Neutrophils secrete proteases at inflammation sites leading to protease/inhibitor imbalance. Among them, neutrophil elastase (NE) is responsible for lung degradation via elastin fragmentation. Monitoring protease/inhibitor status non-invasively would be an important diagnostic tool. We present Meo-Suc-(Ala)2-Pro-Val-nitroxide, a line-shifting elastase activity probe for Electronic Paramagnetic Resonance (EPR) spectroscopy and Overhauser-enhanced Magnetic Resonance Imaging (OMRI). Fast and sensitive with Km= 15 ± 2.9 µM, kcat/Km= 930000 s-1.M-1, this substrate was assessed with bronchoalveolar lavage samples from Pseudomonas pneumonia mouse model. We observed a clear difference between wild type and NE deficient animals. These results can lead to new in vivo diagnostic methods and lung protection.