IMMUNOASSAY
    • An immunoassay is a test that uses antibody               
             and antigen complexes as a mean of generating 
     a measurable result
       • Immuno refers to an immune response that 
         causes the body to generate antibodies, 
       • And assay refers to a test 
    • An antibody-antigen complex is also known as an 
     immuno-complex
    • The assay takes advantage of the specific binding of 
     an antibody to its antigen
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     The antibodies used must have a high affinity for the 
     antigen
                                Dr. M. Alzaharna 2015
                      IMMUNOASSAYS
       • Immunoassays derive their unique specificity, 
        sensitivity, and flexibility from three important 
        properties of antibodies:
          • Their ability to bind to an extremely wide range 
            of natural and man-made chemicals, 
            biomolecules, cells, and viruses
          • Exceptional specificity for the substance to 
            which each antibody binds.
          • The strength of the binding between an antibody 
            and its target
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                                Dr. M. Alzaharna 2015
                 Immunoassay
     • Both the presence of antigen or antibody can be 
       measured
        • For measuring hormones such as insulin, the insulin 
          acts as the antigen
        • When detecting infection the presence of antibody 
          against the pathogen is measured
     • For numerical results the response of the object being 
       measured must be compared to standards of a known 
       concentration
     • This is usually done through the plotting of a standard 
       curve on a graph paper, and then the quantity of the 
       unknown is found from the curve
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                                Dr. M. Alzaharna 2015
                      TYPES OF TESTS
     • Many methods of varying sophistication are used 
       for immunodiagnostic studies 
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                                Dr. M. Alzaharna 2015
                                      TYPES OF TESTS
   Visible Change                                Observable Reaction                        Name of Test
   Clumping                        Particulate antigen reacts with corresponding 
                                       antibody; antigen may be in form of RBCs            Agglutination
                                       (hemagglutination, latex, or charcoal coated 
                                       with antigen).
   Precipitates                    Soluble antigen reacts with corresponding                Precipitation
                                       antibody
   Complement activation,          Competition between two antigen-antibody             Complement fixation 
        hemolysis                      systems (test and indicator systems)                       (CF)
   Visible microscopic             Fluorescent-tagged antibody reacts with 
        fluorescence                   antigen-antibody complex in the presence of      Immunofluorescence
                                       ultraviolet light.
   Color change indicates          Indirect EIA for quantification of an antigen or        Enzyme-linked 
        enzyme substrate               antibody enzyme and substrate                    immunosorbent assay 
        reaction.                                                                             (ELISA)
   Detection of antibodies of      Electrophoresis separation of antigen subspecies       Immunoblot (eg, 
        specific mobility                                                               Western blot [WB])
   Light scatter                   Measures either antigen or antibody in solution 
        proportionately                through the scattering of a light beam; 
        increases as numbered          antibody reagent used to detect antigen IgA,    Rate nephelometry
        size of immune                 IgG, IgM; concurrent controls are run to 
        complexes increases.           establish amount of background scatter in 
                                       reagents and test samples.
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                                                   Dr. M. Alzaharna 2015