General Characteristics and Types of Reactions
All serological reactions in this group are based on a strict rule: complement molecules participate only when a specific antibody finds its antigen and binds to it. Until this moment, the complement remains inactive.
In laboratory practice, three main types of such reactions are distinguished:
- Complement Fixation Test (CFT). This is the most complex assay, requiring extremely rigorous preparation of all components before analysis.
- Radial Hemolysis (RH). This method is technically based on the diffusion of components in a specialized gel.
- Immune Adherence (IA). Designed for working with large particulate antigens and evaluating their interaction with cells.
Mechanism of the Complement Fixation Test (CFT)
CFT is a multi-step assay. It requires two independent systems of components introduced sequentially.
The first (primary) system includes:
- Known antigen (Ag);
- Patient's test serum (which may or may not contain the target antibodies);
- Exogenous complement (C).
The second (indicator or hemolytic) system is necessary for visualizing results. It consists of:
- Sheep erythrocytes;
- Hemolytic serum (specific antibodies directed against sheep erythrocytes).
The essence of the method lies in the competition for complement between the primary and indicator systems.
Interpretation of CFT Results
The test result depends on whether the complement was consumed (fixed) by the immune complexes from the patient's serum during the first stage. Two scenarios are possible:
Scenario A: Positive Reaction (Patient Has the Disease)
- In the first stage, the patient's serum contains specific antibodies.
- They bind to the antigen, forming a stable immune complex.
- The added complement recognizes this complex and firmly binds to it (is consumed).
- When the hemolytic system is added in the second stage, no free complement remains in solution.
- Outcome: No hemolysis occurs. Sheep erythrocytes remain intact and simply sediment to the bottom of the tube. The diagnosis is confirmed.
Scenario B: Negative Reaction (Patient Is Healthy)
- A healthy person's serum lacks the required antibodies.
- No immune complex forms in the first stage.
- All added complement remains completely free.
- After adding the indicator system, free complement attacks the sensitized sheep erythrocytes.
- Outcome: Total destruction of erythrocytes occurs. Pronounced hemolysis is observed, and the liquid takes on a translucent, bright red appearance ('lake blood'). The diagnosis is not confirmed.
Immune Adherence (IA)
This reaction allows the visualization of interactions between large objects. Its essence lies in the activation of complement by particulate antigens, such as whole bacterial cells or viruses pre-treated with immune serum.
Step-by-step mechanism of IA:
- Primary complex formation occurs between the particulate antigen and the antibody.
- This complex activates the complement system, leading to massive generation of the C3b component.
- C3b molecules deposit on the surface of the complex and begin to function as a biological 'glue'.
Result: The particulate antigen, coated with C3b molecules, adheres to the membranes of target cells bearing corresponding receptors. Such cells include human erythrocytes, platelets, and macrophages. Visually, mixing these cells with immune complexes causes them to clump together, resembling a classic agglutination reaction.