Detecting toxic species in the aggregation of amyloid beta in real time

Abstract Amyloid beta (A β ) is a peptide that accumulates in brain tissue and is highly prone to aggregation and misfolding. The amyloid hypothesis suggests that the primary cause of Alzheimer’s disease (AD) is due to the aggregation of A β . The aggregation of A β occurs through multiple stages, beginning with early-stage soluble intermediates and advancing to fully developed, late-stage insoluble amyloid fibrils. During this progression, the early-stage oligomers (soluble) are highly toxic, which play a crucial role in AD. It is challenging to capture the full timeline (a few µ s to several days) of A β aggregation using conventional biophysical and biochemical techniques due to its transient nature, inherent heterogeneity, inherent complexity, and wide dynamic range of the aggregation kinetics. In this study, we have utilized the closed-aperture Z-scan (CA Z-scan) method, which offers unique insights into the structural nonlinearity of intermediates, ranging from early-stage disordered forms to late-stage fibrils in real time. Furthermore, a remarkable shift from negative to positive sign in the nonlinear refractive index ( n 2 ) is observed, coinciding with the formation of toxic oligomers that play a direct role in the progression of AD. This finding could serve as a marker for diagnosing AD. The results obtained from the CA Z-scan are validated using Thioflavin T (ThT) assay and atomic force microscopy (AFM). Here, we have successfully demonstrated that the CA Z-scan, an affordable, simple, and highly sensitive technique, can effectively provide valuable insights into the kinetics of A β aggregation and the diagnosis of AD. We strongly believe that this tool may have promising future applications in developing new diagnostic and therapeutic strategies for various neurodegenerative diseases associated with different types of protein misfolding and aggregation.