14.Antigensantibodiesandthecomplementsystem.pptx

Microbiology

1. Chapter 14 Antigens, Antibodies and the Complement system Dr Sonal Saxena Dr Arpita Saxena

2. A. ANTIGENS An antigen is any substance that induces an immune response Stimulates antibody production in a specific and observable manner Both T and B cells recognise antigens Complete antigens evoke immune response independently Partial antigens (haptens) require additional carrier proteins to evoke immune response

3. Haptens Immunogenic when combined with a carrier protein (induces antibody production) but reacts specifically with antibodies Complex haptens Polyvalent Precipitate with specific antibodies Simple haptens Univalent Inhibit precipitation with specific antibody

4. DEFINITIONS Antigenicity: Property that allows a substance to combine specifically with antibodies and TCRs (T cell receptors) Immunogen : Induces detectable immune response Immunogenicity: Ability to induce immune response All immunogens are antigens but not all antigens are immunogens

5. DEFINITIONS Epitope or antigenic determinant Smallest unit of antigen Responsible for antigenicity Sequential or linear epitope Single linear segment of primary sequence Recognised by T cells Conformational epitope Formed during peptide chain folding Recognised by B cells Epitopes: (a) linear and (b) conformational

6. Paratope Area on the antibody corresponding to an epitope Epitopes and paratopes determine specificity Bacteria and viruses carry different epitope, presenting an antigenic mosaic Similar epitopes on different antigens leads to antigenic cross reaction Adjuvant: substances that enhance the immunogenicity of an antigen

7. DETERMINANTS OF ANTIGENICITY Foreignness: Non-self antigens elicit immune response Size: Larger molecules are highly antigenic Chemical nature: Antigens are proteins and polysaccharides Lipids and nucleic acids are less antigenic

8. DETERMINANTS OF ANTIGENICITY Susceptibility to tissue enzymes Substances not susceptible to tissue enzymes are not antigenic Other factors Dose, route and time of administration Excess dose: Immunological tolerance Intravenous route elicits more response

9. ANTIGENS Table 14.1 Examples of common antigens

10. TYPES OF SPECIFICITY OF ANTIGENS Basis of specificity : S tereochemistry and spatial configuration Types of specificity Species specificity Isospecificity Blood group — Human erythrocyte antigen Histocompatibility antigen — Transplantation

11. ANTIGENIC SPECIFICITY

12. ANTIGENIC SPECIFICITY

13. BIOLOGICAL CLASSES OF ANTIGENS T cell-dependent (TD) antigens T-B interaction and cytokines provide stimulus for B cell activation T cell-independent (TI) antigens Directly stimulate B cells to produce antibody Type I antigen: Endotoxin Type II antigen: B cells activated by cytokines

14. BIOLOGICAL CLASSES OF ANTIGENS Table 14.2 Comparison between T cell-dependent and T cell-independent antigens

15. SUPERANTIGENS A ntigens that evoke an excessive immune response. D o not get processed by an antigen presenting cell (APC). A ttach directly to the MHC-II α 1 domain and β variable chain of TCR . T wo types: E ndogenous: Intracellular (e.g., virus) Exogenous: Extracellular (e.g., bacteria)

16. SUPERANTIGENS A re potent CD4+ T cell activators Initiate the release of cytokines (IL-2) Action of superantigens - Bind outside antigen-binding groove on the TCR β chain - High levels of IL-2 are released - Symptoms of fever, nausea, vomiting, diarrhea - Excessive IL-2 leads endothelial damage - Intravascular coagulation, shock and multiple organ/system failure

17. SUPERANTIGENS Table 14.3 Human diseases associated with microbial superantigens

18. B. ANTIBODIES Antibodies are proteins: Immunoglobulins that react specifically to an epitope on an antigen Secreted by plasma cells Present on B cell membrane Act on antigens by effector functions like Phagocytosis Antibody-dependent cell-mediated cytotoxicity ( ADCC ) Opsonisation

19. IMMUNOGLOBULINS All antibodies are immunoglobulins but all immunoglobulins may not be antibodies Five classes of immunoglobulins — IgG, IgA, IgM, IgD , and IgE

