Stoychev, S. Development of fully automated pipeline for phosphoproteome profiling
As clinical proteomics applications start to reach maturity, this necessitates the requirement for robust and routine high throughput sample preparation workflows that allow processing of large sample cohorts. However, efficient sample preparation remains the Achilles Heel for mass spectrometry analysis, with current methods lacking the throughput, transferability and reproducibility required to deal with these large clinical sample numbers in a routinized laboratory setting. To address these we focus on the implementation of versatile and automatable magnetic bead based sample preparation. Magnetic beads are considered desirable since these are easy to handle, linearly scalable, and high throughput compatible with the relatively simple integration of a magnetic stand in a variety of liquid handling stations making it fairly agnostic for the liquid handling or magnetic bead handling station that may be present in the sample preparation laboratory. We have previously demonstrated protein and peptide clean-up workflows using magnetic HILIC for Solid Phase Extraction (HILIC SPE) from a broad range of common contaminants, while the use of magnetic beads for highly efficient phosphopeptide enrichment has previously been demonstrated (Tape et al. 2014, Baath et al. 2018). In the current work we demonstrate the coupling of these steps into an automated workflow for phosphoproteome profiling requiring minimal liquid handling.