interactions (non-covalent)
• Strength of Antigen-Antibody Interactions


  • These interactions are individually weak (compared with a covalent bond), a large number of such interactions are required to form a strong Ag-Ab interaction
  • Each of these noncovalent interactions operates over a very short distance, generally about 1×10^7 mm (1 angstrom, Å)
  • A strong Ag-Ab interaction depends on a very close fit between the antigen and antibody
  • Requires a high degree of complementarity between antigen and antibody
• Avidity


  The overall strength of multiple interactions between a multivalent antibody and antigen (multiple epitopes)
• Avidity is a better measure of an antibody's binding capacity within biological systems than the affinity of its individual binding sites
• High avidity can compensate for low affinity
• Cross-Reactivity


  Antibody elicited by one antigen can cross-react with an unrelated antigen if they share an identical or very similar epitope
• Cross-Reactivity


  • Polysaccharide antigens that contain similar oligosaccharide residues
  • ABO blood-group antigens which are glycoproteins expressed on red blood cells
• Subtle differences in the terminal residues of the sugars attached to these surface proteins distinguish the A and B blood-group antigens
• An individual lacking one or both of these antigens will have serum antibodies to the missing antigen(s)
• Antibodies produced to streptococcal M antigens have been shown to cross-react with several myocardial and skeletal muscle proteins and have been implicated in heart and kidney damage following streptococcal infections
• Precipitation Reactions


  Antibody and soluble antigen interacting in aqueous solution form a lattice that eventually develops into a visible precipitate
• Precipitation Reactions


  • Antibodies that aggregate soluble antigens are called precipitins
  • Formation of the visible precipitate occurs more slowly and often takes a day or two to reach completion
  • Formation of an Ag-Ab lattice depends on the valency of both the antibody and antigen: the antibody must be bivalent, the antigen must be either bivalent or polyvalent
• Precipitation Reactions in Gels

  When antigen and antibody diffuse toward one another in agar, or when antibody is incorporated into the agar and antigen diffuses into the antibody-containing matrix, a visible line of precipitation will form
• Immunodiffusion


  An analytic technique in which reactants diffuse to intermingle with each other and react immunologically
• Types of Immunodiffusion Reactions

  • Radial immunodiffusion (the Mancini method)
  • Double immunodiffusion (the Ouchterlony method)
• Radial Immunodiffusion


  1. An antigen sample is placed in a well and allowed to diffuse into agar containing a suitable dilution of an antiserum

  2. As the antigen diffuses into the agar, the region of equivalence is established and a ring of precipitation, a precipitin ring, forms around the well

  3. The area of the precipitin ring is proportional to the concentration of antigen

  4. By comparing the area of the precipitin ring with a standard curve, the concentration of the antigen sample can be determined
• Ouchterlony Method


  1. Both antigen and antibody diffuse radially from wells toward each other, thereby establishing a concentration gradient

  2. As equivalence is reached, a visible line of precipitation, a precipitin line, forms
• Immunoelectrophoresis


  The antigen mixture is first electrophoresed to separate its components by charge, then antibody and antigen diffuse toward each other and produce lines of precipitation where they meet in appropriate proportions
• Immunoelectrophoresis is used in clinical laboratories to detect the presence or absence of proteins in the serum
• Immunoelectrophoresis is a strictly qualitative technique that only detects relatively high antibody concentrations (greater than several hundred g/ml)
• Agglutination


  The interaction between antibody and a particulate antigen results in visible clumping called agglutination
• Agglutination reactions are similar in principle to precipitation reactions; they depend on the crosslinking of polyvalent antigens
• Prozone Effect


  An excess of antibody can inhibit agglutination reactions
• Prozone effects can be encountered in many types of immunoassays
• Causes of Prozone Effect

  1. At high antibody concentrations, the number of antibody binding sites may greatly exceed the number of epitopes, so most antibodies bind antigen only univalently instead of multivalently

  2. Polyclonal antibodies may contain high concentrations of antibodies that bind to the antigen but do not induce agglutination (incomplete antibodies, often of the IgG class)
• Agglutinins


  Reactions that cause agglutination
• Agglutination reactions


  Similar in principle to precipitation reactions, depend on the crosslinking of polyvalent antigens
• Prozone effect


  Inhibition of agglutination reactions when there is an excess of antibody
• Prozone effect


  1. High antibody concentrations lead to most antibodies binding antigen univalently instead of multivalently

  2. Univalent antibodies cannot crosslink one antigen to another
• Prozone effects are readily diagnosed by performing the assay at a variety of antibody (or antigen) concentrations
• Incomplete antibodies


  Antibodies that bind to the antigen but do not induce agglutination, often of the IgG class
• At high concentrations of IgG, incomplete antibodies may occupy most of the antigenic sites, blocking access by IgM, which is a good agglutinin
• Lack of agglutinating activity of incomplete antibodies

  • May be due to restricted flexibility in the hinge region, making it difficult for the antibody to assume the required angle for optimal cross-linking of epitopes
  • May be due to the density of epitope distribution or the location of some epitopes in deep pockets of a particulate antigen, making it difficult for the antibodies to agglutinate
• The solution to problems with incomplete antibodies is to try different antibodies that may react with other epitopes of the antigen that do not present these limitations
• Hemagglutination


  Agglutination reactions used in blood typing
• Blood typing


  1. RBCs are mixed on a slide with antisera to the A or B blood-group antigens

  2. If the antigen is present on the cells, they agglutinate, forming a visible clump on the slide

  3. Determination of which antigens are present on donor and recipient RBCs is the basis for matching blood types for transfusions
• Bacterial agglutination


  Used to diagnose infection
• Bacterial agglutination diagnosis


  1. Serum from a patient is serially diluted in an array of tubes to which the bacteria is added

  2. The agglutinin titer of the antiserum can be used to diagnose a bacterial infection
• Patients with typhoid fever show a significant rise in the agglutination titer to Salmonella typhi
• Agglutination reactions also provide a way to type bacteria, for instance different species of the bacterium Salmonella can be distinguished by agglutination reactions with a panel of typing antisera