Efficient sample processing for proteomics applications--are we there yet?

Abstract

The ability to solubilize and digest protein extracts and recover peptides with high efficiency is of paramount importance in proteomics. A novel proteomic sample preparation protocol by Krijgsveld and colleagues (Hughes et al, 2014) provides significant advantages by enabling all sample processing steps to be carried out in a single tube to minimize sample losses, thereby enhancing sensitivity, throughput, and scalability of proteomics analyses.

Figure 1. Proteomics sample processing using surface-functionalized paramagnetic beads

(A) Proteins released from cell lysis are efficiently captured on carboxylate-coated paramagnetic beads by varying the proportion of organic solvent in the sample buffer. This facilitates downstream detergent removal for subsequent trypsin digestion. Following proteolytic digestion, peptides are re-adsorbed and desalted on paramagnetic beads under high organic content for further fractionation. (B) Functionalized paramagnetic beads efficiently capture variable amounts of HeLa cell extracts to expand the coverage of the proteome repertoire. Copy number is from a recent approach based on relative protein distribution (Wisniewski et al, 2014).