Antibodies and antigens PowerPoint Presentation

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Antibodies and antigens Antibodies = immunoglobulins Antibodies bind antigens.

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3 different kinds of antigen-binding molecules; Ig, TCR and MHC.

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Antibodies On the surface of B cells: Antigen receptors for activation of B cells. Secreted: Effector phase of humoral immunity. Cooperation with complement, phagocytes, mast cells and eosinophils.

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Distribution of secreted Ab Plasma Mucosal secretions Interstitial fluid Surface of immune effector cells

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Antiserum: Serum with specific ab. Titer: Reciproke of end point dilution. End point dilution: Highest dilution of serum that gives a positive reaction. Dilution: 1/100 1/200 1/400 1/800 Reaction: + + + − End point dilution: 1/400 Titer: 400

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Effects of antibodies: Neutralization of microbes or toxins Complement activation Opsonization Antibody dependent cytotoxicity Immediate hypersensitivity

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Antibodies are clonally distributed: Clone: Population of identical cells, derived from a single cell by cell division. Clonal distribution of abs: One clone makes only one ab, other clones make different abs. Allelic exclusion.

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Serum: Polyclonal Ab Myeloma tumors: Monoclonal Ab Hybridomas

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Detection of monoclonal bands by serum electrophoresis in agarose gel. + Application - Albumin γ globulin

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Hybridoma monoclonal antibodies

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Some advantages of monoclonal abs: Increased specificity Unlimited supply Standardized reagents; can compare results from different labs. Possible to immunize with impure antigens, e.g. whole cells.

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Structure of antibody molecules: H2L2 (1, 2 or 5; monomer, dimer or pentamer) (H: Heavy chain) (L: Light chain)

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Downloaded from: StudentConsult (on 4 January 2008 01:16 PM) © 2005 Elsevier

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Downloaded from: StudentConsult (on 4 January 2008 01:16 PM) © 2005 Elsevier Structure of an Ab light chain

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The immunoglobulin superfamily

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Proteolytic fragments of an IgG molecule

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Downloaded from: StudentConsult (on 4 January 2008 01:16 PM) © 2005 Elsevier Hypervariable regions in Ab molecules Variability: No. different aa/Frequency of most common aa.

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Downloaded from: StudentConsult (on 4 January 2008 01:16 PM) © 2005 Elsevier Binding of antigen by an antibody’s hypervariable regions of VH and VL.

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Downloaded from: StudentConsult (on 4 January 2008 01:16 PM) © 2005 Elsevier Flexibility of antibody molecules

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Antibodies are divided in classes (isotypes) based on differences in heavy chains.

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Human Ig classes

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Downloaded from: StudentConsult (on 4 January 2008 01:16 PM) © 2005 Elsevier Membrane and secreted forms of Ig heavy chains.

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Two types of light chains: κ (kappa) λ (lambda)

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Synthesis of Ig: Rough endoplasmic reticulum Chaperones (calnexin, BiP) ensure proper folding Golgi complex

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Downloaded from: StudentConsult (on 4 January 2008 01:16 PM) © 2005 Elsevier Ig expression during B lymphocyte maturation.

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Antigens bound by Abs: Proteins Carbohydrates Nucleic acids Phospholipids Small chemicals

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Haptens: Small molecules that must be conjugated to carrier molecules (proteins) to initiate an immune response. They are antigenic (bind Ab) but not immunogenic (cannot by themselves induce an Ab response). Immunogens are macromolecules, and (of course) antigenic.

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Haptens and carriers (Bogen and Munthe) (Carrier molecule) (Hapten-carrier complex) (not) (to)

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Determinants (epitopes) Poly (multi) valent antigens Polyvalent antigens can be immunogenic for B cells without the help of T cells

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Nonoverlapping determinants Overlapping determinants Allosteric effects Neoantigenic determinants (result of postsynthetic modifications, such as proteolysis or phosphorylation)

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Downloaded from: StudentConsult (on 4 January 2008 01:16 PM) © 2005 Elsevier The nature of antigenic determinants

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Antibody-antigen binding: Noncovalent forces Electrostatic forces Hydrogen bonds van der Waals forces Hydrophobic interactions

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Avidity and valency of antibody-antigen interactions

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Bogen and Munthe Avidity Affinity

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Equilibrium dialysis

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Downloaded from: StudentConsult (on 4 January 2008 01:26 PM) © 2005 Elsevier Antigen-antibody complexes

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Three important characteristics of humoral immunity: Specificity Diversity Affinity maturation

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Specificity (distinction of similar antigens) Karl Landsteiner Meta-azoben-zene sulfonate Cross-reactions

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Diversity (specific binding of a large number of antigens). Affinity maturation (late abs and abs in secondary reactions bind better).

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Changes in antibody structure during humoral immune responses

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Isotypes Allotypes Idiotypes

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Binding of two or more adjacent Fc portions is needed to trigger effector functions such as complement activation and phagocytosis.

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Effector functions often mediated by Fc portion. Effector functions are initiated by ab that has bound ag. Isotypes influence how microbes are attacked. Isotypes determine tissue distribution.