To demonstrate the capability of CLP nanoparticles for deep-tissue imaging, the relative bioluminescence intensity of CLP nanoparticles and luminol was compared. Briefly, CLP nanoparticles (0.5 mg/ml) or luminol in aqueous solution was mixed with 50 μM H2O2 and 15 mU MPO with or without 50 mM Cl in a black 96-well plate, which was covered with porcine muscle tissue with a thickness of 3 mm. The same samples without covering with muscle tissue were used as controls. In another study, the in vitro luminescence penetration threshold of CLP nanoparticles was examined. To this end, CLP nanoparticles (1 mg/ml) in aqueous solution was mixed with 100 μM H2O2, 30 mU MPO, and 100 mM Cl in a black 96-well plate, which was covered with muscle tissues with different thicknesses. In all cases, the bioluminescent signals were quantified by an IVIS Spectrum imaging system (exposure time = 5 min, f/stop = 1, binning = 8, FOV = 6.6 cm).

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