A label-free, fast and high-specificity technique for plant cell wall imaging and composition analysis

Abstract

Background New cell wall imaging tools permit direct visualization of the molecular architecture of cell walls and provide detailed chemical information on wall polymers, which will aid efforts to use these polymers in multiple applications; however, detailed imaging and quantification of the native composition and architecture in the cell wall remains challenging. Results Here, we describe a label-free imaging technology, coherent Raman scattering (CRS) microscopy, including coherent anti-Stokes Raman scattering (CARS) microscopy and stimulated Raman scattering (SRS) microscopy, which can be used to visualize the major structures and chemical composition of plant cell walls. We outline the major steps of the procedure, including sample preparation, setting the mapping parameters, analysis of spectral data, and image generation. Applying this rapid approach will help researchers understand the highly heterogeneous structures and organization of plant cell walls. Conclusions This method can potentially be incorporated into label-free microanalyses of plant cell wall chemical composition based on the in situ vibrations of molecules.

Elastic and collapsible: current understanding of cell walls in succulent plants

“…Advancing cell wall analytical methods provide promising prospects, with a growing demand for high-throughput methods for rapid screening and profiling of cell wall components ( Persson et al , 2011 ). Spectroscopic methods have been widely used for cell wall characterization ( Bauer, 2012 ; Mansfield et al , 2012 ; Pettolino et al , 2012 ; Gierlinger, 2018 ; Zhao et al , 2020 ) in combination with imaging techniques ( Zhao et al , 2019 ; Bidhendi et al , 2020 ; DeVree et al , 2021 ; Xu et al , 2021 ). Recent advances in non-destructive real-time imaging, such as light-sheet fluorescence microscopy (LSFM), could allow us to observe changes in the cell walls of succulent tissues under drought in near-physiological conditions ( Grossmann et al , 2018 ; Ovečka et al , 2018 ).…”

Analyses of Microstructure and Dynamic Deposition of Cell Wall Components in Xylem Provide Insights into Differences between Two Black Poplar Cultivars

“…The obtained SRS images were processed for analysis using Fiji software, as described by Xu et al [15]. Briefly, images were opened and changed to 8 bit.…”

“…In order to compare the lignin deposition between the two cultivars, the mature xylems of both cultivars were observed under SRS microscopy and images were acquired at 1600 cm −1 . Subsequently, a quantitative and straightforward SRS image analysis was made, as described by Xu et al [15]. SRS imaging showed that lignin concentrations in the cell corner and compound middle lamella were higher than those in the secondary cell wall.…”

“…To document this variation, major efforts have been made to investigate the content and distribution of the biopolymer components within the xylem cell wall. Studies have shown that lignin displays higher concentrations in the corners of the cells and accumulates in the middle lamella, while cellulose is evenly distributed in primary cell walls and the middle lamella [13][14][15]. As for pectin components, the arabinan-rich pectin was found to be absent from the tangential walls of secondary xylem cells.…”

“…However, time-consuming and generally nonquantifiable analysis techniques make it difficult to follow the changes involved in xylem development at high spatiotemporal resolutions. With the development of cell wall imaging techniques, stimulated Raman scattering (SRS) microscopy was used to visualize the major structures and chemical composition of plant cell walls, providing a label-free, fast, and high-specificity method for plant cell wall imaging and composition analysis [13,15,22].…”