Absorbing-molecule-induced optical clearing enhances Cerenkov luminescence imaging of radiopharmaceutical distribution.

Cerenkov luminescence imaging (CLI) enables noninvasive and real-time tracking of the in vivo distribution of radiopharmaceuticals by detecting Cerenkov optical signals generated during radionuclide decay. However, biological tissues exhibit strong scattering and absorption in the visible spectrum, which not only markedly attenuate Cerenkov signals but also cause discrepancies between the detected optical signals and the actual in vivo distribution of radiopharmaceuticals, thereby limiting the imaging accuracy of CLI. Here, we demonstrate that skin optical clearing alleviates this limitation and enhances the spatial fidelity of CLI images. An absorbing-molecule-based optical clearing treatment markedly decreased the tissue reduced scattering coefficient, increasing visible light transmittance through mouse skin by more than 50-fold (from 0.47% to 24.55%) while reducing skin reflectance by approximately 68.8%. Using 68Ga as the radiotracer, the agreement between CLI and positron emission tomography (PET) was substantially improved, with the Pearson correlation coefficient increasing from 0.55 to 0.78 and the Spearman correlation coefficient from 0.63 to 0.82. After a 15-day recovery period following optical clearing treatment, the treated mouse showed hair regrowth and signs of skin recovery. These results support the potential of optical clearing as a simple and effective strategy for improving the spatial correspondence of CLI with radiopharmaceutical distribution.