Congo Red Derived Covalent Proteotyping Sensor for Amyloid Deposits in Alzheimer’s Disease

Congo Red (CR) is the histochemical staining sensor used to diagnose amyloid tissue deposition in current clinical practice. Its characteristic aryl azo linkage is generally considered to be chemically stable. Here, we discovered by serendipity that neutral borate buffer can activate the inert azo bond in CR to covalently modify amyloid proteins at ambient temperature. Such chemistry allowed us to develop a covalent amyloid sensor to image, enrich, and proteotype amyloid deposits in Alzheimer’s disease (AD) tissue. We first pinpointed the boronic acid in borate buffer triggers such amyloid bioconjugation and found that ultraviolet-light-induced azo trans-to-cis isomerization further enhanced labeling efficiency. Leveraging this boron-azo bioconjugation chemistry, we developed a covalent sensor based on CR for selective detection and microdissection of amyloid deposits from AD mouse brain tissues. By incorporating an alkyne handle into the CR scaffold, we enabled click-chemistry-assisted enrichment of amyloid aggregates, followed by proteotyping via LC-MS/MS. The covalent amyloid sensor proteotyped a complex network of protein interactors including the AD biomarkers Tau and ApoE within amyloid deposits. Overall, this work establishes a borate-activated azo-based covalent sensor for selective imaging, dissecting and proteotyping of amyloid deposits in AD tissues.