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Published on : Mar 14, 2014
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Slide 1 - Antibodies: Structure And Function
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Slide 3 - Antibody Structure Antibodies Are Made Up Of: 2 Light Chains (identical) ~25 KDa 2 Heavy Chains (identical) ~50 KDa Each Light Chain Bound To Heavy Chain By Disulfide (H-L) Heavy Chain Bound to Heavy Chain (H-H) First 100 a/a Of Amino Terminal Vary of Both H and L Chain Are Variable Referred To As VL , VH, CH And CL CDR (Complementarity Determining Regions) Are What Bind Ag Remaining Regions Are Very Similar Within Same Class
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Slide 5 - Repeating Domains of ~110 a/a Intrachain disulfide bonds within each domain Heavy chains 1 VH and either 3 or 4 CH (CH1, CH2, CH3, CH4) Light chains 1 VL and 1 CL Hinge Region Rich in proline residues (flexible) Hinge found in IgG, IgA and IgD Proline residues are target for proteolytic digestion (papain and pepsin) Rich in cysteine residues (disulfide bonds) IgM and IgE lack hinge region They instead have extra CH4 Domain Antibody Structure
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Slide 7 - Digestion With Papain Yields 3 Fragments 2 identical Fab and 1 Fc Fab Because Fragment That is Antigen Binding Fc Because Found To Crystallize In Cold Storage Pepsin Digestion F(ab`)2 No Fc Recovery, Digested Entirely Mercaptoethanol Reduction (Eliminates Disulfide Bonds) And Alkylation Showed Enzymatic Digestion Of Antibodies
Slide 8 - Sequencing Of Several Immunoglobulins Revealed 100-110 Amino Terminus, Highly Variable (V) Five Basic Sequence Patterns ,, , ,  IgA, IgG, IgD, IgE and IgM The Above Classes Are Called Isotype Each class can have either  or  light chains Minor Differences Led To Sub-classes For IgA and IgG IgA1, IgGA2 and IgG1, IgG2, IgG3, IgG4 Sequencing Of Heavy Chains
Slide 9 - CDR Are Hypevariable
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Slide 11 - Immunize Animal With Antigen Multiple Clones Are Generated, Good For In Vivo For Clinical Diagnosis, Research, One Clone That Reacts To Single Epitope Is Preferred Solution By Kohler and Milstein Fuse A Myeloma Cell (Cancerous) With A Normal Plasma Cells Resulting Clones Can Be Cultured Indefinitely Produces An Antibody Recognizing One Epitope Monoclonal Antibodies
Slide 12 - BCR Is An Antibody On Surface Of Cell mIg Very Short Cytoplasmic Tail, Cannot Transduce Signal Heterodimeric Molecule Ig-/Ig- Transduces (long cytoplasmic tail) B-Cell Receptor
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Slide 14 - Fc Receptors (FcR)
Slide 15 - To Transport Abs Across Membranes Secretion of IgA Across Epithelium into lumen Transport of maternal Abs Across Placenta (IgG) Many Cell Types Use FcR Ex. Mast Cells, Macrophages, Neutrophils, B, T, NK Opsonization, ADCC Poly IgR Transport of IgA across epithelium FcRN Transport of maternal IgG to fetus Fc Receptors (FcR) Functions
Slide 16 - IgG Most abundant immunoglobin 80% of serum Ig ~10mg/mL IgG1,2,3,4 (decreasing serum concentration) IgG1, IgG3 and IgG4 cross placenta IgG3 Most effective complement activator IgG1 and IgG3 High affinity for FcR on phagocytic cells, good for opsonization Antibody Classes And Biological Activities
Slide 17 - IgM 5-10% of serum immunoglobulin 1.5mg/mL mIgM (also IgD) expressed on B-cells as BCR Pentameric version is secreted First Ig of primary immune response High valence Ig (10 theoretical), 5 empirical More efficient than IgG in complement activation Antibody Classes And Biological Activities
Slide 18 - IgA 10-15% of serum IgG Predominant Ig in secretions Milk, saliva, tears, mucus 5-15 g of IgA released in secretions!!!! Serum mainly monomeric, polymers possible not common though Secretions, as dimer or tetramer+J-chain polyptetide+secretory component (Poly IgR) Antibody Classes And Biological Activities
Slide 19 - IgA Antibody Transport Across Cell (Transcytosis)
Slide 20 - IgE Very low serum concentration, 0.3g/mL Participate in immediate hypersensitivities reations. Ex. Asthma, anaphylaxis, hives Binds Mast Cells and Blood Basophils thru FcR Binding causes degranulation (Histamine Release) Antibody Classes And Biological Activities
Slide 21 - IgD Expressed on B-cell Surface IgM and IgD, Expressed on B-cell Surface We Do Not Know Any Other Biological Effector Activity Low serum concentrations, ~30g/mL Antibody Classes And Biological Activities
Slide 22 - Cross-Linkage of Bound IgE Antibody With Allergen Causes
Slide 23 - Antibodies Act As Immunogens Antigenic Determinants on Abs Fall in 3 Categories Isotypic Allotypic Idiotypic Isotypic Constant Region Of Ab If you inject Ab in a different species Anti-Isotype is generated If within same species, No Anti-isotype
Slide 24 - Allotype Even though same isotypes within one species small differences (1-4 a/a) arise in different individuals (form of polymorphism) If injected with such Ab you generate anti-allotype Ab Ex. During pregnancy Blood transfusion Antibodies Act As Immunogens
Slide 25 - Idiotype Unique VH AND VL binds antigen but can also behave as antigenic determinant If you inject a monoclonal antibody into a genetically identical recipient then anti-idiotypic antibodies are generated No anti-isotypic and no anti-allotypic Abs will be generated Antibodies Act As Immunogens
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Slide 27 - Monoclonal Antibody Applications Diagnostic Tests Abs are capable to detect tiny amouns (pg/mL) of molecules Ex. Pregnancy hormones Diagnostic Imaging mAbs that recognize tumor antigens are radiolabeled with iodine I-131 Immunotoxins mAbs conjugated with toxins mAbs To Clear Pathogens www.elusys.com