Time-gated luminescence probe for ratiometric and luminescence lifetime detection of Hypochorous acid in lysosomes of live cells

Time-gated luminescence (TGL) probes based on lanthanide complexes have appealed wide attention in the detection of biologically relevant analytes because of their inimitable photophysical properties. In this work, a TGL probe (TR-HOCl) based on intramolecular Forster resonance energy transfer (FRET) system for specific determination of hypochlorous acid (HOCl) was designed and synthesized, in which a rhodamine derivative (energy acceptor) was conjugated to a luminescent Tb3+ complex (energy donor). After reacting with HOCl, the Tb3+ emission of TR-HOCl at 540 nm declined while the rhodamine emission at 580 nm increased, which leaded to an increase of the TGL intensity ratio of rhodamine to Tb3+ complex (I-560/I-540) up to similar to 9-fold. The dose-dependent increase of I-560/I-540 gives a nice linearity in HOCl concentration range of 0.5-45 mu M. The detection limit of for HOCl was determined to be 0.34 mu M. Interestingly, the average luminescence lifetime of the Tb3+ emission decreased (from 588 mu s to 254 mu s) accompanied with the FRET process and the value gave a fine linearity to the variation of HOCl concentration. Additionally, TR-HOCl showed great selectivity for recognition of HOCl over other ROS, RNS, biothiols and other interference. These properties endow TR-HOCl to be conveniently applied for high accurate recognition of HOCl with ratiometric TGL and luminescence lifetime dual signal output. Finally, TR-HOCl was successfully applied for the TGL determination of HOCl in HepG2 cells. The co-localization experiments of TR-HOCl with LysoSensor Green revealed the lysosome-localizing property of the probe in live cells. The study demonstrated that TR-HOCl could be a competent tool for investigating roles of HOCl in various physiological processes.