Fluorescent probes alter miscibility phase boundaries in
ternary vesicles
Sarah L. Veatch, Sherry S. W. Leung , Robert E. W. Hancock, Jenifer L. Thewalt

We use 2H NMR to study the effects of probes on the miscibility transition in multilamellar vesicles
of di(18:1) phosphatidylcholine (PC; DOPC), chain perdeuterated di(16:0)PC (DPPCd62), and
cholesterol both with and without 0.5 mol% of the fluorescent probes DiIC12 or DiOC18. Both
probes raise the miscibility transition temperature in dispersions of 1:1 DOPC/DPPCd62 + 30%
cholesterol, but to differing extents. In membranes containing the popular probe DiIC12, we find that
the fraction of DPPCd62 lipids in the liquid disordered phase is increased, and the ordering of that
phase is reduced even at low temperatures. We examine membranes with smaller DiIC12 fractions and
find a significant shift in transition temperature in samples with 0.05 mol% DiIC12. We conclude that
trace impurities can have a significant effect on membrane physical properties and discuss possible
applications to biological membranes.