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Fluorescent characterization of amyloid deposits in the kidneys of mdx mice

Valeriia Gusel'nikova, Olga Antimonova, Elena Fedorova, Mikhail Shavlovsky, Aleksandr Krutikov, Ekaterina Mikhailova, Aleksandra Gudkova, Vyacheslav Mikhailov, Dmitry Korzhevskii
  • Olga Antimonova
    Institute of Experimental Medicine, Russian Federation
  • Mikhail Shavlovsky
    "Institute of Experimental Medicine, St.Petersburg" "the Pavlov First Saint Petersburg State Medical University" "Institute of cytology RAS" "Northwestern State Medical University I.I. Mechnikov", Russian Federation
  • Aleksandr Krutikov
    Institute of Molecular Biology and Genetics of the Almazov Federal Medical Research Centre, Russian Federation
  • Aleksandra Gudkova http://orcid.org/0000-0003-0156-8821
    "the Pavlov First Saint Petersburg State Medical University" "Institute of Molecular Biology and Genetics of the Almazov Federal Medical Research Centre", Russian Federation

Abstract

Amyloidosis is a group of diseases that occurs when amyloid proteins are deposited in tissues and organs. The traditional way of identifying amyloid in tissue sections is staining with Congo red. However, this method has a number of limitations including background staining (background fluorescence), low fluorescence intensity and false-positive staining. Therefore, a complex of fluorescence-based methods should be applied to characterize tissue localization of amyloid deposits. The aim of this study was to identify amyloid deposits in the kidneys of dystrophin-deficient mdx mice using different fluorescent dyes. We examined 8 cases of renal amyloidosis in aged mdx mice. In all cases, we used traditional methods for amyloid detection (Congo red and Thioflavin T), as well as a new fluorescent dye, disodium salt of 2,7- (1-amino-4-sulfo-2-naphthylazo) fluorene (DSNAF). In our study, we confirmed the amyloid structure of protein deposits in kidneys of aging mdx mice by several fluorescence-based staining methods. We found that fixation method has profound effects on downstream staining procedures, and demonstrated that the application of specific fixative, zinc-ethanol-formaldehyde (ZEF), instead of traditional NBF allow to reduce the background fluorescence. We also illustrated the usefulness of novel fluorescent dye DSNAF for detection of amyloid deposits in mouse tissues. Our results confirmed the strong affinity and high specificity of this dye for amyloid fibrils. The verification of DSNAF for detecting amyloid in human tissues will provide a conclusion on the applicability of the developed staining method in clinical research practice.

Keywords

amyloidosis; mdx mice; Congo red; Thioflavin T; fluorene

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Submitted: 2017-11-12 10:21:17
Published: 2018-04-03 09:01:09
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Copyright (c) 2018 Valeriia Gusel'nikova, Olga Antimonova, Elena Fedorova, Mikhail Shavlovsky, Aleksandr Krutikov, Ekaterina Mikhailova, Aleksandra Gudkova, Vyacheslav Mikhailov, Dmitry Korzhevskii

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