what is enzyme immunoassay?

Answer 1

An assay that uses an enzyme-bound antibody to detect antigen. The enzyme catalyzes a color reaction when exposed to substrate.


An immunoassay is a biochemical test that measures the level of a substance in a biological liquid, typically serum or urine, using the reaction of an antibody or antibodies to its antigen. The assay takes advantage of the specific binding of an antibody to its antigen. Monoclonal antibodies are often used as they only usually bind to one site of a particular molecule, and therefore provide a more specific and accurate test, which is less easily confused by the presence of other molecules. The antibodies picked must have a high affinity for the antigen (if there is antigen available, a very high proportion of it must bind to the antibody).

Answer 2

Usually, antiobdies (I will call these antibodies A1) are used which bind to a specific antigen of interest that has been affixed in some manner to a medium -- usually the assay plate. Then, the assay is "washed", to remove unbound antibodies that have not attached to antigen. After that, secondary antibodies are added (I will call them A2), which will bind to the A1 that are bound to the antigen. Then, it is washed again to remove antibodies that have not bound anything.

Now, connected to the A2-antibodies are enzymes that produce some kind of pigment, so that when the the enzyme substrate is added, pigment is produced in proportion to how much antigen has been bound.

The reason for using 2 different types of antibodies is that the first antibody (A1) is specific for that antigen, and produced ONLY to be used in that particular scenario (because there are literally trillions of antigens in the world). The second (A2) antibody has had the enzyme synthetically attached to it, and the A2 antibody will bind to ANY other antibody, and may thus be used in other assays, not just that particular scenario. So, as you can see, it is more efficient to not have to attach enzymes to every A1 antibody you want to use (expensive thing to do) -- it's much more cost and time-efficient to use A2 antibodies with the enzyme already attached, and then all you have to do is get the A1 antibodies that are specific for whatever antigen you're interested in.