Design of a NIR-I Viscosity-Sensitive Fluorescent Probe for Tracking Nuclear Physiological Changes During Different Cell Cycle Stages

Organic small-molecule fluorescent probes represent an important class of fluorescence imaging tools that play crucial roles in cellular imaging. In this study, we designed a novel nonconventional viscosity-sensitive probe, <b>YY-2</b>, to facilitate live-cell nuclear tracking through intermolecular interactions, such as weak hydrogen bonding between the aniline group and thymine, electrostatic attraction, and π-π stacking. The DNA binding energy of <b>YY-2</b> (-5.24 kJ/mol) is comparable to that of the commercial nuclear dye Hoechst 33342 (-10.98 kJ/mol), while exhibiting lower DNA toxicity. Computational results indicate that the aminophenyl rotor contributes more significantly to the excited-state relaxation process than its nitrophenyl and phenyl counterparts. ESP analysis reveals that <b>YY-2</b> exhibits stronger positive charge, which facilitates its binding to DNA grooves via electrostatic attraction. <b>YY-2</b> intercalates into the groove adjacent to the DNA base pairs, thereby achieving a fluorescence turn-on effect (λ_ex = 610 nm, λ_em = 706 nm). It enables rapid nuclear staining of live cells within 15 min at a low concentration of 0.2 μM in a wash-free manner. In various cell types (including KYSE150 and SiHa cells), <b>YY-2</b> allows for discrimination of nucleoli, chromosomes, nuclear membranes and cytoplasm based on fluorescence brightness, and enables spatiotemporal monitoring of nuclear changes during cell growth.