The fluorescence imaging (FLI) in the second near-infrared window (NIR-II, 1000–1700nm) has attracted considerable attention in the past decade. In contrast to conventional NIR-I window excitation (808nm/980nm), FLI with NIR-II window excitation (1064nm/other wavelength beyond 1000nm) can afford deeper tissue penetration depth with high clarity due to the merits of suppressed photon scattering and diminished autofluorescence. In this review, we have summarized NIR-II window excitable/emissive organic/polymeric fluorophores recently developed. The characteristics of these fluorophores such as chemical structures and photophysical properties have also been critically discussed. Furthermore, the latest development of noninvasive in vivo FLI with NIR-II excitation was highlighted. The ideal imaging results emphasized the importance of NIR-II excitation of these fluorophores in enabling deep tissue penetration and high-resolution imaging. Finally, a perspective on the challenges and prospects of NIR-II excitable/emissive organic/polymeric fluorophores was also discussed. We expected this review will be served as a source of inspiration for researchers, stimulating the creation of novel NIR-II excitable fluorophores and fostering the development of bioimaging applications.