Two recent papers in this Journal address the issue of applying the dichlorofluorescein assay to measure so-called reactive oxygen species (ROS) after irradiating cells, with diametrically opposed conclusions (1 ,2). A number of related earlier papers [e.g. ref. (3)] are based on this assay, which relies on the oxidation by ROS of dichlorodihydrofluorescein (DCFH2) to fluorescent dichlorofluorescein (DCF). While not seeking to adjudicate in the argument (1 ,2) of whether the assay as applied in individual protocols is measuring species originating in extracellular medium or not—more experimental studies are desirable—it does seem remarkable that in most of the over 2000 studies in biology using this assay, little attention has been paid to the chemical parameters that define what the assay reports. Most notable is the general lack of discussion as to which ROS...

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