Modification of methods to use Congo-red stain to simultaneously visualize amyloid plaques and tangles in human and rode
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ORIGINAL ARTICLE
Modification of methods to use Congo-red stain to simultaneously visualize amyloid plaques and tangles in human and rodent brain tissue sections Sumit Sarkar 1
&
James Raymick 1 & Elvis Cuevas 1 & Hector Rosas-Hernandez 1 & Joseph Hanig 2
Received: 24 June 2020 / Accepted: 11 August 2020 # This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply 2020
Abstract Although there are multiple histochemical tracers available to label plaques and tangles in the brain to evaluate neuropathology in Alzheimer disease (AD), few of them are versatile in nature and compatible with immunohistochemical procedures. Congo Red (CR) is an anisotropic organic stain discovered to label amyloid beta (Aβ) plaques in the brain. Unfortunately, its use is underappreciated due to its low resolution and brightness as stated in previous studies using bright field microscopy. Here, we modified a previous method to localize both plaques and tangles in brains from humans and a transgenic rodent model of AD for fluorescence microscopic visualization. The plaque staining affinities displayed by CR were compared with fibrillar pattern labeling seen with Thioflavin S. This study summarizes the optimization of protocols in which various parameters have been finetuned. To determine the target CR potentially binds, we have performed double labeling with different antibodies against Aβ as well as phosphorylated Tau. The plaque staining affinities exhibited by CR are compared with those associated with the diffuse pattern of labeling seen with antibodies directed against different epitopes of Aβ. Neither CP13, TNT2 or TOC1 binds all the neurofibrillary tangles as revealed by CR labeling in the human brain. Additionally, we also evaluated double labeling with AT8, AT180, and PHF1. Interestingly, PHF-1 shows 40% colocalization and AT8 shows 15% colocalization with NFT. Thus, CR is a much better marker to detect AD pathologies in human and rodent brains with higher fluorescence intensity relative to other conventional fluorescence markers. Keywords Congo red . Alzheimer disease . Human brain . Amyloid plaques . Neurofibrillary tangle . Transgenic rat
Introduction Alzheimer disease (AD) is one of the most debilitating diseases of the elderly, presently affecting more than 35.2 million people worldwide; the number of affected individuals is expected to rise to 65.7 million by 2030 (1). Two hallmark features of AD are deposits of senile plaques (SP), which consist primarily of amyloid β-protein (Aβ) peptides and neurofibrillary tangles (NFTs), also commonly referred to as filamentous tau lesions, as well as eventual loss of neurons in the
* Sumit Sarkar [email protected] 1
Division of Neurotoxicology, National Center for Toxicological Research/US FDA, HFT-132, Jefferson, AR 72079, USA
2
Office of Testing & Research, Center for Drug Evaluation Research/ FDA, Silver Spring, MD, USA
brain. It is imperative to note that most of the methods that have been used to detect AD
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