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Updated: Jan 27, 2026

Purification of Ubiquitinated p53 Proteins from Mammalian Cells
Published on: March 21, 2022
Detection of ubiquitinated targets in mammalian and Drosophila models
Elena Maspero1, Valentina Fajner1, Janine Weber1
1IFOM, Fondazione Istituto FIRC di Oncologia Molecolare, Milan, Italy.
Abstract:
In this chapter, we describe techniques to detect ubiquitination events occurring in vivo. We focus on methodologies capable of preserving and detecting target protein ubiquitination in physiological conditions, without overexpressing a tagged version of ubiquitin. We provide detailed protocols for mammalian and Drosophila melanogaster systems using linkage-specific antibodies against ubiquitin. We analyze immunoblotting and immunofluorescence approaches as well as an ELISA-based quantitative approach, which allows comparing more samples and conditions. Finally, we describe the use of in vitro ubiquitination assays with purified proteins that allows a direct analysis of the molecular mechanisms behind specific E3 activity.
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