Antibodies

Introduction Definition: Immunoglobulins represent animal proteins, which possess antibody activity and other proteins with related chemical structure and antigenic specificity. (according WHO)

Antibodies Structure

Immunoglobulin structure Heavy chain IgM, IgG1-4, IgA1-2, IgD, IgE Light chain Domain Ag binding site

Immunoglobulin structure Immunoglobulin domains Variable: VH, VL Constant:CH, CL

Complementarity-determining regions [CDR] hypervariable loops

Antibodies Antigen binding site domains, complementarity determining regions (CDR)- hypervariable regions/loops VL : CDR1, CDR2, CDR3 VH : CDR1, CDR2, CDR3

Class and subclass (isotypes) IgM IgG IgA IgD IgE Antibodies Class and subclass (isotypes) Ig Class Ig subclass IgM no IgG IgG1, IgG2, IgG3, IgG4, IgA IgA1, IgA2 IgD IgE

Immunoglobulin structure Fragments

Immunoglobulin structure Immunoglobulin domain

Immunoglobulin superfamily Immunoglobulin domain

Immunoglobulin structure Antigen binding site

Immunoglobulin domains Antibodies Immunoglobulin domains Light chain Heavy chain domains VL CL VH CH IgM 1 4 IgG 3 IgA IgD IgE

Antigenic Determinants on Immunoglobulins Name Localization of Ag determinants Distribution on cells / molecules Idiotype VH and VL (in and out CDR) several B cell clones >> public i. individual B cell clone >> private i. Isotype -class, subclass CH all members of a species -type and subtype CL Allotype Gm, Am Km [ in animals also VH,L] some, but not all members of a species

Immunoglobulin Isotypes IgM (kappa) IgG1 (kappa) Importance Ig levels B cell tumors Immunodeficiencies

Immunoglobulin Allotypes IgG1 (kappa) Person 2 Person 1 Definition - Antigenic determinants specified by allelic forms of the Ig genes Importance -Monitoring bone marrow grafts -Forensic medicine -Paternity testing

Immunoglobulin Idiotypes Importance V-region marker Regulation of immune responses IgG1 (kappa) Person 1 anti-A IgG1 (kappa) Person 1 anti-B

Antibodies Function

Biological activities of immunoglobulins Property / activity IgG1 IgG2 IgG3 IgG4 IgA1 IgA2 IgM IgE IgD Serum level [mg/ml] 9 3 1 0.5 1.5 0.0003 0.03 Half life [days] 23 8 6 5 2.5 1-st in primary response + Activates classical complement way ++ CH2 +/- - CH4 Crosses placenta Present on mature B cells Binds to mph Fc receptors Present in secretions Induces mast-cell degranulation

Antibodies Ig genes

                                                      Susumu Tonegawa The Nobel Prize in Physiology or Medicine 1987 „for his discovery of the genetic principle for generation of antibody diversity“

Antibody response Cellular lever Antibodies Antibody response Cellular lever

B cell activation Two signals are required for activation of B cell. The first signal: binding of Ag to BCR (surface Ig) The second signal: binding of Th cell to B cell. After activation, B cell differentiate to - plasma cell >> secretion of Ab - memory B cells >> secondary response

B cell activation The second signal represents contact between CD40 receptor on B cell and CD40L ligand expressed on Th cells

B cell activation Only activated Th cell is able to provide the second signal for B cells. Th cells are activated by antigen presenting cell (APC). Activation of Th requires also two signals (steps): 1st> APC engulf Ag, process Ag and express Ag fragments in the frame of MHCII on the cell surface for TCR (T cell receptor) on T cells. 2nd > APC produce IL-1 which binds to IL-1 receptor on Th cell.

Primary and Secondary Responce Antibodies Antibody production Primary and Secondary Responce

Primary and secondary response Primary response - 1st contact with Ag IgM Secondary response - 2nd contact with Ag - based on memory B cells - CLASS SWITCHING Under the influence of cytokines B cells switch from IgM production to IgG and other classes of Ab. afinity maturation: Ab have high afinity to Ag compared with the afinity of Ab produced in primary response faster onset of Ab production higher levels (titers) of Ab longer persistence of Ab (slower drop of Ab titers).

Monoclonal antibodies Principle: 1) “Immortalization” of Ag specific B cell clone by fusion with myeloma cell. 2) Selection of this clone in HAT medium. 3) Propagation of Ag specific clone in vitro or in mice. ____________________________ (Myeloma is a cancer cell “without limitation” of proliferation)