ⓘ Cloned enzyme donor immunoassay

                                     

ⓘ Cloned enzyme donor immunoassay

A cloned enzyme donor immunoassay is a competitive homogenous enzyme immunoassay. This assay makes use of two component fragments of an enzyme which are each individually inactive. Under the right conditions in solution these fragments can spontaneously reassemble to form the active enzyme. For use in biochemical assays one of the enzyme fragments is attached to an analyte of interest. The analyte-enzyme-fragment-conjugate is still able to reassemble with the other enzyme fragment to form an active enzyme. However it is unable to do this if the analyte is bound to an antibody.

To determine the amount of analyte in the sample, an aliquot of the sample should be added to the solution containing the enzyme-fragment-analyte-conjugate, the other enzyme fragment, an antibody directed against analyte and a substrate for the enzymatic reaction. A competition for the antibody occurs between the analyte in the sample and the enzyme-fragment-analyte-conjugate. High concentration of analyte in the sample lead to a relatively small number of enzyme-fragment-analyte-conjugate that inhibits the formation of active enzyme and, therefore, high activity of the enzyme. Conversely, lower concentration of analyte in the sample lead to a relatively large amount of the enzyme-fragment-analyte-conjugate that inhibits the formation of active enzymes and consequently lower activity of the enzymes.