A simple, sensitive, non-stimulated assay was developed to measure the superoxide anion concentration in whole blood, using an ultra-sensitive chemiluminescence (CL) analyzer and lucigenin amplification. The assay system can be performed without leukocyte isolation or stimulant administration. The blood CL levels of healthy males (362.8 +/- 337.7 counts /10 sec) were not different from those of females (335 +/- 308.7 counts/10 sec) (p=0.64), whereas the CL levels in whole blood in patients with acute pancreatitis (2522 +/- 2014 counts/10 sec) were significantly higher than those of healthy controls (p<0.001). This assay system may be valuable in the future for quantitative measurement of reactive oxygen species in various disorders.