20. ANTIBODY STRUCTURE S tructural components of immunoglobulins can be separated by enzyme digestion i ) Papain digestion : T wo types of fractions (three parts) Fc (fragment crystallisable ) Fab (fragment antigen binding) ii) Pepsin digestion : T wo Fab fragments held together in position: F ( ab )2 Fc portion: D igested by pepsin into smaller fragments

21. ANTIBODY STRUCTURE IMMUNOGLOBULIN CHAINS Table 14.4 Immunoglobulin classes and H chains Fig. 14.2 Structure of an immunoglobulin molecule with light and heavy chains

22. ANTIBODY STRUCTURE

23. IMMUNOGLOBULIN CLASSES Table 14.5 Characteristics of immunoglobulin classes

24. IMMUNOGLOBULIN CLASSES

25. IMMUNOGLOBULIN CLASSES IgA Serum Ig Secretory IgA (dimeric with J chain) Colostrum and breast milk are rich in IgA Fig. 14.3 Secretory IgA molecule

26. IgM Millionaire molecule ( pentamer ) Intravascular distribution First antibody to appear Earliest antibody synthesised by fetus Effective in opsonisation and agglutination Deficiency associated with septicemias Fig. 14.4 IgM molecule

27. IMMUNOGLOBULIN CLASSES

28. PROTECTIVE FUNCTIONS OF ANTIBODIES Phagocytosis Virus neutralisation Complement-mediated lysis of cells Antibody-mediated cell cytotoxicity Neutralisation of toxins

29. FACTORS INFLUENCING ANTIBODY PRODUCTION 1. Genetic factors: ‘Responder’ and ‘non-responder’ 2. Age 3. Immunocompetence 4. Nutritional status: Malnutrition affects immune response adversely

30. ABNORMAL IMMUNOGLOBULINS Multiple myeloma Bence-Jones proteins Light chain of immunoglobulin Heavy chain disease Cryoglobulinemia Precipitate is formed on cooling the serum which re-dissolves on warming Found in myelomas, macroglobulinemias SLE (systemic multiple sclerosis)

31. IMMUNOGLOBULIN SPECIFICITIES

32. IMMUNOGLOBULIN SPECIFICITIES Idiotype: Each individual antigenic determinant of the variable region (paratope) is referred to as an idiotope Immunisation with Fab fragments Anti-idiotypic antibodies can be produced

33. ANTIBODY DIVERSITY Presence of a large number of antibodies that bind to different antigens/pathogens T he total collection of antibodies with different specificities is called the antibody repertoire Genetic basis of antibody diversity S plit genes for immunoglobulins An individual has the capacity to produce an estimated 108 different antibody molecules Plasma cells produce antigen-specific antibody of IgM class Later gene arrangement occurs and different antibody class is produced

34. GENETIC REGULATION OF IMMUNOGLOBULINS Two regulator sequences in the Ig DNA. 1. Promoters: a group of short nucleotide sequences that initiate transcription 2. Enhancers: activate transcription from the promoter sequence Genetic Characteristics of Immunoglobulins Respond to an array of foreign antigens. Multiple genes code for single immunoglobulin heavy or light chain. Genes are reorganised during maturation of B-cell

35. MONOCLONAL ANTIBODIES Antibodies produced by a single clone and directed against a single antigenic determinant Kohler and Milstein developed hybridoma technique for monoclonal antibody production Chimeric antibody: Murine variable regions and human constant regions Fig. 14.5 Monoclonal antibody production by hybridoma technique

36. MONOCLONAL ANTIBODIES Monoclonal antibodies have important clinical uses: Therapeutic — for treatment of lymphomas, rheumatoid arthritis Diagnostic imaging — radiolabelled monoclonal antibodies to detect tumour antigens Diagnostic reagents — diagnosing infections (immunoassays), measuring blood levels of various drugs Research — in vitro production of immunoglobulins  

37. HUMANISED (CHIMERIC) ANTIBODIES Mouse monoclonals humanised by genetic manipulation C himeric antibodies with murine variable regions and human constant regions U sed therapeutically to treat patients with refractory multiple myeloma ANTIBODY ENGINEERING A ntibody libraries built by fusing genes coding for antibodies with bacteriophage genomes. D esired human antibody prepared by infecting bacteria with the appropriate bacteriophage.

