The biology of highly reactive oxygen radical species is of great interest in many biomedical research disciplines, including neurodegeneration, aging, cancer, and infectious diseases.[1] There are a number of fluorescent reagents, such as 2,7-dichlorodihydrofluorescein (DCDHF), that can be used to detect free radicals, but they have significant limitations due to their facile oxidation by light and numerous non-radical oxidants such as hydrogen peroxide (H2O2). [2] HPF is a cell-permeable aromatic amino-fluorescein derivative that has little intrinsic fluorescence. [3] It undergoes oxidation only by highly reactive oxygen species (hROS) such as the hydroxyl radical, peroxynitrite, and hROS generated from a peroxidase/H2O2 system. It is inert to hypochlorite ion, nitric oxide, hydrogen peroxide (H2O2), superoxide, and other oxidants. Upon oxidation, HPF is converted to the highly fluorescent molecule fluorescein, with excitation/emission maxima of 490/515 nm, respectively, allowing the simple direct detection of highly reactive biological radicals.