Chemical Imaging of the Lipid and Cholesterol Distribution in the Plasma Membranes of Intact Cells

Abstract

Domains enriched with cholesterol and sphingolipids, often referred to as lipid rafts, are believed to be present in the plasma membranes of eukaryotic cells and mediate important cellular functions, including cell signaling and virus budding. Changes in the abundances of cholesterol and sphingolipids influence many cellular processes. Yet, the distributions of cholesterol and sphingolipids within the plasma membrane have not been established. The inability to directly visualize lipids and cholesterol without the use of labels that may induce clustering renders characterizing the precise distribution of cholesterol and sphingolipids within the plasma membrane especially challenging. To address this problem, we have previously combined a chemically specific and spatially well-resolved imaging technique, high-resolution secondary ion mass spectrometry (SIMS), with metabolic labeling in order to image the distributions of sphingolipids within the plasma membranes of intact cells with at least 100 nm lateral resolution. We now report the use of this approach to image the sphingolipid and cholesterol distributions in parallel on the surface of intact fibroblast cells. We also quantitatively assess whether the cholesterol and sphingolipids are heterogeneously distributed and co-localized within the membrane. The results of these studies and their implications on models of membrane organization will be discussed.