ENHANCED CHEMILUMINESCENCE ASSAY FOR PEROXIDASE

Horseradish peroxidase can be assayed chemiluminescently using a mixture of luminol and an oxidant, eg hydrogen peroxide or perborate. Assay characteristics can be dramatically improved by the addition of small amounts of enhancers. These substances, eg, para-iodophenol, para-hydroxycinnamic acid, and various naphthols and aromatic amines increase the light emission from the reaction by several orders of magnitude. In addition, the reagent background is diminished which contributes to the large increase in signal to background in enhanced chemiluminescence (ECL) assays, The detection limit for peroxidase is 50 attomoles and the assay has been used in a wide range of assay formats (tube, bead, particle, microtiter well, membrane, dipstick). A large number of immunoassays have been developed (peptide and steroid hormones, drugs, antibodies, cancer markers) and implemented on a semi-automatic analyzer. The long-lived signal is ideal for automated instruments because initiation and measurement of light emission can be separated. It is also ideally suited to photographic assays and these have been used in extra-laboratory applications. More recently, ECL assays have been exploited in DNA hybridization assays.