Unveiling the Hydrogen Bonding Network of Intracellular Water by Fluorescence Lifetime Imaging Microscopy

Monitoring subtle changes in the local hydrogen bonding networks, which surround molecular crowding and guide cell functionality, is of paramount importance. Although the fluorescence based microscope technique could be useful, it faces a great challenge due to the nonavailability of a suitable probe, which not only can stain the whole cell but also can provide a contrast optical readout based on the extent of hydrogen bonding in different regions of the cell. Here, we demonstrate a unique amphiphilic fluorescent nanodot (FND) that has a sharp contrast in lifetime imaging due to the different extents of hydrogen bonding of intracellular water in different compartments of the cell. The amphiphilic nature guides the amount of aggregation in FND in different local environments producing contrast images in the lifetime and helps to directly monitor the extent of hydrogen bonding network.