The endoplasmic reticulum (ER) forms an extensive network in plant cells. In leaf cells and vacuolated root cells it is mainly restricted to the cortex whereas in the root meristem the cortical and cytoplasmic ER takes up a large volume throughout the entire cell. Only 3D electron microscopy provides sufficient resolution to understand the spatial organization of the ER in the root. However, high contrast staining and optimally ER specific staining is essential. Here we describe a protocol for selective ER staining that allows automated or semiautomated segmentation of the organelle in 3D datasets obtained from serial sections, Array Tomography, Serial Block Face Scanning Electron Microscopy (SBFSEM), or Focused Ion Beam (FIB) SEM.
Keywords: 3D EM; Automated segmentation; Endoplasmic reticulum; Serial Block Face SEM; ZIO.