Specific turn-on detection of enzyme activities is of fundamental importance in drug discovery research, as well as medical diagnostics. Although magnetic resonance imaging (MRI) is one of the most powerful techniques for noninvasive visualization of enzyme activity, both in vivo and ex vivo, promising strategies for imaging specific enzymes with high contrast have been very limited to date. We report herein a novel signal-amplifiable self-assembling (FNMR)-F-19/MRI probe for turn-on detection and imaging of specific enzymatic activity. In NMR spectroscopy, these designed probes are silent when aggregated, but exhibit a disassembly driven turn-on signal change upon cleavage of the substrate part by the catalytic enzyme. Using these F-19 probes, nanomolar levels of two different target enzymes, nitroreductase (NTR) and matrix metalloproteinase (MMP), could be detected and visualized by (FNMR)-F-19 spectroscopy and MRI. Furthermore, we have succeeded in imaging the activity of endogenously secreted MMP in cultured media of tumor cells by (FMRI)-F-19, depending on the cell lines and the cellular conditions. These results clearly demonstrate that our turn-on F-19 probes may serve as a screening platform for the activity of MMPs.