Agglutination Reaction

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Agglutination Reaction is the immunological reaction where the specific antigen and antibody reaction takes place in vitro.  Agglutination can define as the clumping reaction where clumps or aggregates are formed.  As a result of agglutination, a cross-linked structure forms which refer as “Lattice” that appears in the form of visible aggregates.

Agglutination reaction is a sensitive, easy, cost-effective method to carry out and does not require technical skills to operate sophisticated equipment. The lattice or the network of antigen and antibody can be seen by macroscopically and microscopically depending upon the type of agglutination reaction.

Content: Agglutination Reaction

  1. Definition of Agglutination Reaction
  2. History
  3. Principle of Agglutination Reaction
  4. Steps of Agglutination
  5. Types of Agglutination Reaction
  6. Applications
  7. Conclusion

Definition of Agglutination Reaction

Agglutination reaction can define as the serological reaction where the large or particulate antigen is mixed with the antiserum containing antibodies in solid support like glass side, microtitre plate or test tubes. The agglutination between antigen and antibody occurs when the conditions are optimal like temperature, pH and ionic strength of the solution.

The binding or interaction of both the reactants, i.e. antigen and antibody will result in the formation of antigen-antibody network or lattice which appears as “visible clumps” to the naked eye. The most common example of the agglutination reaction is “Blood typing”.

History

YearDiscoverer Discovery
1896Herbert Edward Durham and Max Von GruberIntroduced the theory of “Specific- agglutination”
1862-1926Fernand WidalIntroduced Widal agglutination for the diagnosis of typhoid fever
1900Karl LandsteinerIntroduced blood typing based on the principle of “Agglutination”

Principle of Agglutination Reaction

The principle of agglutination reaction is based on the “Clumping of antigen and antibody”. Like precipitation reaction, it also involves the binding of antigen and antibody at the zone of equivalence, where both are present in equal proportion.

As the antibodies are y-shaped structure and there are two Fab sites made of “Hypervariable region” will target the specific antigenic determinants or epitopes of an antigen. The binding of antigen and antibody is similar to the “Lock-key model”. Therefore, the epitopes of an antigen are the key which fits into the cleft of the Fab sites of the antibody that works as a lock.

Steps of Agglutination

Agglutination reaction involves two steps:

Sensitization: It can define as the primary stage where binding of antigen and antibody takes place. Temperature, pH, ionic strength and incubation period influence the efficiency of sensitization or binding. Sensitization is not a visible reaction.

Lattice formation: It can define as the secondary stage where the antibody and multivalent antigen forms a Stable network refers as “Lattice”. A lattice is a net-like configuration which consists of a network between sensitized antigen and antibody. It takes much time to occur than that of sensitization. Lattice is a visible reaction.

Types of Agglutination Reaction

Based on the nature of the antigen, agglutination reactions are of two types namely active and passive agglutination.

Active agglutination

It involves direct interaction of antibodies present in the serum with the particulate antigens which carry epitopes of interest. Active agglutination is used to quantify antibodies against the particulate antigens like RBCs, Pathogenic microorganisms like bacteria, fungi etc.

active agglutination

In this method first, add the particulate antigens into the wells of a microtitre plate. Then serially dilute the antibodies and add to the wells coated with an antigen. By increasing the concentration of antibody, the rate of agglutination will also increase.

Positive result: Antibodies present in the serum will bind with the particulate antigen as a result of which antigen-antibody mat forms that settle at the bottom of the well.

Negative result: It occurs due to the presence of an insufficient amount of antibodies to agglutinate the antigens. Therefore, the agglutination reaction will not occur. In this case, instead of clumping the antigens appears as “Pellets” at the bottom of the well.

Example: Slide agglutination test(Blood-grouping), Widal test for the diagnosis of typhoid fever, Coomb’s test for the detection of anti-Rh antibodies etc.

Passive agglutination

It involves indirect interaction of antibody with the soluble antigen by the means of carrier particle.

passive agglutination

Passive agglutination first, consists of the binding of soluble antigen with the carrier matrix like latex bead, polystyrene etc. In this type, the antigens do not carry epitopes of interest that have to be agglutinated.

There is another type, where in place of antigen, known antibodies attach with the carrier matrix refers as Reverse passive agglutination. And after the attachment of antibody with the carrier molecule, the antigens attach to the Fab sites of the antibody to agglutinate it.

reverse passive agglutination

Example: Latex bead agglutination for both antigen and antibody.

Applications

Agglutination test is widely applicable in the field of Clinical Microbiology, where some of the applications are given below:

  1. Blood typing of recipient and donor at the time of blood transfusion.
  2. Helps in the detection of antibody presence and to quantify the amount of antibody present in the patient’s blood.
  3. Active agglutination helps in serodiagnosis of Toxoplasmosis, brucellosis etc.
  4. Passive agglutination helps in the determination of the Rh-factor
  5. Widely used in techniques like latex and haemagglutination.
  6. Helps to detect the type of antigen and to quantify the amount of antigen present in the patient’s blood.

Conclusion

Therefore, active and passive are the two main types of agglutination based on the nature of an antigen. Other than active, passive and reverse passive there is another type refers as “Miscellaneous agglutination”. When clumping occurs between the antibodies against a particulate antigen of different but the related organism will refer as “Cross agglutination”.

Agglutination which occurs between the antigens of the biologically related organism with the specific antibody related to that particular group will refer as “Group agglutination”. Clumping of particulate antigenic elements with the blood cells like RBCs within the blood vessels will refer as “Intravascular agglutination”.

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