Isolation of dictyosomes fromEuglena gracilis

Summary A method for the isolation of dictyosomes fromEuglena gracilis Klebs strain Z (Pringsheim) is described. An extensive Golgi system, with the individual dictyosomes commonly containing ten to twenty cisternae is present. Log phase cells are broken in a French pressure cell at 105 to 120 kg/cm₂ in a breaking mix containing sucrose, sorbitol and ficoll. Addition of 0.3% of glutaraldehyde or formaldehyde to the breaking mix increases the number of stacked cisternae present in the final preparation. In addition to membrane stacks, the fractions contain numerous smooth vesicles. Swollen cisternae, which are also present, may account for these vesicles. Three dictyosome-enriched fractions are obtained by centrifugation in a discontinuous sucrose gradient. Fractions differ morphologically in the degree of stacking of cisternae. Further identification of the membrane fractions was accomplished by measuring IDPase activities in each of the fractions. Inosine diphosphatase activity is enriched 8–10-fold relative to the initial homogenate. The highest IDPase activity was present in the fraction containing the greatest number of stacked cisternae.