Molecule specific imaging of freeze-fractured, frozen-hydrated model membrane systems using mass spectrometry

Imaging time-of-flight secondary ion mass spectrometry is used to chemicall y resolve the spatial distribution of lipids in submicrometer sections of p hospholipid membranes. The results show that it is possible to unravel dyna mical events such as chemical fluctuations associated with domain structure in cellular membranes. In this work, a liposome model system has been used to capture the stages of membrane fusion between two merging bilayer syste ms. Fracturing criteria for preserving chemical distributions are shown to be much more stringent than morphological electron cryomicroscopy studies. Images of membrane heterogeneity, induced via mixing various liposomes foll owed by fast freezing, demonstrate the necessary sample preparation groundw ork to investigate complex, heterogeneous membrane domains. Clear delineati on of membrane structure provides direct evidence that specific domains or "rafts" can exist. Moreover, low concentrations of each phospholipid are di stributed throughout newly fused liposomes despite the existence of distinc t domains. In the liposome model, membrane structure ranges from specific d omains to a fluid mosaic of the phospholipids during the fusion event. The availability of mass spectrometric imaging is proposed to facilitate the di scovery of functional rafts or substructure in cell membranes before, durin g, and after events including cell division, exocytosis, endocytosis, intra cellular transport, infection of membrane-bound viruses, and receptor clust ering. This technology holds the promise to define the biology of cell memb ranes at the molecular level.