Fluorescence from dual molecular rotors allows sensitive detection of widespread brain pathology in a mouse model of Alzheimer's disease.

Amyloid fibrils formed by the misfolding and aggregation of proteins are a pathological hallmark of many neurodegenerative conditions including Alzheimer's disease (AD). Although recent studies have shown that pre-fibrillar species including low molecular-weight oligomers are more toxic in vitro than mature fibrils, and correlate better with cognitive decline in AD patients, techniques to study this subtle pathology remain limited. Here, we describe the use of the dye pair 9-(dicyanovinyl)julolidine (DCVJ) and crystal violet (CV), two fluorescent molecular rotors, to detect widespread pathology in the 5xFAD mouse model of AD via fluorescence spectroscopy. DCVJ and CV individually displayed a limited ability to detect spectral differences between WT and non-plaque areas of 5xFAD brain samples. However, when used in combination, the two probes discerned subtle but significant differences in a much higher proportion of tissue compared to either dye alone. These spectral differences were eliminated after treatment to disaggregate macromolecular protein assemblies, providing evidence that the dye pair was able to detect subtle pathology present in the parenchyma of the 5xFAD mouse brain. These findings demonstrate that the combined use of DCVJ and CV could be a valuable addition to the tools currently available to study the early stages of protein misfolding, which is essential for advancing therapeutics and diagnostic technologies in many neurodegenerative diseases.