Fluorescence imaging plays an important role in researching the biological function of lipid droplets (LDs). However, the short-wave emission, tedious synthesis process and insufficient specificity have significantly limited the applications of commercially available probes. Herein, we have prepared a novel one-step synthesized near-infrared (NIR) fluorescent probe, TNBD, with a very low emission in aqueous solution and the solid state, but a significantly enhanced fluorescence emission is exhibited in oleic acid. Moreover, TNBD exhibited an impressive lipid droplet (LD) specific fluorescence turn-on ability in cells, fatty liver and atherosclerosis (AS) samples with a good biocompatibility and high signal-to-noise ratio. Our study not only establishes a novel LD turn-on fluorescence probe, but also provides a novel way to prepare a NIR LD targeted fluorescence probe.