Stable and General-Purpose Chemiluminescent Detection System for Horseradish Peroxidase Employing a Thiazole Compound Enhancer and Some Additives

Abstract A stable and highly sensitive chemiluminescent detection system for horseradish peroxidase (HRP)/luminol/hydrogen peroxide using a newly designed thiazole compound enhancer has been established. Some additives for the chemiluminescent reaction were explored to overcome some defects of the reaction such as rapid decay and high background of light emission. Recrystallization of luminol and the addition of several detergents into the reacting solution were effective to increase specific light emissions. The addition of skim milk into the reacting solution reduced the background. Consequently, skim milk combined with a detergent increased the signal to noise ratio about 20 times compared with the reactions in the absence of both additives. The optimal concentration of enhancer and the addition of egg albumin stabilized the emission. In the new method, 6 × 10 −18 mol of HRP was detectable. This would be the most sensitive enhanced chemiluminescent detection system for HRP. Furthermore, we could detect picogram per milliliter (10 −17 mol) concentrations of a trace component in biological materials such as endothelin-1 by employing this reaction.