Imaging Plant Cells by High-Pressure Freezing and Serial block-face scanning electron microscopy.

Abstract

This chapter describes methods to enhanced contrast of plant material processed by high-pressure freezing and freeze substitution for improved visualization by serial block-face scanning electron microscopy (SBEM). The contrast enhancing steps are based on a protocol involving the sequential incubation of samples in heavy metals and sodium thiocarbohydrazide (OTO staining). We also describe the pipeline for imaging plant tissues in a commercial SBEM system (Gatan 3View ® ) and routines for the image analysis and three-dimensional reconstructions using open-source and commercial software packages.

Plant phenotyping relevance

植物組織の高圧凍結・SBEM撮像、画像解析、三次元再構築の一連の技術パイプラインが中心で、細胞・組織形態の取得に用いる方法論的研究である。

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