Synaptic vesicles have been isolated from the electirc organ of the bony fish Electrophorus electricus using sucrose step gradients and zonal centrifugation. Although the acetylcholine (ACh) content of the Electrophorus electric organ is only 2% of that of Torpedo, ACh and ATP can readily be measured in the peak fractions using the leech microassay and the firefly luciferin luciferase assay respectively. The protein content of the vesicle fraction in experiments with Electrophorus was much higher than with Torpedo, but a possible contamination of this fraction with mitochondrial or cytoplasmic particles could be excluded. The ACh to ATP ratio of 10.8 is close to that found for cholinergic vesicles isolated from Torpedo and also to that of other amine storing granules.