A high-pressure liquid chromatographic procedure using fluorescence detection for the analysis of prazosin in whole blood and plasma is described. It employs a simple and rapid method of sample preparation. Prazosin and an internal standard are chromatographed as ion-pairs with pentane sulfonic acid. The method is sensitive and reproducible with accurate detection as low as 0.2 ng/ml in whole blood and 0.5 ng/ml in plasma with a coefficient of variation of 5.6% and 5.4% respectively. If propranolol or quinidine is present, modifications of chromatographic conditions are used which separate prazosin from these two drugs. Prazosin is stable when frozen for a long period or refrigerated for short periods.