Fixation of Phytomonad cysts for electron microscopy

This thesis has been concerned with a chemical fixation method appropriate for Polytomella agilis cyst forms. P.agilis is a flagel­ late algae which during encystment builds a cyst coat of high tensile strength(3 9). P.agilis cultures were grown in the dark,at 25°C,in a complex medium, 0.1% (w/v) tryptone, 0.2% (w/v) yeast extract, 0.2% (w/v)sodium acetate(37). Samples of cells were collected for examination under the electron microscope at various stages in the growth cycle, from one day old, twenty one day old, four day old, ten-day old cultures. The chemicals used as single and double fixatives were Afzelius Os04 in CCl4 non-aqueous solution(la), Luft's aqueous unbuffered KMn04 solution (24) and aqueous unbuffered glutaraldehyde solution. Osmium tetroxide has also been used in sodium phosphate buffered aqueous solution for standard double fixation procedures, as applied by Sjostrand (40a). Evaluation of the results obtained has been based on the ability of the fixative to stabilize the ultrastructural features regularly observed. For the first time in the cyst forms of this organism without distorting its cellular morphology, the fixation of the non - homogeneous multilayered cyst wall, the preservation of starch and lipid forms of food reserves, and the honey- comb microbodies has been accomplished.