Basic Principles and Types of Reactions
The core principle of serological testing involves detecting specific antigen-antibody complexes in the patient's blood serum. Depending on the clinical goal, the physician may search for either traces of the pathogen itself (antigens) or the immune system's response (antibodies).
To visualize and evaluate this interaction in microbiology, various classes of reactions are used:
- Classical tests: agglutination reactions (including modifications such as indirect hemagglutination [IHA] and hemagglutination inhibition [HI]), complement fixation test (CFT), neutralization test (NT), and precipitation reaction (PR).
- High-sensitivity methods: radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA). The latter provide maximum accuracy when detecting minimal concentrations of target molecules.
Antibody Detection: The Paired Sera Method
The classical approach for confirming an acute infection is the paired sera method. Its protocol requires strict adherence to time intervals and storage conditions for the biological material.
- The first blood sample is collected at the very onset of the disease (acute phase). The obtained serum must be stored in a refrigerator at 4–8 °C.
- The second sample is collected 10–14 days later.
- The golden rule: both sera must be tested simultaneously in a single laboratory run. This eliminates external variables and errors caused by different reagent lots.
Positive confirmation of a current infection is defined by seroconversion—a rise in antibody titer in the second sample by at least 4-fold compared to the first.
It is important to understand that this approach provides retrospective diagnostics. Laboratory results arrive two weeks after disease onset, by which time the patient is often already in recovery. An exception includes certain infections where a single test showing extremely high antibody titers is sufficient for diagnosis.
Determination of Immunoglobulin Classes
Differentiating synthesized antibodies by class (IgM, IgG, IgA) provides the clinician with valuable information regarding the nature of the immune response and the phase of the pathological process. It is also an excellent tool for distinguishing an active infectious disease from established post-vaccination immunity.
- Acute infection: blood is dominated by IgM antibodies. Early antibodies of other classes (such as low-avidity IgG specific to early pathogen proteins) may also be detected during this phase.
- Chronic process, secondary infection, or carrier state: characterized by the stable presence of IgG and IgA.
- Past infection (immunological memory): persistently low titers of IgG are detected without a rising trend.
A significant limitation of antibody-based serology is severe patient immunosuppression. In a suppressed immune system, the synthesis of protective proteins is sharply reduced, creating a high risk of false-negative results.
Antigen Detection and Test Reliability
Unlike antibody screening, detecting pathogen antigens allows for rapid diagnostics. Antigens can be identified in the earliest stages of infection, before the immune system has had time to produce its own antibodies. Quantitative antigen determination is actively used to monitor disease dynamics: a steady decline in concentration serves as a reliable indicator of therapeutic efficacy.
Regarding the reliability of serological methods, specificity ranges between 70% and 90%. Analytical errors are divided into two categories:
- False-positive results: occur due to non-specific molecular binding within the sample or cross-reactivity (when evolutionary similar antigens are present across different microbial species).
- False-negative results: most commonly associated with the concentration of target antigens or antibodies in the patient's blood falling below the detection threshold of the test system.