Resin-Embedded Multicycle Imaging of Cells and Isolated Plasma Membranes

Abstract

Studies of the plasma membrane protein organization have undergone numerous advances; however, the large number of protein species involved complicates their examination. One approach which has the ability to resolve this complication is that of resin-embedded multicycle imaging (REMI), which stabilizes protein structure and antigenicity while labels are removed and replaced, allowing an arbitrarily large number of proteins of interest to be imaged in the same area. The implementation spearheading this technique is array tomography, which is ideal for many applications, but certain organelles or spatially restricted volumes of interest, such as the plasma membrane, present considerable difficulty. We have developed a method by which cells or isolated plasma membranes unroofed by osmotic shock can be embedded in thin films of acrylic resin (LR White, Lowicryl), making their spatially dependent proteomic information amenable to repeated immunostaining and other methods of analysis. Here we present our latest work in the development of this method.