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Differential effects of hydrogen peroxide (H2O2) treatment on epitope recognition in western blotting.

Authors
  • Han, Shuting1
  • Cui, Yan2
  • Helbing, Dario-Lucas3
  • 1 Leibniz Institute on Aging, Fritz Lipmann Institute, 07745, Jena, Germany; Department of Medical Oncology, Sir Run Run Shaw Hospital, Medical School of Zhejiang University, Hangzhou, 310016, China. , (China)
  • 2 Leibniz Institute on Aging, Fritz Lipmann Institute, 07745, Jena, Germany. Electronic address: [email protected] , (Germany)
  • 3 Leibniz Institute on Aging, Fritz Lipmann Institute, 07745, Jena, Germany; Faculty of Medicine, Friedrich-Schiller-University Jena, 07743, Jena, Germany. , (Germany)
Type
Published Article
Journal
Analytical Biochemistry
Publisher
Elsevier
Publication Date
Dec 01, 2019
Volume
586
Pages
113417–113417
Identifiers
DOI: 10.1016/j.ab.2019.113417
PMID: 31505161
Source
Medline
Keywords
Language
English
License
Unknown

Abstract

Inactivation of horseradish peroxidase by hydrogen peroxide (H2O2) treatment is a convenient alternative to stripping for sequential chemiluminescent western blotting (WB). However, little evidence exists on whether H2O2 treatment affects epitope recognition. Here we show that H2O2 treatment had a negligible effect for most of the tested antibodies, whereas it could also eliminate or enhance antibody binding. Thus, H2O2 treatment has unpredictable effects on epitope recognition. Moreover, we demonstrate potential steric hindrance from previously bound antibodies. Hence, it would be advantageous to pre-test the suitability of antibodies for H2O2-treated WB, and to optimize conditions to mitigate steric interference with re-probing. Copyright © 2019 Elsevier Inc. All rights reserved.

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