Passive Agglutination Reaction

Agglutination is an antigen–antibody reaction in which a particulate antigen combines with its antibody in presence of electrolytes at a specified temperature and pH resulting in formation of visible clumps. Depending on the nature of antigens, agglutination reaction can be active or passive.


Passive agglutination employs carrier particles that are coated with soluble antigens. In this either antibody or antigen is attached to certain inert carrier thereby, particles or cells gets agglutinated when corresponding antigen or antibody reacts. Latex particles, Carbon particles, Bantonite etc. are used as inert carriers. E.g. Antigens coated in latex particles used in ASO test.


Haemagglutination Test types


When antibody instead of antigens is adsorbed on carrier particle for detection of antigens, it is called reverse passive agglutination. Instead of antigen, specific antibody can be coated on carrier particles along their Fc end. The Fab portion remains free to bind with antigen resulting in agglutination. Such a method is called reverse passive agglutination. This is useful in detection of HBsAg in serum, pneumococcal antigens in urine, or Hemophilus, Neisseria, pneumococcus or Cryptococcus antigens in CSF.


Latex Agglutination: It employs latex particles as carrier of antigen or antibodies. In latex agglutination, many antibody or antigen molecules are bound to latex beads (particles), which increases the number of antigen-binding sites. If corresponding antigen or antibody is present in a test specimen, antigen antibody bind and form visible, cross-linked aggregates. Latex agglutination can also be performed with antigen conjugated to the beads for testing presence of antibodies in a serum specimen.


Latex carrier agglutination


Hemagglutination: RBCs are used as carrier particles in hemagglutination tests. RBCs of sheep, human, chick, etc. are commonly used in this test. When RBCs are coated with antigen to detect antibodies in serum, the test is called indirect hemagglutination (IHA) test. Hemagglutination uses erythrocytes as biological carriers of bacterial antigens, and purified polysaccharides or proteins for determining presence of corresponding antibodies in a specimen. When antibodies are attached to RBCs to detect microbial antigen, it is known as reverse passive hemagglutination (RPHA). They are used to detect the antigens in the test samples, such as HBsAg in the serum.


Rose-Waaller Test: This is a passive hemagglutination test to detect rheumatoid factor. A feature of human rheumatoid factor is it can bind with human IgG as well as rabbit IgG. Sheep erythrocytes are coated with sub-agglutinating amounts of anti-sheep IgG antibodies. The rheumatoid factors agglutinate sheep erythrocytes coated with rabbit anti-sheep IgG.


 Haemagglutination and Haemagglutination Inhibition


Viral hemagglutination: Many viruses including influenza, mumps, and measles have an ability to agglutinate RBCs without antigen–antibody reactions. This process is called viral hemagglutination. This hemagglutination can be inhibited by antibody specifically directed against virus, and this phenomenon is called hemagglutination inhibition and is used to quantitate viral antibodies in serum of patients.


Coagglutination test: Coagglutination is a type of agglutination reaction in which Cowan I strain of S. aureus is used as carrier particle to coat antibodies. Cowan I strain of S. aureus contains protein A, an anti-antibody, that combines with Fc portion of immunoglobulin, IgG, leaving Fab region free to react with antigen present in the specimens.


In a positive test, protein A bearing S. aureus coated with antibodies will be agglutinated if mixed with specific antigen. The advantage of this test is that these particles show greater stability than latex particles and are more refractory to changes in ionic strength.

 Coagglutination test


It is not sensitive for detecting small quantities of antigen. Thus, it is not usually used for direct antigen detection.

 Uses of Coagglutination test:

  1. Grouping of streptococci and mycobacteria and for typing of Neisseria gonorrhoeae.
  2. Detection of amoebic and hydatid antigens in serum for diagnosis of amoebiasis and cystic echinococcosis.
  3. Detection of cryptococcal antigen in CSF for diagnosis of cryptococcal meningitis.


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