A simple naphthalimide-based fluorescent probe was designed and synthesized for the determination of mercury ion (Hg2+ ). The probe showed a noticeable fluorescence quenching response for Hg2+ . When added with Hg2+ , the fluorescence intensity of the probe at 560 nm was remarkably decreased with the color changed from yellow to colorless under ultraviolet (UV) light. The probe had a notable selectivity and sensitivity for Hg2+ and displayed an excellent sensing performance when detecting Hg2+ at low concentration (19.5nM). The binding phenomenon between the probe and Hg2+ was identified by Job's method and high-resolution mass spectrometry (HRMS). Moreover, the probe was not only utilized to identify Hg2+ in real samples with satisfactory results (92.00%-110.00%) but also was successfully used for bioimaging in cells and zebrafish. The recognition mechanism has been verified by transmission electron microscopy (TEM) for the first time. All the results showed that the probe could be used as a potent useful tool for detection of Hg2+ .