Mapping Membrane Topology Label Free and Corrected for Changes in the Refractive Index of the Membrane on a Nanometer Scale

Abstract

The plasma membrane is the outer limit of the animal cell. As such, it is both a border separating inside from outside and a signaling platform for interactions with the surroundings. Among these interactions are extracellular matrix contacts and adhesion sites. The membrane and its contact sites together with the underlying cytoskeleton undergo constant remodeling, which leads to changes of the cell shape. In addition to spatial information micro-topographical maps provide, information about the z-dimension and describe the position of the plasma membrane with respect to the distance to a given substrate. Here we address how to measure height differences in the plasma membrane and how to create topographical maps of the plasma membrane with nanometer resolution. We address the currently used methodologies along with their advantages and drawbacks. Moreover, we delineate a label-free method to obtain topographic maps of the plasma membrane that are corrected for differences in the refractive index of the membrane utilizing an interferometric approach with multiple wavelengths and a normalization procedure to account for changes in the refractive index in the membrane.