IVIS fluorescence imaging

ApoE^−/− mice fed a Western diet for 3.5 months received IV injections of p5RHH-Cy5.5-labeled siRNA nanoparticles or control nanoparticles without fluorescence labeling. At 24 hours after injection, aortas were collected for ex vivo IVIS imaging (Xenogen IVIS Spectrum; PerkinElmer, Waltham, MA, USA). The following settings were used for image acquisition: excitation 675 nm, emission 720 nm, binning factor 8, field of view 12.9, exposure time 0.5 seconds, and f-value 2. The pseudocolor “efficiency image” was presented to illustrate fluorescence-labeled nanoparticle distribution in the aortas. Fluorescence data were calibrated quantitatively in units of radiant efficiency or photon flux per unit excitation intensity (photons/sec/cm²/steradian)/(μW/cm²).