Immunoassay

Immunoassays are based on the principle that a particular antigen will stimulate a very specific (unique) immune response, and proteins (called antibodies) produced through an immune response can be used to detect the presence of a target compound in a sample. Immunoassays are fast and accurate tests used to check biological systems by tracking different proteins and antibodies. Immunoassays rely on the ability of an antibody to bind to a specific molecular structure and can be used to detect specific molecules in the laboratory.

Labeled Immunoassay

1. Radioimmunoassay(RIA)

RIA is probably the oldest type of immunoassay. The radioactive isotope is used to label the antibody/antigen. The amount of radioactive signals is inversely proportional to that of target antigens.

2. Counting immunoassay (CIA)

In CIA, polystyrene beads are coated with a number of antibodies that are complementary to the target antigens. During incubation, the beads bind to a variety of antigens and jointly form a large mass, but some beads are not bound. The whole solution passes through a cell counter, with only unbound beads counted. The amount of unbound beads is inversely proportional to that of antigens.

3. Enzyme immunoassays (EIA) or enzyme-linked immunosorbent assays (ELISA)

In the ELISA, the antibody is linked to an enzyme. After incubation with the antigen, the unbound antibody is eluted. The bound antibody-enzyme linked to the target antigen is observed by adding substrates to the solution. The enzyme catalyzes the chemical reactions of the substrate to produce quantifiable color changes.

4. Fluorescence immunoassay (FIA)

In FIA, antibodies are labeled with fluorescent probes. After incubation with the antigen, the antibody-antigen complex is isolated and the fluorescence intensity is measured.

5. Chemiluminescence immunoassay (CLIA)

CLIA is the same as ELISA or fluorescent immunoassay, but its reporter gene is different. CLIA combines the CL system with an immune response. Some reagents are used as CL labels. The system produces chemiluminescence after the introduction of the CL substrate. Therefore, the sample can be quantitatively detected. The most commonly used CL substrates are luminol, isoluminol and its derivatives, acridinium ester derivatives, peroxidase and alkaline phosphatase (ALP).

6. Immunochromatographic analysis (ICA)

ICA, also known as lateral flow immunoassay, is a rapid testing immunoassay for detecting the presence (or absence) of a target analyte in a sample (matrix). In this assay, the capture antibody is immobilized as a crossover wire on a porous hydrophilic material. The analyte in the buffer is then added to one side of the test strip and driven by lateral capillary forces. The assay flows through the capture antibody line and is captured by the antibody. Finally, the complex can be revealed by a nanoparticle label (usually colloidal gold).

7. Liposome immunoassay (LIA)

LIA is an assay involving liposome encapsulation markers. In LIA, liposomes are prepared and then coupled to the analyte or antibody by a suitable method and then assayed in the normal manner. Detection in LIA relies on liposome cleavage and the release of the encapsulated label, which is then measured and correlated with analyte concentration.

Label-free Immunoassay

Although certain markers are commonly used in immunoassays, certain types of assays do not rely on markers. Instead, detection methods that do not require modification or labeling of the analytical components are employed. Surface plasmon resonance (SPR) and quartz crystal microbalance (QCM) are two typical unlabeled methods.

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