Amyloids are thought to be involved in various types of neurodegenerative disorders. Several kinds of intermediates, differing in morphology, size, and toxicity, have been identified in the multistep amyloidogenesis process. However, the mechanisms explaining amyloid toxicity remain unclear. We previously generated a toxic mutant of the nontoxic HET-s((218-289)) amyloid in yeast. Here we report that toxic and nontoxic amyloids differ not only in their structures but also in their assembling process. We used multiple and complementary methods to investigate the intermediates formed by these two amyloids. With the methods used, no intermediates were observed for the nontoxic amyloid; however, under the same experimental conditions, the toxic mutant displayed visible oligomeric and fibrillar intermediates.