Basics

Immunoassays are a very important type of biochemical test. Basically, an immunoassay uses antibodies against a particular compound (usually a protein). If these antibodies are of high enough quality they will bind to this chemical, and only to this chemical, the antibodies are then in turn bound to a detectable indicator, such a fluorescent chemical, or radioactive isotope, finally some method is used to separate the bound antibodies from the rest of the sample.

ELISAs

The most common type of immunoassay is the Enzyme-Linked ImmunoSorbant Assay, or ELISA, which is essentially a refinement of the basic immunoassay mentioned above. A specially treated plate is coated with antibodies for the chemical, then the samples are added. The first set of antibodies binds the chemical to the plate. Then the plate is washed so all the chemicals except the bound target chemical are removed. Then antibodies against the first set of antibodies, which are bound to an enzyme, often horseradish peroxidase. The second antiboy binds to the first and the plate is washed again. Finally a chemical is added to the sample with is catalyzed by the enzyme to produce a coloured product. The amount of colour can then be read by a spectrophotometer. The main purpose of the enzyme is to amplify the results of the immunoassay: normal immunoassays require more sensitive instruments to detect small changes in a single-stage detection system such as fluorescent labelling, ELISAs require a relatively cheap plate reader, and some results are obvious to the naked eye.

Advantages

The major advantage of immunoassays, and in particular ELISAs are their specificity: a properly designed immunoassay can detect small amounts of protein in blood, urine and just about anything else you want to try. ELISAs are also cheap compared to other ways of analyzing a sample, and can be designed to be very easy to use, so they can be sold as single-use diagnostic tests. In fact, some forms of home pregnancy test use immunoassay techniques because of this.

Disadvantages

The major disadvantage of all immunoassays is the difficulty of getting good quality antibodies, as the accuracy of the entire reaction depends upon them binding properly to their target. This is a particular problem when there is no antibody for a particular chemical, or when the only one available binds with other chemicals (cross-reaction). Also experimental immunoassays are apparently one of the hardest types of assay to automate, an increasingly important concern in biochemistry.

Uses

Although most immunoassays are used in research, they are now increasingly used as diagnostic tests, for example for HIV, and the most well-known use of an immunoassay is the home pregnancy test.

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