Selective fluorescent probes for live-cell monitoring of sulphide

Abstract

Aqueous sulphides, including hydrogen sulphide, have important roles in biological signalling and metabolic processes. Here we develop a selective sulphide-trapping strategy involving sulphide addition to an aldehyde; the resulting hemithioacetal undergoes a michael addition with an adjacent unsaturated acrylate ester to form a thioacetal at neutral pH in aqueous solution. Employing this new strategy, two sulphide-selective fluorescent probes, sFP-1 and sFP-2, were synthesized on the basis of two different fluorophore templates. These probes exhibit an excellent fluorescence increase and an emission maximum shift (sFP-1) in response to na 2 s and H 2 s in a high thiol background as found under physiological conditions. We show the utility of the probes for the selective detection of sulphides, and the capacity of our probes to monitor enzymatic H 2 s biogenesis and image free sulphide in living cells.

Highly sensitive sulphide mapping in live cells by kinetic spectral analysis of single Au-Ag core-shell nanoparticles

“…Although the indirectly measured sulphide concentrations may deviate more or less from the true values due to some systematic errors, the magnitudes as well as the trends of the calculated values are nevertheless reliable. Most previous reports regarding fluorescent sulphide probes were only able to provide static whole-cell images and reveal qualitative fluorescence intensity differences (among these images) after 15-30 min external stimulations with various sulphide concentrations in the mM range 24,26,28,29 To our knowledge, ours is not only the first demonstration of live cell sulphide imaging with nM sensitivity but is also the first time that local variations in intracellular sulphide levels have been tracked dynamically.…”

“…Although the PNP probes have performed as well as the fluorescent probes reported previously [23][24][25][26][27][28] , due to the ensemble average effect, bulk measurements are intrinsically not as sensitive as single-particle measurements. The AuNR-Ag PNP with a 2.1-nm thick silver shell, for example, contains B6 Â 10 5 silver atoms, which translates to B30 mM Ag for a B50 pM PNP solution.…”

“…During the past several years, several research groups have developed sulphide-responsive fluorescent organic molecules to detect H 2 S in live cells and have achieved selective intracellular sulphide imaging with limits of detection of 1-10 mM (ref. [23][24][25][26][27][28]. Although most publications suggest that the average endogenous H 2 S level is in the mM range, much lower sulphide concentrations have been reported 29 .…”

“…More importantly, the anabolism and catabolism of cellular sulphide are known to be rapid, which means that the sulphide concentration could fluctuate continuously from as high as submM during its explosive production to as low as a few nM after its rapid consumption 30 . However, almost all existing H 2 Ssensing methods are based on some irreversible stoichiometric reaction between the probe and sulphide species [23][24][25][26][27][28] and cannot be utilized to monitor the dynamics of sulphide generation and elimination inside cells, especially at the single-molecule level. Therefore, more sensitive methods that are capable of following time-dependent changes in H 2 S concentrations over a large dynamic range are needed for in-depth studies of this gaseous transmitter.…”

Expanding the chemistry of fluorescent protein biosensors through genetic incorporation of unnatural amino acids

“…81,[84][85][86][87][88] Recent reports described several small-molecule fluorescent probes for H 2 S detection in living systems. [89][90][91][92][93][94] All these designs are based on H 2 S-specific reactions. A reaction-based FP sensor for H 2 S was also constructed by Chen and co-workers.…”