1. Lecture 14 Antibody-Antigen Reactions

2. Binding of the epitope in the antigen binding site POOR FIT GOOD FIT antibody combining site antigen determinant high attraction low repulsion high repulsion low attraction

3. Antibody Avidity Multiple interactions between antigen binding sites and epitopes Multiple interactions between antigen binding sites and epitopes Greater than additive Greater than additive Similar to Velcro™ Similar to Velcro™ More relevant to biologic systems than affinity More relevant to biologic systems than affinity

4. Effect of multivalent interactions antibodyFabIgGIgGIgM effective antibody valence 112 up to 10 antigen valence 11nn equilibrium constant (L/M) 10 4 10 4 10 7 10 11 advantage of multi-valence --10 3 -fold10 7 -fold definition of bindng affinityaffinityavidityavidity intrinsic affinity functional affinity

5. Biological Consequences of Antibody Affinity/Avidity Neutralization of toxins Neutralization of toxins Complement activation Complement activation Immune elimination of antigen Immune elimination of antigen Virus neutralization Virus neutralization More intense immune complex disease in animals More intense immune complex disease in animals higher levels of circulating antigen-antibody complexes higher levels of circulating antigen-antibody complexes more intense localization of immune complexes on basement membranes. more intense localization of immune complexes on basement membranes. more severe impairment of organ function more severe impairment of organ function

6. Cross-Reaction ABCD anti-A anti-A anti-B anti-B anti-C anti-C anti-D anti-D Immunize Immunize Immunogen Immunogen Cross-reactive antigen AEFG ABCD Reactivity with rabbit antiserum

7. Primary Interactions Between Antigen and Antibody Determined by the interaction between epitopes and antigen-binding sites Determined by the interaction between epitopes and antigen-binding sites Independent of biochemical and biologic phenomena that may follow Independent of biochemical and biologic phenomena that may follow Depend only on the quantity and quality (affinity/avidity) of antibodies Depend only on the quantity and quality (affinity/avidity) of antibodies The basis for ELISA, RIA, IFA assays The basis for ELISA, RIA, IFA assays

8. Enzyme (ELISA) Immunoassay Antigen anti-antigen antibody Enzyme conjugated to anti-Ig antibody (“second antibody”) Add substrate for enzyme

9. Immune Precipitation Antigen Antibody

10. Radial Immunodiffusion Ab-containing gel Ag/8 Ag/8 Ag/4 Ag/4 Ag/2 Ag/2 Ag/1 Ag/1 Precipitin Precipitin ring ring Antigen Concentration Antigen Concentration D 2 Ring

11. Agglutination RBC IgM Antibody IgG Antibody RBC

12. Complement activation A system of plasma proteins that interact with A system of plasma proteins that interact with Antigen/antibody complexes Antigen/antibody complexes Pathogen surface motifs (alternative and lectin pathways Pathogen surface motifs (alternative and lectin pathways Activation of complement results in Activation of complement results in Chemo-attraction of inflammatory cells Chemo-attraction of inflammatory cells Peptide mediators of inflammation (anaphylatoxins) Peptide mediators of inflammation (anaphylatoxins) Increased blood vessel permeability Increased blood vessel permeability Smooth muscle contraction Smooth muscle contraction Mast cell degranulation Mast cell degranulation Opsonization of pathogens (enhances phagocytosis) Opsonization of pathogens (enhances phagocytosis) Killing of pathogens (membrane attack complex) Killing of pathogens (membrane attack complex)

13. Overview of the Complement Cascade

14. Effector Actions of Complement

15. Terms to remember Affinity Affinity Agglutination Agglutination Avidity Avidity Chemotaxis Chemotaxis Complement Complement Classical pathway Classical pathway Alternative (properdin) pathway Alternative (properdin) pathway Cross-reaction Cross-reaction Epitope/Antigenic Determinant Epitope/Antigenic Determinant Opsonization Opsonization Precipitation Precipitation Second antibody Specificity Second antibody Specificity Valence Valence