論文

査読有り 国際誌
2020年9月15日

Practical method for superresolution imaging of primary cilia and centrioles by expansion microscopy using an amplibody for fluorescence signal amplification.

Molecular biology of the cell
  • Yohei Katoh
  • ,
  • Shuhei Chiba
  • ,
  • Kazuhisa Nakayama

31
20
開始ページ
2195
終了ページ
2206
記述言語
英語
掲載種別
研究論文(学術雑誌)
DOI
10.1091/mbc.E20-04-0250

Primary cilia are microtubule-based protrusions from the cell surface that are approximately 0.3 µm in diameter and 3 µm in length. Because size approximates the optical diffraction limit, ciliary structures at the subdiffraction level can be observed only by using a superresolution microscope or electron microscope. Expansion microscopy (ExM) is an alternative superresolution imaging technique that uses a swellable hydrogel that enables the physical expansion of specimens. However, the efficacy of ExM has not been fully verified, and further improvements in the method are anticipated. In this study, we applied ExM to the observation of primary cilia and centrioles and compared the acquired images with those obtained using conventional superresolution microscopy. Furthermore, we developed a new tool, called the amplibody, for fluorescence signal amplification, to compensate for the substantial decrease in fluorescence signal per unit volume inherent to physical expansion and for the partial proteolytic digestion of cellular proteins before expansion. We also demonstrate that the combinatorial use of the ExM protocol optimized for amplibodies and Airyscan superresolution microscopy enables the practical observation of cilia and centrioles with high brightness and resolution.

リンク情報
DOI
https://doi.org/10.1091/mbc.E20-04-0250
PubMed
https://www.ncbi.nlm.nih.gov/pubmed/32726175
PubMed Central
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7550703
ID情報
  • DOI : 10.1091/mbc.E20-04-0250
  • PubMed ID : 32726175
  • PubMed Central 記事ID : PMC7550703

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