BIOl 460 1st Edition Lecture 3 Outline of Last Lecture I. Antigenicity Vs. Immunogenicity Outline of Current Lecture II.Immunology ReactionsIII. Motifs, epitopes, and tetramersIV. DNP.BSA conjugateCurrent Lecture Immunology Lecture 3- Characteristic of Ag- Ab interaction:BSA- 50 K DaAlbumin- 150 K Da 1 Ab- 2 Ag binding site.BSA immunized mouse - collect anti- serumAdd Ag ( BSA) in increasing amounts. Extensive Lattice formation - Immuno precipitate.A divalent (reactant) component or atleast a trivalent component forms larger compound with many branches.These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.Immuno precipitate formed with a small molecule called precipitate.Immuno precipitate formed with a cellular component called agglutinization (bacteria, RBC).Motifs- on surface for Ab to bind.Epitope- Part of molecule Ab to bind.- antigenic determinant Immunization of mouse -alpha (1) alpha (2) alpha (3) or polyclonal antiserum or polyclonal Ab- alpha(4) collection of antibodies. Anti-serum called alpha (5) antibody.Alpha (1) to alpha(5) differ in antigen recognition site.Every immunization is polyclonal in nature.Monoclonal Ab- all antibody molecules are identical (not through immunization but lab).Monovalent Ag- one epitope.Tetramers- 4 repeated epitopes.Epitopes- may be 1-4 structure of protein. Long stretch of hydrophilic AA on protein surface. Can be sequential. Can be conformational or non- sequential and destroyed by denaturation. Lipids or polysaccharides on surface of proteins.We can make artificial epitopes:Protein carrier + small molecule(hapten) ---- Conjugate (small molecule added to Surface of protein)Ab can recognize either epitope or hapten or conjugate.Favorite hapten- DNP- Dinitro-phenol.BSA- DNP conjugate is polyvalent for DNP.BSA- DNP conjugate immunized mouse ---- alpha 1 alpha 2 alpha 3 alpha 4 alpha 5alpha DNPProtein AProtein B immunized mouse --------- alpha A - antiserum alpha BAntiserum + protein A --- Protein A alpha A immuno precipitate Alpha B in supernatantDNP (monovalent) + alpha (DNP) --- No extensive lattice is formed.DNP.BSA (conjugate bivalent) + alpha (DNP) ---- precipitate formed.In a test tube containing alpha DNP add equal amounts of DNP.BSA conjugate. Then add increasing amount of free DNP.Result observed is decreasing conc. of precipitate formed because of competition due to free DNP.Competition: free DNP and DNP.BSA are competing for Ag binding site.DNP.BSA immunized mouse ---- alpha BSA and alpha DNP in test tube.Absorbant Precipitate SupernatantDNP.BSA alpha(DNP.BSA) DNP.BSA alpha(BSA).BSABSA alpha BSA.BSA alpha DNPDNP No precipitate free alpha BSA alpha DNP.DNPDNP.Ova alpha DNP(Ova.DNP) alpha BSA DNP.Ova is ovalbumin conjugate DNPAbsorption- purposeful addition of Ag to antiserum. Antiserum was absorbed on (name of