Reaction mechanism
The essence of the reaction lies in the formation of a unified network (lattice) of antigens and antibodies. Antibodies act as specific "bridges" connecting particulate antigens together.
The scale of clumping depends on the class of participating immunoglobulins:
- IgM have a pentameric structure, allowing them to capture multiple bacteria or cells at once, forming large clusters.
- IgG are represented by monomers, so they only bind adjacent cells, also building a network.
Types of diagnostic agents
Depending on the antigen to be detected, different types of diagnostic agents (microbial suspensions) are used in the reactions:
- O-diagnostic agent: prepared from bacteria killed by heating. This preserves the thermostable somatic O-antigen. The reaction with such a diagnostic agent produces fine-granular agglutination.
- H-diagnostic agent: created from bacteria killed with formalin. In this case, the thermolabile flagellar H-antigen is preserved. Agglutination is large-floccular and occurs faster than O-agglutination.
Clinical significance
The agglutination reaction is widely used in medical practice:
- Serodiagnosis. Allows for the quantitative determination of the titer of specific antimicrobial antibodies in the patient's blood serum.
- Pathogen identification (serotyping). Using known diagnostic sera, the species of bacteria isolated from the patient can be precisely established.
- Immunohematology. Used for blood typing using monoclonal antibodies against erythrocyte alloantigens.
Expanded agglutination reaction
This method of setting up the reaction in test tubes is used for the precise quantification of antibodies.
Setup stages:
- A series of consecutive dilutions of blood serum is prepared.
- An equal amount of diagnostic agent (suspension of killed microbes) is added to all test tubes.
- The mixture is incubated for several hours at 37 °C.
As a result, the serum titer is determined—the maximum dilution at which the formation of a precipitate (clumping) is still visually noticeable.
Cross-reactions
A problem may arise during serotyping: closely related microorganisms often share common antigens. This leads to false-positive results—the same serum clumps different bacteria.
To solve this problem, Castellani's method (1902) proposes using absorbed agglutinating sera. All cross-reacting antibodies are removed from standard serum by precipitating them with related bacteria. Afterward, only specific antibodies reacting strictly to the specific pathogen remain in the serum.
Briefly: answers to topical questions
What types of agglutination reactions exist by method of implementation, besides the tube agglutination test?
In addition to the tube test (in test tubes), slide agglutination (on a glass slide), indirect agglutination, and other modifications are distinguished. The slide test is performed by mixing a drop of serum and a pure culture of the pathogen on a glass slide.
What is the indirect (passive) hemagglutination test (IHA)?
The indirect (passive) hemagglutination test is based on using erythrocytes as carriers to visualize the interaction of antigen with antibody. In the classic version for detecting antibodies, an antigenic erythrocyte diagnostic agent is used, consisting of erythrocytes with known antigens. In the reverse indirect hemagglutination test for detecting antigens, an antibody erythrocyte diagnostic agent with known antibodies is used.
What physicochemical environmental conditions are necessary for the optimal course of the agglutination reaction?
The optimal course of the agglutination reaction requires the mandatory presence of electrolytes, such as isotonic sodium chloride solution, as well as maintaining the optimal pH value of the medium.
Class and exam questions
What is a serum titer?
This is the highest dilution of the patient's blood serum at which a precipitate is still visually determined in the tube agglutination test.
Why are absorbed sera needed?
They are needed to exclude cross-reactions with related bacteria sharing common antigens. Such sera contain only specific antibodies against the target pathogen.
What else to study on this topic
- Slide agglutination test
- Indirect agglutination reaction
- Immunofluorescence, ELISA, and RIA methods (overview)
- Monoclonal antibodies in immunohematology
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