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Sci Rep. 2019 Oct 23;9(1):15182. doi: 10.1038/s41598-019-51624-9.

A versatile microfluidic device for highly inclined thin illumination microscopy in the moss Physcomitrella patens.

Author information

1
Division of Biological Science, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, 464-8602, Japan. kozgunova@gmail.com.
2
Division of Biological Science, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, 464-8602, Japan.

Abstract

High-resolution microscopy is a valuable tool for studying cellular processes, such as signalling, membrane trafficking, or cytoskeleton remodelling. Several techniques of inclined illumination microscopy allow imaging at a near single molecular level; however, the application of these methods to plant cells is limited, owing to thick cell walls as well as the necessity to excise a part of the tissue for sample preparation. In this study, we utilised a simple, easy-to-use microfluidic device for highly inclined and laminated optical sheet (HILO) microscopy using a model plant Physcomitrella patens. We demonstrated that the shallow microfluidic device can be used for long-term culture of living cells and enables high-resolution HILO imaging of microtubules without perturbing their dynamics. In addition, our microdevice allows the supply and robust washout of compounds during HILO microscopy imaging, for example, to perform a microtubule regrowth assay. Furthermore, we tested long-term (48 h) HILO imaging using a microdevice and visualised the developmental changes in the microtubule dynamics during tissue regeneration. These novel applications of the microfluidic device provide a valuable resource for studying molecular dynamics in living plant cells.

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