Aggregation-induced emission-twisted intramolecular charge transfer-activated fluorescent probe for analyzing mitochondrial viscosity in cells and zebrafish.

Mitochondria are crucial energy-supplying organelles that support cellular activities and play vital roles in cell metabolism, aging, autophagy, and apoptosis. Abnormal viscosity can alter the mitochondrial microenvironment, disrupt normal mitochondrial function, and lead to disease. To address this, we designed and developed two aggregation-induced emission-twisted intramolecular charge transfer fluorescent probes, namely, (E)-1,1,3-trimethyl-2-(4-(1,2,2-triphenylvinyl)styryl)-1H-benzo[e]indol-3-ium (HSL-1) and (E)-2-(4-(di-p-tolylamino)styryl)-1,3,3-trimethyl-1H-benzo[e]indol-3-ium (HSL-2). In vitro fluorescence detection revealed that both HSL-1 and HSL-2 were sensitive to viscosity and demonstrated a strong log-linear relationship, with linear coefficients of 0.982 and 0.980, respectively. Notably, the responses of HSL-1 and HSL-2 to viscosity changes were unaffected by pH, polarity, or interfering ions. HSL-1 exhibited stronger resistance to background interference than HSL-2 and significantly enhanced fluorescence intensity; thus, it was selected for cell experiments and animal fluorescence intensity assessments. Furthermore, HSL-1 showed excellent biocompatibility, enabling real-time detection of mitochondrial viscosity changes and identification of viscosity abnormalities triggered by mitophagy in HeLa cells. It could also monitor changes in mitochondrial viscosity in zebrafish. In conclusion, HSL-1 is a valuable tool for studying viscosity and understanding diseases associated with abnormal mitochondrial viscosity.