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Journal of Histochemistry and Cytochemistry, Vol. 48, 493-498, April 2000, Copyright © 2000, The Histochemical Society, Inc.

SYMPOSIUM

A Practical Technique to Postfix Nanogold-immunolabeled Specimens with Osmium and to Embed Them in Epon for Electron Microscopy

Hajime Sawadaa and Michiyo Esakia
a Department of Anatomy, Yokohama City University, School of Medicine, Yokohama, Japan

Correspondence to: Hajime Sawada, Dept. of Anatomy, Yokahama City University, School of Medicine, Fukuura 3-9, Kanazawa-ku, Yokohama, Japan 236-0004. E-mail: hsawada@med.yokohama-cu.ac.jp

Nanogold is a tiny gold probe, freely diffusible in cells and tissues, and is suitable for pre-embedding immunohistochemistry. However, it is necessary to develop Nanogold to a larger size so that it can be observed by conventional transmission electron microscopy. Silver enhancement is usually used for visualizing Nanogold, but the silver shell produced is unstable in OsO4 and often becomes invisible after OsO4 postfixation, which is necessary for good visualization of ultrastructure. We used silver enhancement with silver acetate, followed by gold toning with chloroauric acid, to replace the silver shell with a more stable gold in order to observe Nanogold after osmium fixation and Epon embedding. This technique is applicable to various intra- and extracellular antigens. For correlative observation of immunolabled specimens by light and electron microscopy, specimens adhered to slideglasses were embedded in Epon under non-adhesive plastic film. By heating the Epon sheets after polymerization, these supports were removed without difficulty and provided easy correlative observation. (J Histochem Cytochem 48:493–498, 2000)

Key Words: Nanogold, silver enhancement, gold toning, flat-embedding, osmium fixation


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