Lipid domains as obstacles for lateral diffusion in supported bilayers probed at different time and length scales by two-dimensional exchange and field gradient solid state NMR

Employing two new solid-state NMR techniques for measuring lateral diffusion of lipids in supported bilayers, the deuterium two-dimensional exchange deuterium NMR (2D-NMR) and the Supercon fringe field proton NMR, we have studied bilayer domain connectivity in the coexistence region of a 1:1 DMPC-DSPC mixture. The advantage of the NMR methods is that they permit the study of percolation in lipid mixtures as a function of the experimental time and length scale and that they do not rely on the use of any labels attached to the lipids that change their molecular packing properties. Both NMR methods show an excellent agreement of their results at comparable mixing times. Moreover, a simulation of the effect of restricted diffusion on the 2D-NMR spectra shows clearly that the sensitivity of 2D-NMR for gel domains as diffusion obstacles in terms of a simple domain model depends on the mixing time and thus on the length scale of the experiment. In contrast to previous findings made by fluorescence and spin label studies, we do not observe any discontinuity of the lateral diffusion coefficient D indicative of a disconnection of the fluid phase down to a temperature which is only 4 ^∘{}C above the solidus temperature of the mixture. At experimental length scales below 200 nm, D is not significantly reduced due to the presence of gel phase domains up to an area fraction of 0.5. At length scales between 500 nm and 1 {μ}m both methods give a rather linear decrease of D with increasing gel phase area fraction, starting in the vicinity of the liquidus point. This surprising result is discussed in terms of the relationship between experimental length scale and detectability of percolation.