Basics of Immunology

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1 Basics of Immunology

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3 Basics of Immunology What is the immune system? Biological mechanism for identifying and destroying pathogens within a larger organism. Pathogens: agents that cause disease Bacteria, viruses, fungi, worms, etc. Types: Innate Immunity Adaptive Immunity 3

4 Immunity - Definition & Types Innate Immunity Adaptive Immunity Definition: Resistance to infectious diseases and is mediated by immune system Types: Innate or Non Specific Anatomic barriers: Skin, mucous membrane Physiologic barriers: Temperature, ph Phagocytic Barriers: Inflammatory barriers: redness, swelling, heat and pain Adaptive or Specific Antigen specificity Diversity Immunological Memory Self or non self recognition 4

5 Innate immunity

6 Adaptive Immunity - Types Naturally acquired Immunity Arises as a result of accidental interaction of a disease causing agent Artificially acquired Immunity Immunity due to deliberate interaction of a disease causing agent. e.g.., vaccination The colors of the receptors indicate specificity: each can bind to one specific antigen. Adaptive immunity can only attack targets that it has prepared for 6

7 Adaptive Immunity Adaptive Immunity Naturally acquired Artificially acquired Active Passive Active Passive Antigens enter the body naturally; body induces antibodies and specialized lymphocytes Antibodies pass from mother to fetus via placenta or to infant via the mother s milk Antigens are introduced by vaccination; body produces antibodies and specialized lymphocytes Preformed antibodies in immune serum are introduced by injection 7

8 Components of Immune System Cells Antigen presenting cells Polymorphic mono nuclear Leukocytes Cytotoxic leukocytes Natural Killer (NK) Cells Lymphokine activated Killers (LAK) Lymphocytes, B cells and T Cells Proteins Cytokines Antibodies Acute phase Reactants Antibodies Each B cell produces a specific type of antibody Antigen: Antibody Generator Any substance that can elicit an immune response when administered parenterally Reacts with the antibody thus produced 8

9 Structure of Antibody Composed of 2 heavy and 2 light chains encoded by different genes Single disulfide bond between heavy and light, two disulfide bonds between 2 heavy chains. Light chain is either ƙ (kappa) or λ (lambda); 98% of mouse antibodies are ƙ. Heavy chain constant region varies depending on isotype. 9

10 Epitope Paratope Interaction Variable and constant regions are encoded by different gene segments variable region binds antigen constant region binds receptors, bacterial products, complement, etc. 10

11 Types of Antibodies - IgG Induced by protein antigens Constitutes about 80% of antibody in serum Composed of two light chains (κ or λ) and one type of heavy chain (γ) Coats Ag for its speedy uptake Four isotypes IgG1, IgG2, IgG3, IgG4 Most of BioGenex Antibodies are IgG Type 11

12 Types of Antibodies - IgA Constitutes 13% of total antibodies Predominant class of Abin extravascular secretions Present in saliva, tears, nasal secretions, bronchial and digestive tract mucus and mammary gland secretions Exists either as monomer or dimer Dimer is formed by two IgA monomers, a J chain and secretory component Has two isotypesiga1 and IgA2 12

13 Types of Antibodies - IgM Largest antibody Constitutes upto8% of total Abs It is a pentamerformed by five monomers of IgMAbs joined by disulphidebridges First Abthat appears during immune response Activates complement system 13

14 Types of Antibodies - IgD Membrane bound Constitutes less than 1% of total Abs Membrane bound and one of the main receptors on mature B cells May play an important role in regulation of immune responses. 14

15 Types of Antibodies - IgE Constitutes less than 0.003% of total Abs Triggers allergies Binds to Mast cells and Basophils Subsequent exposure to the same antigen triggers allergic reaction Responsible for release of factors that attracts Eisonophils 15

16 Polyclonal vs. Monoclonal Antibodies Polyclonal Abs are produced from different clones of B cells for a given Ag Recognizes the different epitopes on an Ag Monoclonal Abare produced against a single epitope 16

17 Polyclonal vs. Monoclonal Antibodies Polyclonal Abs is produced from different clones of B Lymphocytes and hence different antibodies with distinct specificity for given antigen (i.esame antigen different epitopes).it s robust, exhibit cross reactivity Monoclonal Abs is produced from a single clone and directed against a single epitope. It show lot to lot consistency and is specific 17

18 Polyclonal vs. Monoclonal Antibodies Monoclonal Single plasma cell population / single epitope Higher Specificity Hybridoma technology Lot to lot stability Longer time to generate Multiple epitopes Higher Sensitivity Polyclonal Produced in lab animals like Rabbit, Goat, etc. Lot to lot variations Cross Reactivity 18

19 Applications of Antibodies Western blotting: Detection of protein of interest in a mixture Flow cytometry: Detection of proteins in live cells ELISA: Binding characteristics of proteins and quantitation IHC: localization of proteins of interest with in a cell In situ Hybridization: detect modification and abnormalities at genetic level Western IHC Direct Bind ELISA Denatyred/linear epitope IP/ChIP Sandwich IF/Flow Cytometry Functional Modulation folded/discontinuous epitope 19

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