Electron tomography and immunogold labelling as tools to analyse de novo assembly of plant cell walls.

Abstract

High-resolution imaging of the membranous intermediates and cytoskeletal arrays involved in the assembly of a new cell wall during plant cytokinesis requires state-of-the-art electron microscopy techniques. The combination of cryofixation/freeze-substitution methods with electron tomography (ET) has revealed amazing structural details of this unique cellular process. This chapter deals with the main steps associated with these imaging techniques: selection of samples suitable for studying plant cytokinesis, sample preparation by high-pressure freezing/freeze substitution, and ET of plastic sections. In addition, immunogold approaches for the identification of proteins and polysaccharides during cell wall assembly are discussed.