IMPORTANCE OF THE FIXATIVE FOR RELIABLE ULTRASTRUCTURAL PRESERVATION OF POIKILOHYDRIC PLANT-TISSUES - OBSERVATIONS ON DRY, PARTIALLY, AND FULLY HYDRATED TISSUES OF SELAGINELLA-LEPIDOPHYLLA

Leaves of desiccated 'resurrection plants', Selaginella lepidophylla, were hydrated either through the roots of intact plants or as isolated organs. Air-dry tissue and samples at 1, 4, 8 and 24 h (both detached and intact) of hydration were prepared for electron microscopy using aldehyde fixatives of different osmotic strengths. Both dry and hydrat ed tissues were also prepared using freeze substitution. Significant d ifferences in the ultrastructural preservation of these different samp les were noted. There was a direct correlation between the osmolality of both the fixative and the tissue with the quality of ultrastructura l preservation. When the osmolality of the fixative was slightly (or e ven considerably) higher than that of the tissue, optimal preservation was achieved. Freeze substitution, however, gave the most faithful pr eservation of all subcellular compartments, despite the frequent prese nce of small ice crystals. Additionally, hydration of detached leaves for more than 4 h resulted in swelling damage of the organelles and cy toplasm, regardless of the fixation protocol. Broadly interpreted, the results of this study indicate that an optimal preservation of plant cell and organelle ultrastructure can be achieved by the use of high o smolality fixatives or, preferably, freeze substitution. These results are also important in determining the method of hydration of poikiloh ydric samples for physiological studies and for interpretation of func tional changes as related to the structural condition of the organelle s. (C) 1997 Annals of Botany Company.