Aug, 2014
Electron microscopy of primary cell cultures in solution and correlative optical microscopy using ASEM
ULTRAMICROSCOPY
- Volume
- 143
- Number
- First page
- 52
- Last page
- 66
- Language
- English
- Publishing type
- Research paper (scientific journal)
- DOI
- 10.1016/J.ultramic.2013.10.010
- Publisher
- ELSEVIER SCIENCE BV
Correlative light-electron microscopy of cells in a natural environment of aqueous liquid facilitates highthroughput observation of protein complex formation. ASEM allows the inverted SEM to observe the wet sample from below, while an optical microscope observes it from above quasi-simultaneously. The disposable ASEM dish with a silicon nitride (SiN) film window can be coated variously to realize the primary-culture of substrate-sensitive cells in a few milliliters of culture medium in a stable incubator environment. Neuron differentiation, neural networking, proplatelet-formation and phagocytosis were captured by optical or fluorescence microscopy, and imaged at high resolution by gold-labeled immunoASEM with/without metal staining. Fas expression on the cell surface was visualized, correlated to the spatial distribution of F-actin. Axonal partitioning was studied using primary-culture neurons, and presynaptic induction by GluR82-N-terminus-linked fluorescent magnetic beads was correlated to the presynaptic-marker Bassoon. Further, megakaryocytes secreting proplatelets were captured, and P-selectins with adherence activity were localized to some of the granules present by immuno-ASEM. The phagocytosis of lactic acid bacteria by denthitic cells was also imaged. Based on these studies, ASEM correlative microscopy promises to allow the study of various mesoscopic-scale dynamics in the near future. (C) 2013 Elsevier B.V. All rights reserved.
- Link information
- ID information
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- DOI : 10.1016/J.ultramic.2013.10.010
- ISSN : 0304-3991
- eISSN : 1879-2723
- Web of Science ID : WOS:000336377100007