How to detect ubiquitination

Detection of ubiquitination can be used to identify ubiquitination of specific amino acid residues or to confirm the action of specific ubiquitin ligases/conjugating enzymes. Ubiquitination detection strategy: First, ubiquitinated total proteins in the sample are enriched and purified through ubiquitination enrichment strategies. The obtained ubiquitinated proteins are then treated with deubiquitinating enzymes, followed by SDS-PAGE/immunoblotting with specific antibodies against the target protein. Finally, qualitative and semi-quantitative analysis of ubiquitinated proteins is achieved through band comparison analysis.

Detection of protein ubiquitination is mainly divided into in vivo and in vitro detection. The in vivo ubiquitination detection method involves co-immunoprecipitation (CO-IP), followed by SDS-PAGE/immunoblotting to observe the typical ladder pattern of ubiquitinated proteins. In vitro ubiquitination detection requires providing the target protein, ubiquitin, and related ligases and conjugating enzymes, followed by SDS-PAGE detection and direct detection of ubiquitination modifications of the target protein through protein immunoblotting experiments.

Biotytec uses Thermo Fisher's Q ExactiveHF mass spectrometry platform, Orbitrap Fusion mass spectrometry platform, and Orbitrap Fusion Lumos mass spectrometry platform combined with Nano-LC technology to providequantitative ubiquitination proteomics analysisservice technology package. You only need to send the samples and inform us of the experimental purpose. We will handle all subsequent matters of the project, including protein extraction, protein digestion, ubiquitinated peptide enrichment, peptide separation, mass spectrometry analysis, raw mass spectrometry data analysis, and bioinformatics analysis.

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