Manual of basic techniques for a health laboratory - libdoc.who.int

11. Immunological and serological techniques 333IncubatethenwashFFFFFFAntigen(cell or tissue specimen)AntibodyFluorescence-labelledanti-immunoglobulinIncubatethen washFFFFFig. 11.6 Principle of indirect immunofluorescenceFluorescenceIncubatethenwashAntigen(insoluble)Fig. 11.7 Principle of agglutinationAntibody-containingserumPrecipitationAgglutinationAgglutination involves the reaction of an antibody with a particulate (insoluble)antigen leading to visible clumping of these particles (Fig. 11.7). The interaction ofsurface antigens and antibodies directed against them leads to cross-linking ofadjacent particles, e.g. bacteria, to form a lattice of agglutinated cells.Active agglutination (direct)Active agglutination involves antigenic determinants that are an intrinsic constituentof the particle, e.g. haemagglutination reactions used for blood grouping.Passive agglutination (indirect)Passive agglutination involves antigenic determinants that are not an intrinsic constituentof the particle. A soluble antigen is combined with insoluble particles suchas erythrocytes. The erythrocytes are usually treated with tannic acid, which alterstheir surface properties so that the antigen can bind firmly. Other insoluble particlesinclude bacteria, charcoal, bentonite (clay) and polyvinyl latex where the antigenis simply adsorbed.Semi-quantitative titrations can be carried out to determine the amount of antibodypresent in a sample. A constant volume of suspended particles (antigen) isadded to a constant volume of serially diluted antiserum. The presence of antibodyin the serum causes agglutination of the particles (Fig. 11.8) and the reaction isusually scored on a scale of 0 to 4 +. The antibody content is expressed as a titre —