Improved clearing method contributes to deep imaging of plant organs

Tissue clearing methods are increasingly essential for the microscopic observation of internal tissues of thick biological organs. We previously developed TOMEI, a clearing method for plant tissues; however, it could not entirely remove chlorophylls nor reduce the fluorescent signal of fluorescent p...

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Bibliographic Details
Published in:Communications biology 2022-01, Vol.5 (1), p.12-12, Article 12
Main Authors: Sakamoto, Yuki, Ishimoto, Anna, Sakai, Yuuki, Sato, Moeko, Nishihama, Ryuichi, Abe, Konami, Sano, Yoshitake, Furuichi, Teiichi, Tsuji, Hiroyuki, Kohchi, Takayuki, Matsunaga, Sachihiro
Format: Article
Language:English
Subjects:
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14
14/19
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631/449/448
Animals
Arabidopsis
Biology
Biomedical and Life Sciences
Botany - instrumentation
Botany - methods
Brain - diagnostic imaging
Brain research
Diagnostic Imaging - instrumentation
Diagnostic Imaging - methods
Fluorescence
Green fluorescent protein
Hydrogels
Life Sciences
Methods
Mice
Neuroimaging
Proteins
Science
Seedlings
Sodium
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Summary:Tissue clearing methods are increasingly essential for the microscopic observation of internal tissues of thick biological organs. We previously developed TOMEI, a clearing method for plant tissues; however, it could not entirely remove chlorophylls nor reduce the fluorescent signal of fluorescent proteins. Here, we developed an improved TOMEI method (iTOMEI) to overcome these limitations. First, a caprylyl sulfobetaine was determined to efficiently remove chlorophylls from Arabidopsis thaliana seedlings without GFP quenching. Next, a weak alkaline solution restored GFP fluorescence, which was mainly lost during fixation, and an iohexol solution with a high refractive index increased sample transparency. These procedures were integrated to form iTOMEI. iTOMEI enables the detection of much brighter fluorescence than previous methods in tissues of A. thaliana , Oryza sativa , and Marchantia polymorpha . Moreover, a mouse brain was also efficiently cleared by the iTOMEI-Brain method within 48 h, and strong fluorescent signals were detected in the cleared brain. Sakamoto et al. demonstrate an improved optical clearing method, iTOMEI, for plant imaging. The new method can achieve fast clearing and effective removal of autofluorescence signals, and at the same time preserve signals from desired fluorescence proteins.
ISSN:2399-3642
2399-3642
DOI:10.1038/s42003-021-02955-9