Fluorescent substances absorb a characteristic wavelength of that substance and then emit a fluorescent light with a wavelength longer than the absorbed light. The ratio of photons absorbed versus emitted as fluorescent light is referred to as thefluorescence quantum yield or quantum efficiency and is associated with the fluorescence intensity of fluorescent substances.Two methods are used to determine this ratio, either a relative or absolute method. The relative method involves comparison of fluorescence intensity to a standard fluorescent substance with a known ratio to calculate a relative value. In this article, the ratio determined using this method is referred to as fluorescence quantum yield. On the other hand, the absolute method involves calculating the ratio directly from a fluorescence spectrum measured from the fluorescent substance using an integrating sphere. In this article, the ratio determined by this method is referred to as fluorescence quantum efficiency. Theabsolute method only involves the measurement sample, which makes it easier than the relative method. This article describes using an RF-6000 spectrofluorophotometer with an integrating sphere attached to determine the fluorescence quantum efficiency andmeasure quinine sulfate.