Journal
JOURNAL OF ELECTRON MICROSCOPY
Volume 61, Issue 5, Pages 321-326Publisher
OXFORD UNIV PRESS
DOI: 10.1093/jmicro/dfs055
Keywords
atomic force; cytoskeleton; actin; freeze-etching; unroofing; electron microscopy
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Funding
- Japan Society for the Promotion of Science [22370056]
- Ministry of Knowledge Economy in Korea [ISTDP 10033633]
- Grants-in-Aid for Scientific Research [22370056] Funding Source: KAKEN
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Atomic force microscopy (AFM) combined with unroofing techniques enabled clear imaging of the intracellular cytoskeleton and the cytoplasmic surface of the cell membrane under aqueous condition. Many actin filaments were found to form a complex meshwork on the cytoplasmic surface of the membrane, as observed in freezeetching electron microscopy. Characteristic periodic striations of about 5 nm formed by the assembly of Gactin were detected along actin filaments at higher magnification. Actin filaments aggregated and dispersed at several points, thereby dividing the cytoplasmic surface of the membrane into several large domains. Microtubules were also easily detected and were often tethered to the membrane surface by fine filaments. Furthermore, clathrin coats on the membrane were clearly visualized for the first time in water by AFM. Although the resolution of these images is lower than electron micrographs of freezeetched samples processed similarly, the measurement capabilities of the AFM in a more biologically relevant conditions demonstrate that it is an important tool for imaging intracellular structures and cell surfaces in the native, aqueous state.
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