ag ab reactions.pdf
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Antigen- Antibody reactions Antigen: a molecule (or parts of one) that causes antibody generation (Immunoglobulin) The specific region on an antigen recognized by an antibody Affinity Combined strength...
Antigen- Antibody reactions Antigen: a molecule (or parts of one) that causes antibody generation (Immunoglobulin) The specific region on an antigen recognized by an antibody Affinity Combined strength of total non-covalent interactions between single Ag- binding site of Ab and single epitope is affinity of Ab for that epitope. Low affinity Ab: Bind Ag weakly and dissociates readily. High affinity Ab: Bind Ag tightly and remain bound longer. Avidity Strength of multiple interactions between multivalent Ab and Ag is avidity. Avidity is better measure of binding capacity of antibody than affinity. High avidity can compensate low affinity. Cross reactivity Antibody elicited by one Ag can cross react with unrelated Ag if they share identical epitopeor have similar chemical properties. Immune complex formation: Ab have two Ag binding sites and most Ag have at least two epitopes, cross linking can occur producing large aggregates called immune complex. If the Ag s are soluble molecules and the compex becomes large enough to settle out of soution a precipitation occurs When the immune complex involves the crosslinking of cells or particles an agglutination reaction occurs Multivalent antigens combine with bivalent antibodies in varying proportions, depending on the antigen-antibody ratio in the reacting mixture. Precipitation results with the formation of large lattice consisting of alternating antigens and antibody molecules (Mancini) This is a standard method for quantitative estimation of Ab. Slide test WIDAL test Tube agglutination test to detect bacterial infection Applications of Agglutination Reactions Cross-matching and grouping of blood. Identification of Bacteria. E.g. Serotyping of Vibrio cholera, Serotyping of Salmonella typhi and paratyphi. Serological diagnosis of various diseases. E.g Rapid plasma regains (RPR) test for Syphilis, Antistreptolysin O (ASO) test for rheumatic fever. Detection of unknown antigen in various clinical specimens. E.g. detection of Vi antigen of Salmonella typhi in the urine. Complement fixation A group of serum proteins collectively called complement. During most antigen- antibody reactions, complement binds to the antigen-antibody complex and is used up, or fixed. This process of complement fixation can be used to detect very small amounts of antibody. Antibodies that do not produce a visible reaction, such as precipitation or agglutination, can be demonstrated by the fixing of complement during the antigen- antibody reaction. Complement fixation Complement fixation was once used in the diagnosis of syphilis (Wassermann test) and is still used to diagnose certain viral, fungal, and rickettsial diseases. The complement-fixation test requires great care and good controls, one reason the trend is to replace it with newer, simpler tests. The test is performed in two stages: complement fixation and indicator ` http://highered.mheducation.com/sites/0072556781/student_view0/chapter31/animation _quiz_4.html ELISA TYPES OF ELISA 1 Non competitivebinding assay A. Sandwich or Direct ELISA: Antigen measuring system [Titrewellscoated with antibodies ; Enzyme labelled antibodies] B. Indirect ELISA: Antibody measuring system [Titrewellscoated with antigens ; Enzyme labelled antiantibodies] 2. Competitive binding assay [Titrewellscoated with antibodies ; Enzyme labelled antigens] IMMUNOFLUORESCENCE: Fluorescent dyes (fluoresceinor rhodamine) attached to known specific Abs, used to detect presence of Abs in serum or Ag (microorganisms) in a sample Direct fluorescent Abtest: used to detect Ag or microorganism Indirect fluorescent Abtest: used to find a specific Abin the serum Quantitative Immunofluorescence-flow cytometry
