ClearSee: a rapid optical clearing reagent for whole-plant fluorescence imaging

• DOI: 10.1242/dev.127613
• Publication Date: 2015-10-23T01:38:17Z
• Authors: Daisuke Kurihara; Yoko Mizuta; Yoshikatsu Sato; Tetsuya Higashiyama
• Keywords: Chlorophyll; 0301 basic medicine; Photons; Microscopy, Confocal; Green Fluorescent Proteins; Arabidopsis; Phloem; Plants; Plants, Genetically Modified; Plant Roots; Fluorescence; Plant Leaves; 03 medical and health sciences; Techniques and Resources; Microscopy, Fluorescence; Urea; Indicators and Reagents; Cloning, Molecular; Xylitol; Fluorescent Dyes

Imaging techniques for visualizing and analyzing precise morphology and gene expression patterns are essential for understanding biological processes during development in all organisms. With the aid of chemical screening, we developed a clearing method using chemical solutions, termed ClearSee, for deep imaging of morphology and gene expression in plant tissues. ClearSee rapidly diminishes chlorophyll autofluorescence while maintaining fluorescent protein stability. By adjusting the refractive index mismatch, whole-organ and plant imaging can be performed both by confocal and two-photon excitation microscopy in ClearSee-treated samples. Moreover, ClearSee is applicable to multicolor imaging of fluorescent proteins to allow structural analysis of multiple gene expression. Given that ClearSee is compatible with staining by chemical dyes, the technique is useful for deep imaging in conjunction with genetic markers and for plant species not amenable to transgenic approaches. This method is useful for whole imaging for intact morphology and will help to accelerate the discovery of new phenomena in plant biological research.