高美华《医学免疫学》precipitation.mbb.ppt

Experiment 2,Precipitation,Definition:Soluble Ag+Ab Ag-Ab complexClassification:Precipitation reaction in solution Precipitation reaction in gels Immunoelectrophoresis Technique Immunoturbidimetry,Precipitation Reactions In Gels,Agar、AgarosePrecipitation reactions between soluble antigens and antibodies can take place in semisolid media such as agar gels.When soluble antigen and antibodies are placed in wells cut in the gel,the reactants diffuse in the gel and form gradients of concentration,with the highest concentrations closest to the wells.Some where between the two wells,the reacting antigen and antibodies will be present at proportions that are optimal for formation of a precipitate.,Double Agar Diffusion Test,analysis of the properties of AgPatterns of IdentityPatterns of NonidentityPatterns of Partial Identity:spur,single agar diffusion test,The wells contain antigen at different concentrations,while the antibodies are distributed uniformly in the agar gel.Thus,the precipitin line is replaced by a precipitin ring around the well.The distance the precipitin ring migrates from the center of the antigen well is directly proportional to the concentration of antigen in the well.,Method,Result,Counter Immunoelectrophoresis,pH88.6 buffer,Ag：PI=5 MW：small Electric field force:cathodeanode，strongElectroosmosis:anodecathodeElectric field force ElectroosmosisDirection of phoresis:cathodeanode,Ab：PI=7 MW：largeElectric field force:cathodeanode，weakElectroosmosis:anodecathodeElectric field force Electroosmosisanodecathode,Cathode,+Anode,Ag,Ab,Result,Rocket Immunoelectrophoresis,Immunoelectrophoresis,Immunoelectrophoresis involves separating a mixture of proteins in an electrical field(electrophoresis)followed by their detection with antibodies diffusing into the gel.,a small drop of human serum is placed in a well cut in the center of a slide that is coated with agar gel.The serum is then subjected to electrophoresis,which separates the various components according to their mobilities in the electrical field.After electrophoresis,a trough is cut along the side of the slides,and antibodies to human serum proteins are placed in the trough.,The antibodies diffuse in the agar,as do the separated serum proteins.At an optimal antigen:antibody ratio for each antigen and its corresponding antibodies,they form precipitin lines.Comparison of the pattern and intensity of lines of normal human serum with the patterns and intensity of lines obtained with sera of patients may reveal an absence,overabundance,or other abnormality of one or more serum proteins.,