Direct fluorescence imaging of cell walls in plant roots and stems

Microscopy methods developed in the 1960s are still commonly used in plant science today but new low-cost techniques offer big advantages.

Investigating plant structure has long been fundamental to botanical science. Across disciplines, including plant development, ecophysiology, and biotechnology, particular interest has focused on the complex structure of lignocellulosic walls. The lignified and suberized cell walls have vital physiological functions in structural support, defence, and water transport, and no less importantly, constitute a major part of the terrestrial biomass. Microscopy is critical to studies focussed on plant cell walls, with classical methods developed in the 1960s still popular to this day. However, recent innovations in tissue preparation, fluorescence staining and microscopy equipment offer advantages over traditional practices.

Hand-cut section of a Rhizophora apiculata root visualized after double staining with Congo red and fluorol yellow. Cell walls with suberin are shown in yellow, lignified walls are green, and the cortical parenchyma is red. Image credit: Kitin et al.

In their new paper published in AoBP, Kitin et al. review recent innovations in tissue preparation and fluorescence staining for lignin and suberin. They demonstrate simple and cost-effective protocols for preparation of plant samples for microscopy using hand-cut sections cleared with glycerol. Autofluorescence of lignin in combination with direct stains, such as Congo red and fluorol yellow, was shown to clearly differentiate between lignified, suberized, and unlignified cell walls in stem and root tissue samples. They also managed to use these methods for three-dimensional imaging of plant cells, using wide-field fluorescence or confocal laser scanning microscopy. The authors hope that their methods will help future plant anatomical and histochemical studies by providing data the variety of plant cell types and cell wall components.

William Salter

William (Tam) Salter is a Postdoctoral Research Fellow in the School of Life and Environmental Sciences and Sydney Institute of Agriculture at the University of Sydney. He has a bachelor degree in Ecological Science (Hons) from the University of Edinburgh and a PhD in plant ecophysiology from the University of Sydney. Tam is interested in the identification and elucidation of plant traits that could be useful for ecosystem resilience and future food security under global environmental change. He is also very interested in effective scientific communication.

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