Dr. Nabil MTIRAOUI, M.Sc, Ph.D

1. Antigens and Immunogens Properties and Characteristics Dr. Nabil MTIRAOUI, M.Sc, Ph.D Lecture 7

2. Definition and Properties

3. Introduction • In 1899 Ladislas Deutsch named the hypothetical substances halfway between bacterial constituents and antibodies "substances immunogens or antigens". He originally believed those substances to be precursors of antibodies, just like zymogen is a precursor of zymase. But by 1903 he understood that an antigen induces the production of immune bodies (antibodies) and wrote that the word antigen was a contraction of "Antisomatogen.

4. Definition of Immunogen and Antigenicity • Immunogensare substances that generate immune response • Antigenicity is the ability to combine specifically with the final products of the immune responses ( antibodies and/or cell surface receptors).

5. Definition of Antigen • Antigenis substance which when introduced parentally into the body stimulates the production of an antibody with which it reacts specifically and in an observable manner • simple to macromolecules e.g. carbohydrates, phospholiplids,nucleic acids and proteins.

6. Definition of Hapten • Haptens are small molecules which could never induce an immune response unless coupled to a carrier molecule. • e.g. dinitrophenyl, aminobenzene sulphonate, arsonate • Haptein-carrier molecule, unlike free haptein, can acts as an immunogen.

7. Definition of Epitope • An epitope is the small site on the antigen which is recognized by the antibody. • Usually between one and six sugars or amino acids on the surface of the antigen.

8. Definition of Epitope • Antigenic determinant is a cluster of epitopes on the surface of an antigen. • Antigen has several determinants each structurally different from each other. • A monoclonal antibody reacts with only one determinant on the same antigen.

9. Definition of Epitope

10. Substance, which when mixed with an antigen, enhances the magnitude and duration of the immune response • Functions: • Prolong retention of immunogen • Increase the effective size of the immunogen • Stimulate local influx of macrophages or immune cells to the injection site Definition of Adjuvant

11. Classification of Antigens

12. Classification of Ag : Based on Immunogenicity • Complete antigen : Substances which can induce antibody formation by themselves and can react specifically with these antibodies • Incomplete antigen (haptens): substances unable to induce antibody formation on its own but can become immunogenic when covalently linked to proteins, called carrier proteins.

13. Classification of Ag : based on origin • Exogenous antigens Exogenous antigens are antigens that have entered the body from the outside, for example by inhalation, ingestion, or injection. The immune system's response to exogenous antigens is often subclinical. • Endogenous antigens Endogenous antigens are antigens that have been generated within previously normal cells as a result of normal cell metabolism, or because of viral or intracellular bacterial infection.

14. Classification of Ag : based on origin • Auto-antigens An autoantigen is usually a normal protein or complex of proteins (and sometimes DNA or RNA) that is recognized by the immune system of patients suffering from a specific autoimmune disease. These antigens should, under normal conditions, not be the target of the immune system, but, due to mainly genetic and environmental factors, the normal immunological tolerance for such an antigen has been lost in these patients.

15. Types of Antigens

16. Types of Antigens • T-independent Antigens • Can directly stimulate B cells to produce antibodies • Polysaccharides in general • Generally more resistant to degradation  persist for longer periods of time  continue to stimulate immune system • T-dependent Antigens • Do not directly stimulate antibody production; need help of T cells • Usually proteins

17. T-independent Antigen • Polysaccharides • Properties • Polymeric structure • Polyclonal B cell activation • Resistance to degradation • Examples • Pneumococcal polysaccharide, lipopolysaccharide • Flagella

18. T-dependent Antigen • Proteins • Structure • Examples • Microbial proteins • Non-self or Altered-self proteins

19. Types of Antigens • Bacterial Antigens: these may be: • Soluble Ags: which are products excreted into the environment e.g. exotoxins, enzymes, haemolysin, ect.. • Cellular Ags: which correspond to the different structure of the bacterial cell e.g. • Capsular Agsin capsulated organisms. • Flagellaror HAgs in flagellated(motile)organisms. • Somaticor O Ags are parts of the cell wall of gram negative bacteria. • Virulenceor ViAgs are surface Ags. • FimbrialAgs are surface Ags in fimriated gram negative bacteria.

20. Types of Antigens • Viral Antigens: these may be: • Protein coat viral Ags. • Soluble Ags: which diffuse in the surrounding fluids during virus growth e.g. soluble nucleoprotein as in influenza and mumps viruses.

21. Types of Antigens • Human tissue antigens (isoantigens): • Blood group Ags: A and B as well as RhAgs on red cells. These are important in blood transfusion reactions. • HistocompatibilityAgs: these are glycoprotein molecules present on membranes of tissue cells. They are called the major histocompatibility complex (MHC) Ags or human leucocyteAgs (HLA). There are 2 classes of MHC: class1 MHC are present on all nucleated cells, class2 MHC are present on immunocompetent APC.

22. Antibody-Antigen Interactions

23. Antibody-Antigen Interactions • Binding of antibody to antigen is dependent on • hydrogen bonds, electrostatic attractions and Van der Waals attractions. • These bonds are weak compared to covalent bonds but the large number of weak bonds result in a stable complex. • Antibody-antigen binding is reversible.

