Nanometer-scale molecular organization in lipid membranes studied by time-of-flight secondary ion mass spectrometry.

Jonas K Hannestad,Fredrik Höök,Peter Sjövall

doi:10.1116/1.5019794

Abstract

The organization of lipid membranes plays an important role in a wide range of biological processes at different length scales. Herein, the authors present a procedure based on time-of-flight secondary ion mass spectrometry (ToF-SIMS) to characterize the nanometer-scale ordering of lipids in lipid membrane structures on surfaces. While ToF-SIMS is a powerful tool for label-free analysis of lipid-containing samples, its limited spatial resolution prevents in-depth knowledge of how lipid properties affect the molecular assembly of the membrane. The authors overcome this limitation by measuring the formation of lipid dimers, originating in the same nanometer-sized primary ion impact areas. The lipid dimers reflect the local lipid environment and thus allow us to characterize the membrane miscibility on the nanometer level. Using this technique, the authors show that the chemical properties of the constituting lipids are critical for the structure and organization of the membrane on both the nanometer and micrometer length scales. Our results show that even at lipid surface compositions favoring two-phase systems, lipids are still extracted from solid, gel phase, domains into the surrounding fluid supported lipid bilayer surrounding the gel phase domains. The technique offers a means to obtain detailed knowledge of the chemical composition and organization of lipid membranes with potential application in systems where labeling is not possible, such as cell-derived supported lipid bilayers.

Highlights

We present a method to measure the lipid organization in supported lipid bilayers in regions as small as a few nanometers based on time-of-flight secondary ion mass spectrometry (ToF-SIMS)
The overall composition of the lipid membrane was primarily evaluated using the relative intensities of the different headgroup fragments, determined through the peak areas measured in high mass resolution mode
Images obtained in high spatial resolution mode show whether the different lipids are present at the surface either in a uniform lipid film with complete miscibility, or in two phase systems with discrete single-lipid domains

Summary

Introduction

We present a method to measure the lipid organization in supported lipid bilayers in regions as small as a few nanometers based on time-of-flight secondary ion mass spectrometry (ToF-SIMS). ToF-SIMS is a technique that is well suited for analysis of lipids, for which the detection and imaging of a wide range of molecular species with down to submicrometer spatial resolution have been demonstrated, with applications covering both synthetic samples and tissues.. ToF-SIMS provides a means to represent lipid surfaces in high detail, its spatial resolution is not sufficient to elucidate the finer detail of the lipid membrane composition. NanoSIMS has previously been employed to enable imaging of lipid films with nanoscale resolution.. NanoSIMS allows for a more finegrained understanding of lipid membrane organization, the technique has the downside of lacking the general applicability of ToF-SIMS, as it typically requires predefined isotopic labels in order to enable unambiguous identification of analytes. NanoSIMS has previously been employed to enable imaging of lipid films with nanoscale resolution. NanoSIMS allows for a more finegrained understanding of lipid membrane organization, the technique has the downside of lacking the general applicability of ToF-SIMS, as it typically requires predefined isotopic labels in order to enable unambiguous identification of analytes.

Methods

Results

Conclusion