Protocol for live imaging of intracellular nanoscale structures using atomic force microscopy with nanoneedle probes

Takehiko Ichikawa; Mohammad Shahidul Alam; Marcos Penedo; Kyosuke Matsumoto; Sou Fujita; Keisuke Miyazawa; Hirotoshi Furusho; Kazuki Miyata; Chikashi Nakamura; Takeshi Fukuma

doi:10.1016/j.xpro.2023.102468

Protocol for live imaging of intracellular nanoscale structures using atomic force microscopy with nanoneedle probes

STAR Protoc. 2023 Sep 15;4(3):102468. doi: 10.1016/j.xpro.2023.102468. Epub 2023 Jul 22.

Authors

Affiliations

¹ Nano Life Science Institute (WPI-NanoLSI), Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan. Electronic address: tichikawa@staff.kanazawa-u.ac.jp.
² Division of Nano Life Science, Graduate School of Frontier Science Initiative, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan.
³ École Polytechnique Fédérale de Lausanne, Institute for Bioengineering, Laboratory for Bio and Nanoinstrumentation, Lausanne, CH 1015, Switzerland.
⁴ Faculty of Frontier Engineering, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan.
⁵ Nano Life Science Institute (WPI-NanoLSI), Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan; Faculty of Frontier Engineering, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan.
⁶ Nano Life Science Institute (WPI-NanoLSI), Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan.
⁷ Nano Life Science Institute (WPI-NanoLSI), Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan; Division of Nano Life Science, Graduate School of Frontier Science Initiative, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan; Faculty of Frontier Engineering, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan.
⁸ AIST-INDIA Diverse Assets and Applications International Laboratory (DAILAB), Cellular and Molecular Biotechnology Research Institute (CMB), National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8565, Japan.
⁹ Nano Life Science Institute (WPI-NanoLSI), Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan; Division of Nano Life Science, Graduate School of Frontier Science Initiative, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan; Faculty of Frontier Engineering, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan. Electronic address: fukuma@staff.kanazawa-u.ac.jp.

Abstract

Atomic force microscopy (AFM) is capable of nanoscale imaging but has so far only been used on cell surfaces when applied to a living cell. Here, we describe a step-by-step protocol for nanoendoscopy-AFM, which enables the imaging of nanoscale structures inside living cells. The protocol consists of cell staining, fabrication of the nanoneedle probes, observation inside living cells using 2D and 3D nanoendoscopy-AFM, and visualization of the 3D data. For complete details on the use and execution of this protocol, please refer to Penedo et al. (2021)¹ and Penedo et al. (2021).².

Keywords: AFM; Atomic Force Microscopy; Cell Biology; Microscopy.

MeSH terms

Cell Membrane / chemistry
Microscopy, Atomic Force / methods
Nanotechnology* / methods