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Artificial biomaterials can significantly increase the rate of tissue regeneration. However, implantation of scaffolds leads not only to the accelerated tissue healing, but also to an immune response of the organism, which results in the degradation of the biomaterial. The synergy of the immune response and scaffold degradation processes largely determine the efficiency of the tissue regeneration. Still, methods suitable for fast, accurate and non-invasive characterization of the degradation degree of biomaterial, which are applicable both for in vitro and in vivo, are highly demandable. In this work we show the possibility of monitoring of the degradation of decellularized bovine pericardium scaffolds under conditions mimicking the immune response and oxidation processes using multiphoton tomography combined with fluorescence lifetime imaging (MPT-FLIM). We found that the average fluorescence lifetimes of genipin-induced crosslinks in collagen and oxidation products of collagen are prominent markers of oxidative degradation of scaffolds. This was verified in model experiments, where the oxidation was induced with hypochlorous acid or by exposureexposed to activated neutrophils. The fluorescence decay parameters also correlated with the changes of micromechanical properties of the scaffolds as assessed using atomic force microscopy (AFM). Our results suggest that FLIM can be used for quantitative assessments of the properties and degradation of the scaffolds essential for the wound healing processes in vivo.