Tumor Stiffness‐Enhanced Room Temperature Phosphorescence Imaging Using Dual‐Ligand Nanoprobes

Organic room temperature phosphorescence (RTP) materials offer high tumor‐to‐background imaging contrast owing to their long afterglow and minimal autofluorescence interference. In this work, a targeted phosphorescent nanoprobe is prepared based on a pyrene/benzophenone (PyBP) guest–host system and further functionalized it with folic acid and cyclic Arg‐Gly‐Asp peptide for enhanced tumor specificity. In oral squamous cell carcinoma (OSCC) xenograft models, this dual‐ligand nanoprobe can help achieve a four–fivefold increase in the tumor signal over the background. Notably, the previously unexplored relationship between tumor stiffness and RTP emission is studied. Mechanical and histological analyses confirmed increased stiffness in OSCC tissues. By modulating the rigidity of tumor phantoms and treating the xenografts with collagenase, a direct correlation is revealed between tumor stiffness and phosphorescence intensity. This study suggests the effectiveness of dual targeting in improving tumor visualization using guest–host‐doped RTP nanoprobes. The findings demonstrate that the biomechanical features of the tumor microenvironment enhance phosphorescence emission, highlighting tumor stiffness as a natural amplifier for RTP imaging.