Sonication-assisted protein extraction improves proteomic detection of membrane-bound and DNA-binding proteins from tumor tissues

Tissue lysis using urea buffer was adopted in our previously published CPTAC protocol for mass spectrometry (MS)-based quantitative global proteomic and phosphoproteomic analysis. Herein, we reported an update to this initial protocol by implementing a sonication step into urea-based tissue lysis. Similar to the initial CPTAC protocol, we identified >12,000 proteins and >25,000 phosphopeptides in a TMT set containing both non-sonicated and sonicated tumor tissues from patient-derived xenograft mouse models. However, an improvement in the detection of membrane-bound and DNA-binding proteins was observed by including the sonication. We also offered recommendations for optimal sonication conditions such as buffer composition, timing, and instrumentation settings, and a troubleshooting section for potential users. Additionally, the protocol is equally applicable to other biological specimens.