Evaluation of a high-intensity focused ultrasound-immobilized trypsin digestion and ¹⁸O-labeling method for quantitative proteomics

A new method that uses immobilized trypsin concomitant with ultrasonic irradiation results in ultrarapid digestion and more thorough ¹⁸O labeling for quantitative protein comparisons. The method was reproducible and provided effective digestions within <1 min with lower amounts of enzyme, compared to traditional methods. This method was demonstrated for digestion of both simple and complex protein mixtures, including bovine serum albumin, a global proteome extract from the bacteria Shewanella oneidensis, and mouse plasma, as well as ¹⁸O labeling of complex protein mixtures, validating this method for differential proteomic measurements. This approach is simple, reproducible, cost-effective, rapid, and well suited for automation.