Initial Diagnostics (Screening)
The first stage of laboratory evaluation is screening. Its primary objective is to identify potentially infected serum samples for subsequent in-depth analysis.
ELISA (Enzyme-Linked Immunosorbent Assay) is the primary method used at this stage. The goal of this test is twofold: test systems are designed to simultaneously detect specific human antibodies against the virus as well as the viral p24 antigen. Although this combined approach increases the reliability of the initial diagnostic tier, a positive ELISA result is not a definitive diagnosis and requires mandatory verification.
Confirmatory Diagnostics (Reference Method)
Samples that test positive during screening are sent for confirmatory testing. Western blotting serves as the reference method that finalizes the microbiological diagnosis. This method allows for the detailed separation of viral proteins and the detection of antibodies directed against specific structural components of the pathogen.
A strict criterion for establishing a positive diagnosis is the detection in the patient's serum of antibodies against at least two viral envelope glycoproteins. These surface proteins are encoded by genes of the env group. A crucial rule of the algorithm is that the diagnosis is confirmed based on envelope proteins regardless of whether antibodies against core virion proteins (gag group) or viral enzymes (pol group) are present in the patient's serum.
The Role of PCR in the Testing Algorithm
Molecular genetic methods, such as polymerase chain reaction (PCR), play an important but strictly auxiliary role in the microbiological algorithm. PCR is definitively not used for primary diagnostics. The main reason for this lies in the extreme genetic variability of the virus—due to constant mutations, primers may fail to recognize the altered genome, leading to false-negative screening results.
PCR is only utilized after the diagnosis has been definitively confirmed via serological methods. The method is necessary to address three clinical tasks:
- Viral load quantification: measuring the number of viral copies helps predict disease progression and assess the severity of the infectious process.
- Subtype identification: determining the specific genetic variant of the circulating virus.
- Resistance testing: screening for mutations that render the virus resistant to antiretroviral drugs.
Immunological Monitoring
Because the virus systematically destroys the patient's immune system, microbiological diagnosis is inextricably linked with immunological monitoring. Clinicians must continuously assess the degree of preservation of the body's defenses.
The basis of monitoring is the regular enumeration of key T-lymphocyte subpopulations—cells bearing CD4 and CD8 receptors. A decline in the CD4+ T-cell count is considered the primary marker of progressive immunosuppression. If this parameter drops below the critical threshold of 500 cells/µL, it indicates pronounced immune depression and necessitates immediate adjustment of the patient management strategy.