Chemiluminescent flow injection determination of alkaline phosphatase and its applications to enzyme immunoassays

A Chemiluminescent flow injection method based on the reaction of cortisol-21-phosphate with alkaline phosphatase (ALP) is described. The cortisol produced by the enzyme-catalyzed reaction reacts with lucigenin (N,N′-dimethyl-9,9′-biacridinium dinitrate) in basic aqueous solution to give intense chemiluminescence. The method allows ALP to be determined in the range 4 × 10−12–5 × 10−11 mol 1−1. The detection limit for ALP is 1 × 10−12 mol 1−1. Experimental conditions, such as flow rates, pH, incubation time and reactant concentrations have been studied. The applications of the method to enzyme immunoassay were demonstrated by the determination of 17 α-hydroxyprogesterone (17-OHP) and human chorionic gonadotropin (hCG) in urine or serum samples using ALP as the enzyme label. Good correlations were obtained between the results of serum hCG or 17-OHP assay by the proposed method and other methods.