Heterogeneous XXXX fibril growth mechanisms imaged by total internal reflection fluorescence microscopy.

@entity3793 @entity2532 @entity7732 @entity6
Total internal reflection fluorescence microscopy has been used to visualize the fibrillization of @entity2532 , a hormone which in aggregated forms plays a role in type 2 @entity6 pathology. Data were obtained at acidic pH where fibrillization is hindered by the charging of @entity3793 18 and at slightly basic pH where the loss of charge on the @entity3793 promotes aggregation. The experiments show three types of aggregate growth processes. In the earliest steps globular seeds are formed with some expanding radially during the course of the reaction. The dimensions of the globular seeds as well as their staining with the amyloid-specific dye @entity7732 indicate that they are plaques of short fibrils. The next species observed are fibrils that invariably grow from large globular seeds or smaller punctate granules. Fibril elongation appears to be unidirectional, although in some cases multiple fibrils radiate from a single seed or granule. After fibrils are formed, some show an increase in fluorescence intensity that we attribute to the growth of new fibrils alongside those previously formed. All three aggregation processes are suggestive of secondary (heterogeneous) nucleation mechanisms in which nucleation occurs on preformed fibrils. Consistently, electron micrographs show changes in fibril morphology well after fibrils are first formed, and the growth processes observed by fluorescence microscopy occur after the corresponding solution reactions have reached an initial apparent plateau. Taken together, the results highlight the importance of secondary nucleation in the fibrillization of @entity2532 , as this could provide a pathway to continue fibril growth once an initial population of fibrils is established.
