We have studied fluorescence in rubrene single crystals by use of fluorescence up-conversion, fluorescence anisotropy, and temperature-dependent time-resolved fluorescence techniques. Thermally activated singlet fission was demonstrated to play an important role in the quenching of two intrinsic fluorescence bands, 565 and 610 nm. At low temperatures, singlet fission is suppressed while another process, namely energy trapping, becomes pronounced. The 650 nm fluorescence originates from the hole trap states located 0.27 eV above the valence band. Rubrene (5,6,11, has attracted increasing attention in recent years due to its applications in organic light emitting diodes (OLEDs) 1 and organic field effect transistors (OFETs). 2 Rubrene single crystals are well known for their high hole mobility (up to 40 cm 2 /V s) 3,4 and photoconductivity. 5 Observations of singlet fission 6,7 and long exciton diffusion lengths 8 makes it potential to improve the efficiency of organic photovoltaics. Although there has been much done on the applications, some fundamental questions such as the origin of the fluorescence in rubrene single crystals remain controversial. In particular, in the case of steady-state fluorescence spectra, both experimental results and interpretations contradict depending on experimental conditions, e.g., the polarization of the excitation beam, the crystal quality, and the degree of photo-oxidation.Rubrene crystals grown by physical vapor transport (PVT) are orthorhombic, with space group Cmca [Figs. 1(a)-1(d)]. 9 The reported steady-state fluorescence spectra of rubrene single crystals can be summarized into three types [Fig. 1(e)]:(1) 565 nm type. For incident light polarized parallel to the c axis, a strong fluorescence band centered at 565 nm dominates the fluorescence spectrum, which is explained to be due to the M-polarized transition 10 or c-polarized emission. 11 When the excitation polarization is parallel to the ab facet, the 565 nm fluorescence band disappears, and instead, the fluorescence spectra are present in two other types: (2) 610 nm type. The fluorescence spectrum centered at 610 nm is usually considered as the fluorescence spectra from pristine rubrene. 12 (3) 650 nm type. Another type of fluorescence spectrum under b polarized excitation is centered at around 650 nm. It is disputed to be due to either the oxidation related defects, 10 the band gap states, 13 or the amorphous phase embedded in the crystal. 12 In this work, we discuss the origin of the steady-state fluorescence in rubrene single crystals by stating the important role of singlet fission and energy trapping. The fluorescence kinetics observed was on a picosecond time scale due to the quenching by singlet fission. Thus the majority (99.9%) of the intrinsic fluorescence is quenched due to singlet fission. At low temperatures, energy trapping is observed while the thermally activated singlet fission is suppressed. The hole trap states located 0.27 eV above the valence band produce the 650 nm fluorescence band. Based on the fl...