Planing frozen hydrated plant specimens for SEM observation and EDX microanalysis.

Huang, C. X. and Canny, M. J. and Oates, K. and Mccully, M. E. (1994) Planing frozen hydrated plant specimens for SEM observation and EDX microanalysis. Microscopy Research and Technique, 28 (1). pp. 67-74. ISSN 1059-910X

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A procedure is described for forming a flat face on a frozen piece of plant tissue, which may then be observed fully-hydrated or lightly etched, and coated or uncoated with a metal film, in scanning electron microscopy (SEM). The frozen sample was planed with a glass knife at -80°C in cryo-ultramicrotome. The sections were discarded, and the planed block face placed on the cold stage in the microscope column, either for observation uncoated at low kV, or for light etching (-90°C) to reveal the cell outlines. If a higher accelerating voltage was needed, the face was given an evaporative coating of Al in the cryo-preparation chamber and returned to the column. The advantages of the planed face over the usual fracture face are illustrated: imaging at a chosen rather than a chance position; clearer cellular and subcellular detail; preservation of hydrated gels like mucilage and swollen cell walls; the possibility of making serial parallel sections through the same piece of tissue; opportunities for accurate morphometric analyses on the planed face; capacity to produce longitudinal sections; preservation of very delicate structures that are destroyed by fixation and drying. A major advantage of the Al-coated planed face is the increased accuracy of energy-dispersive X-ray (EDX) microanalyses on a smooth rather than a rough surface. Tests are included which show that neither the light etching employed, nor successive planing, interferes with the analyses of elements in the frozen face.

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Journal Article
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Microscopy Research and Technique
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30 Jan 2009 09:31
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21 Nov 2022 19:02