Naturstoff reagent A (diphenylboric acid 2-aminoethyl ester [DPBA]) has been used historically in plant science to observe polyphenolic pigments, such as flavonoids, whose fluorescence requires enhancement to be visible by microscopy. Flavonoids are common dietary constituents and are the focus of considerable attention because of their potential as novel therapies for numerous diseases. The molecular basis of therapeutic activity is only gradually being established, and one strand of such research is making use of the social amoeba Dictyostelium discoideum. We extended the application of DPBA to flavonoid imaging in these preclinical studies, and report the first method for use of DPBA in this eukaryotic model microbe and its applicability alongside subcellular markers. This in vivo fluorescence imaging provided a useful adjunct to parallel chemical and genetic studies.