pH-Independent two-photon fluorescent lysotrackers for real-time monitoring autophagy

Two carbazole-based two-photon fluorescent lysotrackers with different electron-donating groups (a methoxyl group for Lyso-MCO and a dimethylamino group for Lyso-NCO, respectively) have been developed from simple starting materials via an only 2-step procedure. Both of them exhibit pH-independent and specific lysosome location with a rapid staining rate, high photostability and deep issue penetration along with large two-photon absorption action cross-sections. By virtue of the better two-photon absorption properties of Lyso-NCO, it was chosen to visually monitor lysosomal tracking and autophagy. Compared with the approach of GFP-LC3 for autophagy detection, lysotracker Lyso-NCO achieved efficient and real-time visualization of the membrane fusion period in the autophagy process through detecting the level of the co-localization coefficients between Lyso-NCO and Mitotracker Red FM (MTR) in live cells.