Abstract
Lignocellulosic biomass is a complex network of polymers making up the cell walls of plants. It represents a feedstock of sustainable resources to be converted into fuels, chemicals, and materials. Because of its complex architecture, lignocellulose is a recalcitrant material that requires some pretreatments and several types of catalysts to be transformed efficiently. Gaining more knowledge in the architecture of plant cell walls is therefore important to understand and optimize transformation processes. For the first time, super-resolution imaging of poplar wood samples has been performed using the Stimulated Emission Depletion (STED) technique. In comparison to standard confocal images, STED reveals new details in cell wall structure, allowing the identification of secondary walls and middle lamella with fine details, while keeping open the possibility to perform topochemistry by the use of relevant fluorescent nano-probes. In particular, the deconvolution of STED images increases the signal-to-noise ratio so that images become very well defined. The obtained results show that the STED super-resolution technique can be easily implemented by using cheap commercial fluorescent rhodamine-PEG nano-probes which outline the architecture of plant cell walls due to their interaction with lignin. Moreover, the sample preparation only requires easily-prepared plant sections of a few tens of micrometers, in addition to an easily-implemented post-treatment of images. Overall, the STED super-resolution technique in combination with a variety of nano-probes can provide a new vision of plant cell wall imaging by filling in the gap between classical photon microscopy and electron microscopy.
Highlights
The implementation of a super-resolution microscopy technique for imaging plant cell walls is a new topic with very few studies [1], whereas the need for accessing nanometer-resolution is well known
The advantages microscopy over conventional confocal microscopy to image gain in image resolution when used in combination with deconvolution treatment
The Stimulated Emission Depletion (STED) technique provides a gain in contrast and resolution cannot reach those of transmission electron microscopy [24], STED benefits image resolution when used in combination with deconvolution treatment
Summary
The implementation of a super-resolution microscopy technique for imaging plant cell walls is a new topic with very few studies [1], whereas the need for accessing nanometer (nm)-resolution is well known. The investigation of the nanoscale lignocellulosic architecture has already been conducted in order to correlate structural modifications with enzymatic digestibility [2] by comparing the mechanisms of different enzymes [3] through the combination of, for example, different microscopy and tomography analyses [4]. All of these studies used electron microscopy or atomic force microscopy techniques, which are complex techniques regarding sample preparation and/or image analysis. They are all based on the possibility of switching fluorescent dye molecules on and off into a dark state, whether by using a targeted switching approach (depletion wavelength), or a stochastic approach
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