Cyanine dyes with tail length asymmetry enhance photoselection: A multiscale study on DiD probes in a liquid disordered membrane.

Visualization of membrane domains like lipid rafts in natural or artificial membranes is a crucial task for cell biology. For this purpose, fluorescence microscopy is often used. Since fluorescing probes in lipid membranes partition specifically in e.g. local liquid disordered or liquid ordered environments, the consequent changes in their orientation and location are both theoretically and experimentally of interest. Here we focused on a liquid disordered membrane phase and performed molecular dynamics (MD) simulations of the indocarbocyanine DiD probes by varying the length of the attached alkyl tails and also the length of the cyanine backbone. From the probed compounds in a DOPC lipid bilayer at ambient temperature, a varying orientation of the transition dipole moment was observed, which is crucial for fluorescence microscopy and which, through photoselection, was found to be surprisingly more effective for asymmetric probes than for the symmetric ones. Furthermore, we observed that the orientation of the probes was dependent on the tail length; with the methyls or propyls attached, DiD oriented with its tails facing the water, contrary to the ones with longer tails. With advanced hybrid QM/MM calculations we show that the different local environment for differently oriented probes affected the one-photon absorption spectra, that was blue-shifted for the short-tailed DiD with respect to the DiDs with longer tails. We show here that the presented probes can be successfully used for fluorescence microscopy and we believe that the described properties bring further insight for the experimental use of these probes.