Automated protein digestion workflows for MS-based proteomics applications

Mass spectrometry (MS) based bottom-up proteomics is built upon large scale identification of peptides, and depends on proteins being efficiently converted to peptides by a protease of known specificity. The most common preparation methods are digestion in solution (ISD) or digestion of Proteins separated on an SDS PAGE gel, in-gel digestion (IGD). Both methods consist of a lengthy sequence of washing and chemical modification steps. To increase throughput and reproducibility, automation of these processes is highly desired. Contrary to other „omic“-applications, proteomics analysis by LC-MS/MS remains time-intensive, making the measurement the ratedetermining step in the pipeline. Thus the preparation of samples does not require a high-but rather a medium-throughput setup.

The normal benchtop methods for the IGD and the ISD were adapted to make automation in a robotic setup possible. In addition, we modified a standard PAL robot setup with a vacuum chamber that can be controlled by the robot's software and facilitates the removal of large volumes of washing solvents required by IGD leading to enhanced sensitivity.