24. Antibody-Antigen Interactions • Avidity is the strength of binding two molecules or cells to one another at multiple sites. It is determined by • heterogeneity of antibodies in serum • heterogeneity of antigenic determinants • Affinity– measure of the functional affinity of an antiserum for the whole antigen. • Cross-reactivity is due to antiserum reacting with a partially related antigen.

25. Low Affinity High Affinity Ab Ab Ag Ag Antibody-Antigen Interactions: Affinity • Strength of the reaction between a single antigenic determinant and a single Ab combining site Affinity = attractive and repulsive forces

26. Y Y Y Y Y Y Y 104 106 1010 Keq = Avidity Affinity Avidity Antibody-Antigen Interactions: Avidity • The overall strength of binding between an Ag with many determinants and multivalent Abs

27. Cross reactions Anti-A Ab Anti-A Ab Anti-A Ab Ag B Ag C Identical epitope Similar epitope Ag A Antibody-Antigen Interactions: Cross Reactivity • The ability of an individual Ab combining site to react with more than one antigenic determinant. • The ability of a population of Ab molecules to react with more than one Ag

28. Properties of Antigens

29. Chemical Nature of Immunogens • Proteins • Polysaccharides • Nucleic Acids • Lipids • Some glycolipids and phosopholipids can be immunogenic for T cells and illicit a cell mediated immune response

30. What Does The B Cell Ig Receptor Recognize? • Proteins (conformational determinants, denatured or proteolyzed determinants) • Nucleic acids • Polysaccharides • Some lipids • Small chemicals (haptens)

31. What Does the αβT Cell Receptor (TCR) Recognize? • Only fragments of proteins (peptides) associated with MHC molecules on surface of cells • Th recognize peptide associated with MHC class II molecules • Tc recognize peptide associated with MHC class I molecules

32. Fe Antigenic Determinants : Recognized by B cells and Ab • Composition • Proteins, polysaccharides, nucleic acids • Sequence (linear) determinants • Conformational determinants • Size • 4-8 residues • Number • Limited (immunodominantepitopes) • Located on the external surfaces of the Ag

33. Antigenic Determinants : Recognized by T cells • Composition • Proteins (some lipids) • Sequence determinants • Processed • MHC presentation (lipid presentation by MHC-like CD1) • Size • 8 -15 residues • Number • Limited to those that can bind to MHC

34. Conventional Antigen Antigen presenting cell α2 β2 α2 β2 CHO CHO MHC Class II α1 β1 α1 β1 CHO CHO CHO CHO Super antigen Antigen αV αV βV βV CHO CHO CHO CHO T cell receptor αC βC αC βC CHO CHO CHO CHO T lymphocyte Superantigen

35. Superantigens • Proteins produced by pathogens • Not processed by antigen presenting cells • Intact protein binds to variable region of β chain on TCR of T cells and to MHC class II on antigen presenting cells (APC) • Large numbers of activated T cells release cytokines having pathological effects

36. Superantigens • Definition • Examples • Staphylococcal enterotoxins • Staphylococcal toxic shock toxin • Staphylococcal exfoliating toxin • Streptococcal pyrogenicexotoxins

37. Antigen presenting cells (APC) • Cells with the capacity to capture, process and present antigenic peptides to T cells • Antigens are presented in the context of MHC class I or II • Also deliver co-stimulatory signal (signal II) to T cells leading to proper activation • Only APCs can activate a naïve T cell • Dendritic cells, Macrophages, B cells

38. MHC Class I and Class II • MHC I on all cellsMHC II on APC • Bind Ag  only small peptides • As an individual you make a small number of different kinds of MHC I and MHC II • Encoded by stable genes inherited; NOTgenerated by rearrangements • But in the population there are lots of genetic variants of MHC I and MHC II • Important in transplants • Hence the name  “Major Histocompatability Complex”

39. T cells • T cell receptors (TCR) – Ag specific • Glycoproteins CD4 (helper T cells) or CD8 (cytotoxic T cells) CD4 CD8 TCR TCR CD8 T cell or cytotoxic T cell CD4 T cell or helper T cell

40. All T cells are “Antigen specific” • Mediated by “T cell receptor”  TCR • Surface molecule analogous to part of Ab • Diversity is generated by rearrangement of TCR gene locus

41. TCR Recognizes its Epitope Only in the Context of MHC • CD4 TCR – peptide/MHC Class II • CD8 TCR – peptide/MHC Class I

42. CD4+ T cells • T cells with CD4 marker (glycoprotein) • 70% of T cells in the periphery • T helper cells • Play central role in modulating cellular immunity via secretion of cytokines that modulate: • B cell activation • Immunoglobulin secretion (quality) • Macrophage and dendritic cell activation • Cellular chemotaxis and inflammation • Th1 versus Th2 cells

43. CD4+ T cells • Helper T cells involved in Ab production • Recognition of “exogenous Ag” • Bacteria • Extracellular Ag • Recognize MHC class II molecule • Present on “antigen presenting cells” = APC • e.g. Macrophages, Dendritic Cells, B cells

44. CD8+ T cells • Cytotoxic T cells  cell killing • Recognition of “endogenous Ag” • Virus infected cells • Cancerous cells • Recognize MHC class I molecule • Present on all cells

45. Thank You