We would like to measure intermolecular associations and proximity in membranes on the length scale of the molecules themselves (i.e. nm or less). FRET is commonly used; however, lipid molecules modified by dyes are very different from the lipids themselves, therefore a label-free method is desirable. We use secondary ion mass spectrometry to probe the nanometer scale structure of supported lipid bilayers by taking advantage of the intermolecular recombination of atomic ions into diatomic species that occurs in dynamic SIMS. In this experiment, one can measure mean distances between molecules on a length scale far below the lateral resolution of the NanoSIMS 50L instrument, which combines high spatial resolution and high mass resolution for chemical imaging. As an example, we show that in lipid bilayers, the efficiency of atomic recombination to form secondary 13C15N- ions depends on the distance between 13C and 15N atoms installed on head groups of different lipid molecules. Specifically, with site-specific isotopic labeling of lipid head groups, we determine the dependence of recombination efficiency on the average distance between 13C- and 15N-labeled phospholipids in supported monolayers and bilayers. We refer to this method of measuring nanometer-scale distances between isotopically-labeled molecules as a chemical ruler, similar in concept to FRET but different in physical phenomenon. High precision isotope ratio analysis with the NanoSIMS 50L makes this a potentially unique way to study proximity between membrane-associated components on the sub-5 nm length scale without the use of fluorescent dyes that perturb bilayer structure. We then apply this chemical ruler to study the structure of supported lipid bilayers of lipid compositions commonly believed to contain nanometer-scale liquid-ordered domains that often serve as models for lipid rafts.

Short range interactions in model membranes measured by atom recombination and mass spectrometry

251st National Meeting (2016)

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