Cx. Huang et al., PLANING FROZEN-HYDRATED PLANT SPECIMENS FOR SEM OBSERVATION AND EDX MICROANALYSIS, Microscopy research and technique, 28(1), 1994, pp. 67-74
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 etc
hed, and coated or uncoated with a metal film, in scanning electron mi
croscopy (SEM). The frozen sample was planed with a glass knife at -80
degrees C in a cryo-ultramicrotome. The sections were discarded, and
the planed block face placed on the cold stage in the microscope colum
n, either for observation uncoated at low kV, or for light etching (-9
0 degrees C) to reveal the cell outlines. If a higher accelerating vol
tage was needed, the face was given an evaporative coating of Al in th
e cryo-preparation chamber and returned to the column. The advantages
of the planed face over the usual fracture face are illustrated: imagi
ng at a chosen rather than a chance position; clearer cellular and sub
cellular detail; preservation of hydrated gels like mucilage and swoll
en cell walls; the possibility of making serial parallel sections thro
ugh 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 fixat
ion 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 tha
t neither the light etching employed, nor successive planing, interfer
es with the analyses of elements in the frozen face. (C) 1994 Wiley-Li
ss, Inc.