LOV-based fluorescent proteins (FPs) are an alternative class of fluorescent reporters with unique properties which complement the well-established proteins of the GFP family. One of the most important features of LOV-based FPs is the independence of molecular oxygen for the development of their specific fluorescence. Furthermore, they are characterized by small size and rapid signal development. Over the last few years, a number of different bacterial and plant LOV-based fluorescent proteins such as FbFP, iLOV and miniSOG have been developed and optimized. In this report, we comparatively have characterized the photophysical properties of nine different LOV-based fluorescent proteins including the excitation and emission maxima, the extinction coefficient, the fluorescence quantum yield, the average fluorescence lifetime and the photostability. The unified characterization of the LOV-based FPs provides a useful guide to apply them as in vivo tools for quantitative analyses and biological imaging.