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 4x10(-12)-5x10(-11) mol l(-1). The detection limit for ALP is 1x10(-12) mol 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 alpha-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.