Dihydrorhodamine 123 (DHR) attached to membranes of granulocytes (PMN) and monocytes is caused to fluoresce by reactive oxygen intermediates (ROI) indicating the ability of phagocytes to produce these microbicide metabolites in a flow microcytofluorimeter. Whole blood samples from five boys with known chronic granulomatous disease (CGD) and from their mothers (and from one father and one grandmother), were examined following erythrocyte lysis in order to test this new method. An incubation period of 10 min with phorbol-myristate-acetate, followed by another 15 min incubation period with DHR before flow microcytofluorimetric analysis of 5 or 10 x 10(3) phagocytes, was sufficient to obtain the following results. PMN and monocytes from four patients with CGD could clearly not produce any ROI whereas cells from one patient displayed decreased activity in ROI production as compared to cells from a healthy donor. The X-linked mode of inheritance was detected in six carriers by the presence of two different cell populations (one normal ROI-producing and one negative or less active population). All the phagocytes from one mother produced ROI in normal amounts suggesting an autosomal mode of inheritance. All in all, the method presented provides a fast and most simple tool to diagnose CGD, to determine a decrease or total lack of ROI production and to establish the mode of inheritance of the disease.