Antigen tests: Methodology review
As previously indicated, antigen tests primarily detect the presence of specific viral antigens (protein markers) found on the outside of the SARS-CoV-2 virus. Antigen tests have similar specificity to molecular tests such as RT-PCR but are less sensitive. However, they are relatively less expensive, can be performed at point-of-care (POC), generate results quickly, and are amendable for use in screening programs to identify those likely to be contagious. Because of the decreased sensitivity of antigen tests, it may be necessary to confirm some antigen test results (e.g., a negative test in persons with symptoms or a positive test in persons without symptoms) with a molecular-based test (NAAT).
Typically, antigen tests are performed on nasopharyngeal or nasal swab specimens and use immunoassay methodology to detect the presence of a specific viral antigen. The main types of immunoassays used for the detection of SARS-CoV-2 include enzyme-linked immunosorbent assay (ELISA), chemiluminescent immunoassay (CLIA), and lateral flow immunoassay (LFIA). For the SARS-CoV-2 virus, antigen tests can often be used to indicate current COVID-19 infection.
The following is a brief description of immunoassays, ELISA, CLIA, and LFIA methodologies. Also, included is a description of the sandwich ELISA methodology, which is one of the most common types of ELISA methods used for SARS-CoV-2 detection.
Immunoassays:
In general, immunoassay methods are based on the principle of specific antibody to antigen binding. They can be used to either identify specific antibodies or specific antigens related to the pathogen of interest. For antigen testing, immunoassays use antibodies to detect and measure a specific antigen or antigens related to the pathogen. Many viruses including the SARS-CoV-2 virus can be directly identified via the targeting of specific viral proteins or antigens.
Although the key feature for all immunoassays is the binding of an antibody to its antigen, the other important aspect is the ability to produce a measurable signal in response to the binding. Most immunoassays measure binding using chemically linking antibodies or antigens with some kind of detectable label. Several different types of labels can be used in immunoassays including enzymes that produce a color change or produce light (chemiluminescence) and fluorescent agents which fluoresce and emit light.
The most popular detection labels for use in many immunoassays are enzymes. An immunoassay that uses an enzyme is referred to as enzyme immunoassay (EIA). One of the most popular EIA methods in general use, and also used for COVID-19 virus detection, is the enzyme-linked immunosorbent assay (ELISA) which is described in the following section.
