A non-peptide-based chemiluminescent probe for sensitively visualizing neutrophil elastase activity in vivo

As a prototypical serine protease, neutrophil elastase (NE) plays a pivotal role in the regulation of inflammation, and its dysregulated activation is closely associated with organ damage related to inflammation. Despite the existence of several NE-responsive fluorescent probes, they are still susceptible to interference from auto-fluorescence caused by real-time excitation light. In this study, we developed a chemiluminescent probe named NE-CL for the detection of endogenous NE activity both in vitro and in vivo. The NE-CL consists of an acrylate-substituted dioxetane scaffold, a self-immolative linker, and a perfluorinated propionamide substrate that acts as the recognition group. In the presence of NE, a significant amount of chemiluminescence is generated by NE-CL with an impressive signal-to-noise ratio (∼342-fold) and high sensitivity (LOD∼20.8 ng/mL). Furthermore, the exceptional selectivity and appropriate luminescence duration time of NE-CL render it suitable for in vivo sensing. More importantly, NE-CL has been successfully utilized to differentiate various cell lines based on endogenous NE activity and visualize NE expression levels in mice inflammation model. These finding suggest that NE-CL possesses great potential for highly sensitive and accessible detection of NE-related diseases in the future.