Abstract
Staining and immunodetection by light microscopy are methods widely used to investigate plant cell walls. The two techniques have been crucial to study the cell wall architecture in planta, its deconstruction by chemicals or cell wall-degrading enzymes. They have been instrumental in detecting the presence of cell types, in deciphering plant cell wall evolution and in characterizing plant mutants and transformants. The success of immunolabeling relies on how plant materials are embedded and sectioned. Agarose coating, wax and resin embedding are, respectively, associated with vibratome, microtome and ultramicrotome sectioning. Here, we have systematically carried out a comparative analysis of these three methods of sample preparation when they are applied for cell wall staining and cell wall immunomicroscopy. In order to help the plant community in understanding and selecting adequate methods of embedding and sectioning for cell wall immunodetection, we review in this article the advantages and limitations of these three methods. Moreover, we offer detailed protocols of embedding for studying plant materials through microscopy.
Highlights
Plant cell walls are fundamental to plant biology and have a strong impact on the use of plant products in industrial processes
We demonstrate how they impact on primary fluorescence detection and how they can affect the accessibility of antibodies and cell wall-degrading enzymes to targeted polymers
Immunolabeling of plant materials requires the use of highquality sections and obtaining that is often not trivial
Summary
Plant cell walls are fundamental to plant biology and have a strong impact on the use of plant products in industrial processes. Immunolabeling procedures have been crucial to study the cell wall architecture in planta (e.g., Knox et al, 1990; Majewska-Sawka et al, 2004; Guillemin et al, 2005; Xue et al, 2013) and its deconstruction by chemicals or cell wall-degrading enzymes (Marcus et al, 2008; Hervé et al, 2010) Both staining and immunodetection of plant polysaccharides enable to focus on specific cell types (Hall et al, 2013; Eeckhout et al, 2014) and immunolabelings even allow to isolate certain cell types from their neighbor cells (Verhertbruggen et al, 2009b). Cell wall immunolabeling has contributed in evaluating plant cell wall evolution and in redefining plant
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