Principle of Immunoassay

The term immunoassay refers to any method for measuring the concentration or amount of analyte in solution based on specific interaction between the antibody and the targeted antigen. This method (1) requires that the recognized analyte be physically separated from the residual biomolecules, and (2) employs a labeled antibody using radiolabel, fluorescent dye, enzyme, chemiluminescent molecule, or other label to measure or estimate the bound targets (Fig. 3). This method is termed competitive when the amount of measurable bound label is inversely proportional to the amount of analyte originally in solution. It is termed non-competitive when the amount of measurable bound label is directly proportional to the amount of analyte originally present in the biological sample.

Large magnetic particles (10-100 pm) have been used in radioimmunoassays (RIAs) to detect and quantify nortriptyline, methotrexate, digoxin, thyroxine, and human placental lactogen. Such large particles must be stirred carefully to avoid sedimentation phenomena. Smaller particles, such as hollow glass or polypropylene or magnetic core (2-10 pm), have also been used and were evaluated in estradiol RIA, with nondrastic problems related to the particle sedimentation [32]. Nowadays various biomedical kits are available that are based on immunodetection and magnetic beads and either enzyme immunoassays (EIAs) or RIAs [34].

FIG. 3 Schematic illustration of immunoassay using labeled antibody.

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