Ion localisation in plant cells using the combined techniques of freeze-substitution and X-ray microanalysis

Abstract
X-ray microanalysis of plant cells Ions play vital roles in the water relations of plant and animal cells and in the regulation of cellular metabolism. An important component of the function of these elements is often a change in subcellular distribution. Investigation of the subcellular distribution of elements such as sodium, potassium, calcium, magnesium and chlorine, which are highly mobile in the aqueous phase, is a most challenging problem in biology. In order to study the distribution and transport of elements within cells a technique with high sensitivity and high resolution is required. The chosen method must not, in itself, alter the native elemental distribution during sample preparation or during measurement. Measurement of the (inorganic) elemental content of compartments (cell walls, cytoplasm and vacuole) within plant cells can be approached by a number of different methods, including the analysis of efflux of radioactive ions, analysis of tissues with different compartmental volume fractions, the use of microelectrodes specific to particular ions, by NMR in some instances and by X-ray microanalysis of individual cells. Of these, X-ray microanalysis offers more than any of the other methods in terms of both resolution and sensitivity. The use of microelectrodes is restricted in plant biology because of the physical strength of the walls of many cell types, and the difficulties of visualising the electrode tip in intact tissues in other than surface cells. As a technique, X-ray microanalysis also has the potential to resolve concentration differences within the cytoplasmic phase (e.g. between different organelles).