Shedding light on imaging safety: Decoding the origin of photocytotoxicity in RhB‐assisted fluorescence imaging

Photocytotoxicity represents a significant limitation in the application of dye-assisted fluorescence imaging (FI), often resulting in undesirable cellular damage or even cell death, thereby restricting their practical utility. The prevalence of Rhodamine B (RhB) in FI underscores the importance of elucidating its photocytotoxicity effects to minimize photodamage. This study identifies the primary cause of photocytotoxicity stems from the generation of cytotoxic singlet oxygen in RhB, utilizing femtosecond transient absorption spectroscopy coupled with quantum chemical calculations. The Laser power-dependent cellular viability reveals a threshold at about 50 mW cm