Males of sapphirinid copepods use regularly alternating layers of hexagonal-shaped guanine crystals and cytoplasm to produce spectacular structural colors. In order to understand the mechanism by which the different colors are produced, we measured the reflectance of live individuals and then characterized the organization of the crystals and the cytoplasm layers in the same individuals using cryo-SEM. On the basis of these measurements, we calculated the expected reflectance spectra and found that they are strikingly similar to the measured ones. We show that variations in the cytoplasm layer thickness are mainly responsible for the different reflected colors and also that the copepod color strongly depends on the angular orientation relative to the incident light, which can account for its appearance and disappearance during spiral swimming in the natural habitat.