A simple radial diffusion technique is described for assaying deoxyribonuclease I (DNase I) activity in whole human plasma and serum. Enzymatic activity was calculated from a standard curve constructed from the studies on the hydrolysis of calf thymus DNA by bovine pancreatic DNase of known concentrations. The assay was shown to be specific for nucleases which are active at neutral pH and require magnesium ions for activation. There was no significant difference in DNase I activity in plasma compared with that in serum from normal individuals (P greater than 0.05). The DNase I activity of 35 normal human sera was 26.1 +/- 9.2 ng/ml and ranged from 10.8-48.5 ng/ml. This technique may prove useful for the evaluation of DNase I activity in crude biological fluids and is not affected by the presence of enzymatic inhibitors and/or activators.