38. C. COMPLEMENT

39. PROPERTIES OF COMPLEMENT Complement of one species can react with antibodies of other species Does not increase after immunisation Heat-labile; inactivated at 56 o C for 30 minutes Binds only to antigen–antibody complexes and not with free antigen or antibody Only IgM, IgG3, IgG1, and IgG2 bind

40. COMPONENTS OF COMPEMENT A t least 30 distinct serum proteins which includes 1. Properdin system 2. Control proteins Complement is a complex of nine different fractions, C1 to C9 E rythrocyte–antibody complex is called EA. When C components are attached to EA, the product is called EAC , followed by the components that have reacted

41. FUNCTIONS OF COMPLEMENT Cytolysis—lysis of cells, bacteria, and viruses Chemotaxis Opsonisation Triggers inflammatory reactions Specific cell functions Secretion of immunoregulatory molecules Immune clearance — removes immune complexes

42. COMPLEMENT ACTIVATION Fig. 14.6 Biological effects of complement activation

43. COMPLEMENT PATHWAYS T hree parallel but independent mechanisms or pathways which differ only in the initial steps Classical pathway Alternate pathway Lectin pathway

44. 1. CLASSICAL COMPLEMENT PATHWAY Activated by: (a) Antigen–antibody complex (b) Aggregated immunoglobulin, (c) DNA, (d) C-reactive protein, (e) Trypsin-like enzymes (f) Some retroviruses.

45. 1. CLASSICAL COMPLEMENT PATHWAY Fig. 14.7 Initiation and outcome of classical complement pathway

46. 2. ALTERNATIVE COMPLEMENT PATHWAY Antibody-independent A ctivators are : i ) Endotoxins and teichoic acid of gram-positiv e bacteria ii) IgA and D iii) Cobra venom factor and the nephritic factor iv) Zymosan v) Viruses & virus-infected cells vi) Parasites ( e.g., trypanosomes) Alternative pathway of complement

47. 3. LECTIN COMPLEMENT PATHWAY (MANNOSE BINDING LECTIN OR MB LECTIN [MBL] PATHWAY) Mannose-binding lectin (MBL) binds to mannose on pathogen Does not depend on antibody for activation Acts on organisms like certain Salmonella, Neisseria, and Candida which have mannose residues on the surface Three complement pathways converge at the membrane attack complex ( MAC)

48. CONVERGENCE OF THE THREE PATHWAYS Fig. 14.8 Convergence of the three complement pathways

49. COMPLEMENT PATHWAYS Table 14.6 Differences between the three complement pathways

50. REGULATION OF COMPLEMENT ACTIVATION

51. BIOLOGICAL EFFECTS OF COMPLEMENT Phagocytosis Inflammatory response Hypersensitivity reaction Autoimmune disease — complement is decreased Endotoxic shock — massive C3 fixation and platelet adherence Immune adherence — a defence against pathogens

52. Fig. 14.9 Outcomes of complement-mediated immunological reactions

53. Table 14.7 Microbial evasion of complement-mediated damage

54. Table 14.8 Sites of synthesis of C components

55. DISEASES ASSOCIATED WITH ABNORMAL/ DEFICIENT COMPLEMENT ACTIVITY Deficiencies of the Complement System

56. DISEASES ASSOCIATED WITH ABNORMAL/ DEFICIENT COMPLEMENT ACTIVITY Complement-associated d iseases Asthma Glomerulonephritis V arious forms of arthritis A utoimmune heart disease Lupus erythematosus Multiple sclerosis I nflammatory bowel disease

57. QUANTITATION OF COMPLEMENT Complement needs to be estimated in Acquired and inherited complement deficiencies Inherited and acquired C1 inhibitor deficiency Disorders with complement activation Estimation of individual complement components Immunoprecipitation assays Nephelometry and turbidimetry Detection of complement activation products — C3a Estimation of complement function — hemolytic assays ELISA — detects all three functional activities of complement