抄録
A sensitive and separative fluorometric analysis of alkaline phosphatase isoenzyme was described. Electrophoretical separation of alkaline phosphatase isoenzyme was based on acetate gel (Cellogel) electrophoresis with the discontinuous buffer system, and the locarization of alkaline phosphatase isoenzyme was fluorometrically visualized with naphthol ASMX-phosphate as substrate. The liberated naphthol ASMX which has a strong fluorescence at 505nm on the acetate gel were scanned by scanning fluorophotometer, and the peak areas obtained from scannings were proportional to the activities of each separated isoenzyme. The standard curves were linear in the rang of 5 to 640 U/1. The coefficient of variation of this method as a quantitative analysis of enzyme separated was 3 to 5 per cent in the same membrane and 8 to 10 per cent in the different membranes.
Using this method, the kinetical inhibition studies of alkaline phosphatase isoenzymes were easilly carried out by the substrate solution containing different concentration of inhibitors, and the kinetical values of the enzymes in the solution of enzyme mixture were clearly coincided with the kinetical values obtained from the purified enzymes. The usefullness and application of this sensitive microtechniques in clinical examination was studied and discussed.
