OBJECTIVE Measurement of serotonin concentrations in blood is considered to be important for the clinical studies of diseases involving vascular endothelial injury. We have developed a high-performance liquid chromatographic (HPLC) method for measurement of serotonin level in platelet-poor plasma (PPP). METHODS Venous blood samples were collected using vacuum tubes containing ethylenediaminetetraacetic acid (EDTA) dipotassium salt; the optimum concentration in blood was 7.4 mmol/l. Serotonin in samples was separated by HPLC with a column-switching system, and was specifically converted into a fluorescent derivative with benzylamine for convenient detection. RESULTS The between-day assay coefficient of variation for a serotonin level (5.1 nmol/l) in PPP was 6.2%. The mean concentration of serotonin in PPP in healthy subjects was 5.7 +/- 3.0 nmol/l. The serotonin levels in PPP in patients with ischemic heart disease were significantly higher than those in healthy subjects (P = 0.001). CONCLUSIONS The developed method had sufficient performance to determine serotonin levels in PPP from patients with ischemic heart disease.