Summary
Oxidative posttranslational protein modifications occur as a normal process of cell biology and to a greater extent during pathogenic conditions. The detection and quantitation of protein oxidation has posed a continuing challenge to bioanalytical chemists because the products of oxidative protein damage are chemically diverse, protein oxidation generally occurs at low background levels, and the complexity of biological samples introduces high background noise when standard techniques such as immunolabeling are applied to “dirty” tissue extracts. A refinement of classic reductive amination methods has been developed, which circumvents these difficulties by incorporating a biotin label at sites of protein carbonylation. Biotin hydrazide-labeled proteins are detectable using standard streptavidin-coupled detection techniques such as peroxidase-catalyzed chemiluminescence of immunoblots. Advantages of the biotin hydrazide-labeling technique are its sensitivity and its lack of reliance upon antibodies that inevitably suffer from nonspecific background noise and contaminating endogenous immunoglobulins.
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Acknowledgements
This work was supported in part by grants from the National Institutes of Health (NS044154), the ALS Association, and the Oklahoma Center for Advancement of Science and Technology (OCAST).
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© 2009 Humana Press, a part of Springer Science+Business Media, LLC
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Hensley, K. (2009). Detection of Protein Carbonyls by Means of Biotin Hydrazide-Streptavidin Affinity Methods. In: Kurien, B., Scofield, R. (eds) Protein Blotting and Detection. Methods in Molecular Biology, vol 536. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-59745-542-8_46
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DOI: https://doi.org/10.1007/978-1-59745-542-8_46
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